1: Am J Physiol Cell Physiol. 2005 Nov 16; [Epub ahead of print] Estradiol-17{beta} stimulates proliferation of mouse embryonic stem cells (ES-E14TG2a): Involvement of MAPKs and CDKs as well as protooncogenes. Han HJ, Heo JS, Lee YJ. Department of Veterinary Physiology, College of Veterinary Medicine,Chonnam National University, Gwangju, Gwangju, Korea, Republic of. Although the importance of estradiol-17beta(E2) in many physiological processes has been reported, no previous study has investigated the effects of E2 on embryonic stem (ES) cell proliferation. In this study, therefore, we examined the effect of E2 on the DNA synthesis of mouse ES cells and its related signaling pathways. The results of this study showed E2 (10(-9) M) significantly increased [(3)H] thymidine incorporation at > 4 hr and E2 (> 10(-12) M) induced an increase of [(3)H] thymidine incorporation following 8 hr incubation. Moreover, E2 (> 10(-12) M) also increased BrdU incorporation and cell number. Indeed, E2 stimulated estrogen receptors (ERalphaand ERbeta) protein levels and increased mRNA expression levels of protooncogenes (c-fos, c-jun, and c-myc). Tamoxifen (antiestrogen) completely inhibited E2-induced increases of [(3)H] thymidine incorporation. In addition, E2-BSA (10(-9) M) also increased [(3)H] thymidine incorporation at > 1 hr and E2-BSA (> 10(-12) M) increased [(3)H] thymidine incorporation at 1 hr incubation. E2-BSA-induced increase of BrdU incorporation also appeared in a dose-dependent manner. Tamoxifen had no effect on E2-BSA-induced increase of [(3)H] thymidine incorporation. Also, E2 and E2-BSA displayed maximal phosphorylation of p44/42 MAPKs at 10 min or 5 min, respectively. E2 increased cyclin D1, cyclin E, cyclin-dependent kinase (CDK) 2 and CDK 4. In contrast, E2 decreased the levels of p21(cip1) and p27(kip1) (CDKs inhibitory proteins). Increases of these cell cycle regulators were blocked by PD 98059 (MEK inhibitor, 10(-5) M). Moreover, E2-induced increase of [(3)H] thymidine incorporation was inhibited by PD 98059 or butyrolactone I (CDK 2 inhibitor). In conclusion, estradiol-17beta stimulates DNA synthesis of mouse ES cells, and this action is mediated by MAPKs, CDKs, or protooncogenes. PMID: 16291822 [PubMed - as supplied by publisher] --------------------------------------------------------------- 2: Biochim Biophys Acta. 2005 Nov 8; [Epub ahead of print] Dietary PUFA modulate the expression of proliferation and differentiation markers in Morris 3924A hepatoma cells. Vecchini A, Ceccarelli V, Nocentini G, Riccardi C, Di Nardo P, Binaglia L. Department of Internal Medicine, Section of Biochemistry, University of Perugia, Via del Giochetto, 3, 06126 Perugia, Italy. The effect of dietary polyunsaturated fatty acids on the expression of differentiation and proliferation markers in Morris 3924A hepatoma cells was investigated. ACT/I rats were conditioned 10 days with diets enriched with linoleic acid or alpha-linolenic acid before subcutaneous hepatoma cell transplantation. After 19 days from the inoculum, the mRNA levels of liver-enriched transcription factors and of their target genes were quantified. Both linoleic acid- and linolenic acid-enriched diets induced a decrease of beta-actin, AFP, PCNA, c-myc and of hepatocyte nuclear factors HNF-1alpha and HNF-4alpha mRNA levels in tumor tissue whereas HNF-3beta expression was induced by both dietary treatments. Only the alpha-linolenic acid-enriched diet was effective in reducing c-jun and increasing albumin mRNA levels. Since albumin is a C/EBPalpha target gene, C/EBPalpha gene transcription was evaluated at both protein and mRNA levels. It was found that alpha-linolenic acid-enriched diet did not enhance the C/EBPalpha mRNA content in hepatoma tissue while inducing C/EBPalpha protein expression with an isoform pattern similar to the hepatic phenotype. This evidence implies that alpha-linolenic acid or one of its metabolic products induce albumin synthesis in hepatoma cells by modulating C/EBPalpha gene expression at post-transcriptional level. PMID: 16290114 [PubMed - as supplied by publisher] --------------------------------------------------------------- 3: Oncogene. 2005 Oct 10; [Epub ahead of print] The hepatitis B virus X protein enhances AP-1 activation through interaction with Jab1. Tanaka Y, Kanai F, Ichimura T, Tateishi K, Asaoka Y, Guleng B, Jazag A, Ohta M, Imamura J, Ikenoue T, Ijichi H, Kawabe T, Isobe T, Omata M. 1Department of Gastroenterology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan. Hepatitis B virus X protein (HBx) has many cellular functions and is a major factor in hepatitis and hepatocellular carcinoma caused by HBV infection. A proteomic approach was used to search for HBx-interacting proteins in order to elucidate the molecular mechanism of hepatocarcinogenesis. HBx was attached to myc and flag tags (MEF tags) and expressed in 293T cells; the protein complex formed within the cells was purified and characterized by mass spectrometry. COP9 signalosome (CSN) subunits 3 and 4 were subsequently identified as HBx-interacting proteins. In addition, CSN subunit 5, Jun activation domain-binding protein 1 (Jab1), was shown to be a novel cellular target of HBx. In vivo and in vitro interactions between HBx and Jab1 were confirmed by standard immunoprecipitation and GST pull-down assays. An analysis of HBx deletion constructs showed that amino acids 30-125 of HBx were responsible for binding to Jab1. Confocal laser microscopy demonstrated that HBx was mainly localized in the cytoplasm, while Jab1 was found mainly in the nucleus and partially in the cytoplasm, and that the two proteins colocalized in the cytoplasm. The cotransfection of HBx and Jab1 resulted in substantial activator protein 1 (AP-1) activation and knockdown of endogenous Jab1 attenuated AP-1 activation caused by HBx. In addition, the coexpression of HBx and Jab1 potentiated phosphorylation of JNK, leading to the subsequent phosphorylation of c-Jun, whereas the level of c-Jun and JNK phosphorylation induced by HBx was decreased in Jab1 knockdown cells. These results suggest that the interaction between HBx and Jab1 enhances HBx-mediated AP-1 activation.Oncogene advance online publication, 10 October 2005; doi:10.1038/sj.onc.1209093. PMID: 16247477 [PubMed - as supplied by publisher] --------------------------------------------------------------- 4: Mol Pharmacol. 2005 Oct 7; [Epub ahead of print] Activation of ERK signaling by epidermal growth factor mediates c-Jun activation and p300 recruitment in keratin 16 gene expression. Wang YN, Chen YJ, Chang WC. National Cheng Kung University. In studies of gene regulation of keratin 16, we reported previously that Sp1 shows a functional cooperation with c-Jun and coactivators p300/CBP in driving the transcriptional regulation of EGF-induced keratin 16 gene expression. In the present study, we found that the stimulated expression of keratin 16 by EGF was mediated mainly through the MEK-ERK signaling pathway. Ser63 and Ser73 on the c-Jun NH2-terminal transactivation domain could be phosphorylated in cells treated with EGF, nevertheless, we found surprisingly that the c-Jun COOH-terminus played a pivotal role in EGF-induced expression of keratin 16. The activation of keratin 16 by EGF treatment could not be enhanced by overexpression of myc-c-JunK3R, in which three putative acetylation lysine residues on the c-Jun COOH-terminus were all mutated into arginines, suggesting that c-Jun acetylation on the COOH-terminus might partially play a functional role in this system. In addition, by using a chromatin immunoprecipitation assay and a DNA affinity precipitation assay, EGF treatment up-regulated the p300 recruitment through ERK signaling to the promoter region in regulating keratin 16 transcriptional activity. Furthermore, the enhancement of acetyl-histone H3 to the keratin 16 chromatin promoter induced by EGF was also mediated via ERK activation. In conclusion, these results strongly suggested both of c-Jun induction and p300 recruitment to gene promoter, mediated through ERK activation, played an essential role in regulating keratin 16 gene expression by EGF. p300 mediated and regulated EGF-induced keratin 16 gene expression, at least in part through multiple mechanisms including a selective acetylation of c-Jun and histone H3. PMID: 16214953 [PubMed - as supplied by publisher] --------------------------------------------------------------- 5: Radiat Res. 2005 Oct;164(4 Pt 1):420-30. Expression of the proto-oncogene Fos after exposure to radiofrequency radiation relevant to wireless communications. Whitehead TD, Brownstein BH, Parry JJ, Thompson D, Cha BA, Moros EG, Rogers BE, Roti Roti JL. Washington University School of Medicine, Radiation Oncology Department, Radiation and Cancer Biology Division, St. Louis, Missouri 63108, USA. In this study the expression levels of the proto-oncogene Fos were measured after exposure to radiofrequency (RF) radiation at two relatively high specific absorption rates (SARs) of 5 and 10 W/kg for three types of modulated signals: 847.74 MHz code division multiple access (CDMA), 835.62 MHz frequency division multiple access (FDMA), and 836.55 MHz time division multiple access (TDMA). This work was undertaken to confirm a previous report by Goswami et al. (Radiat. Res. 151, 300-309, 1999) that CDMA and FDMA radiation caused small but statistically significant increases in Fos levels as cells entered plateau phase during exposure. No effects on Myc or Jun levels were observed in that study. Therefore, in the present study, analyses were restricted to Fos expression during the transition from exponential growth to plateau phase. Fos expression was measured using the real-time polymerase chain reaction (RT-PCR) technique. Serum-stimulated C3H 10T(1/2) cells were used as a positive control for Fos expression. Possible influences of final cell number or pH variability on Fos expression were evaluated. Expression of Fos mRNA in C3H 10T(1/2) cells was not significantly different from that found after sham exposure at either SAR level for any signal modulation. Therefore, the results of Goswami et al. could not be confirmed. PMID: 16187744 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 6: Oncogene. 2005 Sep 12; [Epub ahead of print] Altered regulation of c-jun and its involvement in anchorage-independent growth of human lung cancers. Maeno K, Masuda A, Yanagisawa K, Konishi H, Osada H, Saito T, Ueda R, Takahashi T. [1] 1Division of Molecular Carcinogenesis, Center for Neurological Diseases and Cancer, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan [2] 3Department of Internal Medicine and Molecular Science, Nagoya City University Graduate School of Medical Sciences, Nagoya, Aichi, Japan. The c-jun oncogene is frequently overexpressed in non-small-cell lung cancers (NSCLC), but its functional involvement in lung cancer development has not been clearly elucidated. In this study, we found that among the immediate-early serum responsible genes, exemplified by c-jun, c-fos and c-myc, induction of c-jun in a human bronchial epithelial cell line, BEAS-2B, was dependent on anchorage, in contrast to clear induction of c-fos and c-myc under both anchorage-dependent and -independent conditions. In fact, forced expression of c-jun in BEAS-2B cells significantly increased cell viability and colony formation in soft agar. Furthermore, we also found that such anchorage-dependent regulation of c-jun was lost in a significant fraction of human lung cancer cell lines. Interestingly, suppressed anchorage-independent but not anchorage-dependent growth was noted by constitutive expression of a dominant-negative c-jun mutant in a lung cancer cell line showing dysregulated and sustained c-jun expression in the absence of anchorage. These findings suggest that dysregulated c-jun expression may be involved in the acquisition of anchorage independence in the process of human lung carcinogenesis.Oncogene advance online publication, 12 September 2005; doi:10.1038/sj.onc.1209018. PMID: 16158054 [PubMed - as supplied by publisher] --------------------------------------------------------------- 7: Int J Mol Med. 2005 Oct;16(4):637-43. Overexpression of regucalcin suppresses cell proliferation of cloned normal rat kidney proximal tubular epithelial NRK52E cells. Nakagawa T, Sawada N, Yamaguchi M. Laboratory of Endocrinology and Molecular Metabolism, Graduate School of Nutritional Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan. The role of regucalcin, which is a regulatory protein in intracellular signaling pathway, in the regulation of cell proliferation was investigated by using the cloned normal rat kidney proximal tubular epithelial NRK52E cells over-expressing regucalcin. NRK52E cells were transfected with regucalcin (RC) pCXN2 vector and the multiple neomycin-resistant clones that stably overexpress regucalcin were selected. The regucalcin content of RC/pCXN2-transfected cells used in this study was about 21-fold as compared with that of the parental wild-type NRK52E cells. Wild-type NRK52E cells, pCXN2 vector-transfected cells (mock-type), and RC/pCXN2-transfected cells (transfectants) were cultured for 24, 48, and 72 h in the presence of bovine serum (5%). The cell numbers of wild- and mock-type were significantly increased with the time course of the culture. Cell numbers of transfectants were significantly suppressed as compared with that of wild- and mock-type. The decrease in cell number of wild-type cultured for 72 h in the presence of butyrate (8.3 x 10(-6) or 8.3 x 10(-5) M), rescovitine (10(-8) or 10(-7) M), or sulforaphane (10(-9) M), which is an inhibitor of the cell cycle, was not observed in transfectants. The effect of PD98059 (10(-8) M), staurosporine (10(-10) M) or dibucaine (10(-8)-10(-6) M), which is an inhibitor of protein kinases, in decreasing cell number of wild-type was not seen in transfectants. Moreover, the culture with wortmannin (10(-8) or 10(-7) M), an inhibitor of phosphatidylinositol 3 (PI3)-kinase, or Bay K 8644 (10(-8) or 10(-7) M), an agonist of calcium entry in cells, caused a significant decrease in cell number of the wild-type. This decrease was not observed in transfectants. The result of reverse transcription-polymerase chain reaction (RT-PCR) analysis using specific primers showed that c-jun and chk2 mRNA levels were significantly decreased in transfectant. p53 mRNA level was significantly increased in transfectants. The expression of c-myc, c-fos, cdc2, p21, and G3PDH mRNAs in transfectants was not significantly changed. This study demonstrates that overexpression of regucalcin has a suppressive effect on cell proliferation, which is mediated through various signaling pathways, in the cloned normal rat kidney proximal tubular epithelial NRK52E cells. PMID: 16142398 [PubMed - in process] --------------------------------------------------------------- 8: Cell Cycle. 2005 Oct 27;4(10) [Epub ahead of print] Mechanisms of Tumor Suppression by the SCF(Fbw7). Minella AC, Clurman BE. Divisions of Clinical Research and Human Biology, Fred Hutchinson Cancer Research Center, Seattle, Washinton, USA; Department of Medicine, University of Washington School of Medicine, Seattle, Washinton, USA. SCF ubiquitin ligases regulate the degradation of many proteins involved in the control of cell division and growth. F-box proteins are the SCF components that bind to substrates, and this binding is usually signaled by substrate phosphorylation. The Fbw7/hCdc4 F-box protein was first recognized by its ability to bind cyclin E, and the SCF (Fbw7) is now known to target c-Myc, c-Jun and Notch for degradation in addition to its role in cyclin E proteolysis. Fbw7 thus negatively regulates several key oncoproteins. Accordingly, Fbw7 is a tumor suppressor that is mutated in a wide spectrum of human cancers, and Fbw7 functions as a haploinsufficient tumor suppressor in mice. Because there are three Fbw7 isoforms that reside in different subcellular compartments, as well as multiple Fbw7 substrates that are the products of proto-oncogenes, the mechanisms of tumor suppression by Fbw7 are complex and incompletely understood. In this review we discuss the activities of the SCF(Fbw7) in the context of its role as a tumor suppressor and highlight recent findings demonstrating that dominant oncogenes disable Fbw7 function. PMID: 16131838 [PubMed - as supplied by publisher] --------------------------------------------------------------- 9: Eur J Endocrinol. 2005 Sep;153(3):435-44. ACTH 1-24 inhibits proliferation of adrenocortical tumors in vivo. Zwermann O, Schulte DM, Reincke M, Beuschlein F. Medizinische Klinik-Innenstadt, Ludwig Maximilians University, Munich, Germany. OBJECTIVES: Although several lines of evidence suggest that the overall effects of the ACTH receptor, melanocortin 2 receptor (MC2-R), mediated signal transduction on adrenocortical growth and tumorigenesis are anti-proliferative, activation of MC2-R induces mitogens like jun, fos, and myc and activates the MAPK pathway. In vivo, potential effects of endogenous ACTH on adrenal tumori-genesis can not be separated from effects of other POMC derived peptides. METHODS: Murine adrenocortical tumor cells that lack MC2-R expression (Y6(pcDNA)) and Y6 cells stablely transfected with MC2-R (Y6(MC2-R)) were generated. Presence of functional MC2-R was demonstrated by RT-PCR and Western blot using an antibody for phosphorylated CREB. As a syngenic tumor model, LaHeF1/J mice simultaneously received 10(7) Y6(MC2-R) and Y6(pcDNA) subcutaneously, giving rise to MC2-R positive and negative tumors within the same animal. Animals were treated for 3 weeks in groups of 12 according to the following schedule: group A, control animals receiving saline injection; group B, animals receiving 5.7 ng/injection of a slow release formula of ACTH 1-24 administered i.p. three times a week (aiming at a low physiologic dose); and group C, animals receiving 57 ng/injection of ACTH 1-24 (high physiological dose). RESULTS: Twenty days of ACTH 1-24 treatment did not significantly affect corticosterone levels, endogenous ACTH levels or adrenal and thymus weight compared with saline injection. However, ACTH 1-24 treatment of group B and C mice significantly reduced tumor weight in MC2-R positive tumors in a dose dependent manner (P = 0.03), while no significant difference in tumor mass was observed in MC2-R negative tumors. PCNA and TUNEL staining, together with morphological characterization, demonstrated that these in vivo effects were due to reduced proliferation, while apoptosis and cellular hypertrophy within the tumor remained unchanged. CONCLUSION: MC2-R expression is associated with a less aggressive adrenal tumor phenotype and anti-proliferative effects can be amplified through stimulation with physiological doses of ACTH. PMID: 16131607 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 10: Zhonghua Zheng Xing Wai Ke Za Zhi. 2005 May;21(3):197-200. [Effect of vacuum-assisted closure on the expression of proto-oncogenes and its significance during wound healing] [Article in Chinese] Chen SZ, Cao DY, Li JQ, Tang SY. Department of Plastic Surgery and Burns, Tang Du Hospital, the Fourth Military Medical University, Xi'an 710038, China. cszong@fmmu.edu.cn OBJECTIVE: To study the effects of VAC on starting the process of wound healing and decreasing apoptosis. METHODS: To examine the variations in expression of proto-oncogenes c-myc, c-jun and Bcl-2 in pig wound model with acute full-thickness skin defect and human chronic wounds by immunohistochemistry, calculate the numbers of expressive positive cells and the labelling index (LI), and observe the process of wound healing. RESULTS: (1) In pig experiment, the wound in experimental group was very clean and without obvious exudates, many neoepiderm and granulation tissue rapidly appeared or formed after 6 days, and healed completely by the 25th day. On the contrary, in the wound of control group, more exudates and blood crust could be seen and fewer neoepiderm and granulation tissue appeared after 6 days and was healed by 30th day. Immediately after the wound was created, the expression of c-myc, c-jun and Bcl-2 was lower and mainly situated in nucleus or cytoplasma of the basilar cells. After the wound was created in control group, or after starting the VAC treatment in experimental group, their expression rapidly and obviously increased, the distribution of the positive cells also became enlarged, but the amount of expression decreased rapidly after the expressive peak have reached. In the successive 12 days following the wound was created, the expression of c-myc, c-jun and Bcl-2 in the experimental group was constantly higher than that of the control group. (2) In human chronic wounds, there wasn't obvious secretions and more healthy granulation tissue was rapidly formed after VAC treatment. The expression of c-jun was mainly located in cytoplasma of basilar cells of epithelium, dermal fibroblasts and inflammatory cells, and the positive cell and labelling index obviously decreased. The expression of c-myc and Bcl-2 was mainly in cytoplasma of basilar cells, but the amount of expression and the labelling index became obviously increased after VAC treatment. CONCLUSIONS: VAC could rapidly start the healing course of the pig' s acute skin wound and human chronic wound, decrease atoptosis of the reparative cells, so as to accelerate wound healing. PMID: 16128104 [PubMed - in process] --------------------------------------------------------------- 11: Biochem J. 2005 Aug 26; [Epub ahead of print] PERK is required to activate the stress-activated MAP kinases and induce the expression of the immediate early genes upon disruption of ER calcium homeostasis. Liang SH, Zhang W, McGrath BC, Zhang P, Cavener DR. The eIF2alpha kinase PERK (PEK/EIF2AK3) is essential for the normal function of highly secretory cells in the pancreas and skeletal system, as well as the unfolded protein response (UPR) in mammalian cells. To delineate the regulatory machinery underlying these PERK-dependent stress-responses, gene profiling was employed to assess global changes in gene expression in PERK-deficient mouse embryonic fibroblasts (MEFs). Several immediate-early genes, including c-myc, c-jun, egr-1, and fra-1, displayed PERK-dependent expression in MEFs upon disruption of calcium homeostasis by inhibiting the ER transmembrane SERCA calcium pump. Induction of c-myc and egr-1 by other reagents that elicit the UPR, however, showed variable dependence upon PERK. Induction of c-myc expression by thapsigargin was shown to be linked to key signaling enzymes including phospholipase C, phosphatidylinositol 3-kinase, and p38 mitogen-activated protein kinase (MAPK). Analysis of the phosphorylated status of major components in MAPK signaling pathways indicated that thapsigargin and dithiothreitol but not tunicamycin could trigger the PERK-dependent activation of c-Jun N-terminal kinase (JNK) and p38 MAPK. However, activation of JNK and p38 MAPK by non-ER stress stimuli including UV irradiation, anisomycin, and tumor necrosis factor-alpha was found to be independent of PERK. PERK plays a particularly important role in mediating the global cellular response to ER stress that is elicited by the depletion of calcium from the ER. We suggest that this specificity of PERK function in the UPR is an extension of the normal physiological function of PERK to act as a calcium sensor in the ER. PMID: 16124869 [PubMed - as supplied by publisher] --------------------------------------------------------------- 12: Food Chem Toxicol. 2005 Aug 18; [Epub ahead of print] Molecular mechanism of cell cycle blockage of hepatoma SK-Hep-1 cells by Epimedin C through suppression of mitogen-activated protein kinase activation and increased expression of CDK inhibitors p21(Cip1) and p27(Kip1). Liu TZ, Chen CY, Yiin SJ, Chen CH, Cheng JT, Shih MK, Wang YS, Chern CL. Center for Gerontological Research and Graduate Institute of Medical Biotechnology, Chang-Gang University, Kwei-Shan, Taoyuan, Taiwan, ROC. Reports elsewhere demonstrated that Epimedin C, a constituent isolated from the leaves of Epimedium sagittatum, possessed anti-tumor activity. However, its mechanism of action remains unresolved. Using SK-Hep-1 cells, a poorly-differentiated hepatoma subline, as an experimental model, we present evidence here that the anti-tumor activity of Epimedin C may involve cell cycle blockage. Immunoblotting analyses demonstrated that Epimedin C caused a decreased expression of hyperphosphorylated retinoblastoma (Rb) protein, cyclin D1, c-Myc, and c-Fos. In parallel, we measured the kinase activities and found that CDK2 and CDK4 were suppressed with commensurate increased levels of CDK inhibitors, p21(Cip1) and p27(Kip1). These data suggested that Epimedin C arrested the proliferation of these cells at G(0)/G(1) phase through inhibition of CDK2 and CDK4 activities via an increased induction of p21(Cip1) and p27(Kip1). Alternatively, we investigated whether the anti-proliferative effect of Epimedin C on these cells might involve MAP kinase cascade. Using western blotting technique, we demonstrated that Epimedin C also selectively decreased ERK1/2 phosphorylation. Among the downstream effectors of ERK examined, we found that Epimedin C selectively decreased the expression of c-Fos, but not c-Jun. By EMSA assay, we further demonstrated that decreased c-Fos resulted in the downregulation of AP-1/DNA binding activity. Taken together, the molecular mechanisms of anti-tumor activity of Epimedin C may be proceeded by the combined effects of the cell cycle blockage via either the inhibition of CDK2 and CDK4 activities, with commensurate increase in their inhibitors, p21(Cip1) and p27(Kip1) or negatively modulates the ERK/c-Fos/AP-1 signaling pathway. PMID: 16112786 [PubMed - as supplied by publisher] --------------------------------------------------------------- 13: Toxicol Sci. 2005 Nov;88(1):250-64. Epub 2005 Aug 4. Gene expression profiling of the PPAR-alpha agonist ciprofibrate in the cynomolgus monkey liver. Cariello NF, Romach EH, Colton HM, Ni H, Yoon L, Falls JG, Casey W, Creech D, Anderson SP, Benavides GR, Hoivik DJ, Brown R, Miller RT. GlaxoSmithKline Inc., Safety Assessment, Research Triangle Park, North Carolina 27709, USA. Neal.F.Cariello@gsk.com Fibrates, such as ciprofibrate, fenofibrate, and clofibrate, are peroxisome proliferator-activated receptor-alpha (PPARalpha) agonists that have been in clinical use for many decades for treatment of dyslipidemia. When mice and rats are given PPARalpha agonists, these drugs cause hepatic peroxisome proliferation, hypertrophy, hyperplasia, and eventually hepatocarcinogenesis. Importantly, primates are relatively refractory to these effects; however, the mechanisms for the species differences are not clearly understood. Cynomolgus monkeys were exposed to ciprofibrate at various dose levels for either 4 or 15 days, and the liver transcriptional profiles were examined using Affymetrix human GeneChips. Strong upregulation of many genes relating to fatty acid metabolism and mitochondrial oxidative phosphorylation was observed; this reflects the known pharmacology and activity of the fibrates. In addition, (1) many genes related to ribosome and proteasome biosynthesis were upregulated, (2) a large number of genes downregulated were in the complement and coagulation cascades, (3) a number of key regulatory genes, including members of the JUN, MYC, and NFkappaB families were downregulated, which appears to be in contrast to the rodent, where JUN and MYC are reported to upregulated after PPARalpha agonist treatment, (4) no transcriptional signal for DNA damage or oxidative stress was observed, and (5) transcriptional signals consistent with an anti-proliferative and a pro-apoptotic effect were seen. We also compared the primate data to literature reports of hepatic transcriptional profiling in PPARalpha-treated rodents, which showed that the magnitude of induction in beta-oxidation pathways was substantially greater in the rodent than the primate. PMID: 16081524 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 14: J Am Soc Nephrol. 2005 Oct;16(10):2852-63. Epub 2005 Aug 3. Combined proteomics and pathways analysis of collecting duct reveals a protein regulatory network activated in vasopressin escape. Hoorn EJ, Hoffert JD, Knepper MA. National Institutes of Health, 10 Center Drive, Building 10, Room 6N260, Bethesda, MD 20892, USA. Low sensitivity is characteristic of many proteomics methods. Presented here is an approach that combines proteomics based on difference gel electrophoresis (DIGE) with bioinformatic pathways analysis to identify both abundant and relatively nonabundant proteins in inner medullary collecting duct (IMCD) altered in abundance during escape from vasopressin-induced antidiuresis. Rats received the vasopressin analog dDAVP by osmotic minipump plus either a daily water load (vasopressin escape) or only enough water to replace losses (control). Immunoblotting confirmed the hallmark of vasopressin escape, a decrease in aquaporin-2, and demonstrated a decrease in the abundance of the urea transporter UT-A3. DIGE identified 22 mostly high-abundance proteins regulated during vasopressin escape. These proteins were analyzed using pathways analysis software to reveal protein clusters incorporating the proteins identified by DIGE. A single dominant cluster emerged that included many relatively low-abundance proteins (abundances too low for DIGE identification), including several transcription factors. Immunoblotting confirmed a decrease in total and phosphorylated c-myc, a decrease in c-fos, and increases in c-jun and p53. Furthermore, immunoblotting confirmed hypothesized changes in other proteins in the proposed network: Increases in c-src, receptor for activated C kinase 1, calreticulin, and caspase 3 and decreases in steroid receptor co-activator 1, Grp78/BiP, and annexin A4. This combined approach proved capable of uncovering regulatory proteins that are altered in response to a specific physiologic perturbation without being detected directly by DIGE. The results demonstrate a dominant protein regulatory network in IMCD cells that is altered in association with vasopressin escape, providing a new framework for further studies of signaling in IMCD. PMID: 16079266 [PubMed - in process] --------------------------------------------------------------- 15: J Cell Sci. 2005 Aug 15;118(Pt 16):3717-26. Epub 2005 Aug 2. Abl tyrosine kinase regulates a Rac/JNK and a Rac/Nox pathway for DNA synthesis and Myc expression induced by growth factors. Boureux A, Furstoss O, Simon V, Roche S. CRBM, CNRS FRE2593, 1919 route de Mende, 34293 Montpellier Cedex 05, France. The cytoplasmic tyrosine kinase Abl is a Src substrate required for platelet-derived growth factor (PDGF) receptor signaling leading to Myc expression and DNA synthesis. Abl targets are, however, ill defined. Here we report that the small GTPase Rac is an important effector of its mitogenic function. PDGF-induced Rac activation was impaired in cells with inactive Abl and active Rac overcame the mitogenic defects found in these cells. Rac function required both a Jun N-terminal kinase (JNK) and a NADPH oxidase (Nox) pathway. Furthermore, co-activation of JNK and Nox were sufficient to mimic the Rac mitogenic rescue. Abl also regulated PDGF-induced JNK and Nox activation. Finally, we found that Myc is an important target of this signaling cascade: Myc induction was sensitive to small inhibitors of JNK and Nox activities and forced expression of Myc overcame the G1 block induced by dominant interfering mutants of mitogen-activated protein kinase kinase 4 (MKK4) and Nox2 activating subunit. We concluded that cytoplasmic Abl operates on a Rac/JNK and a Rac/Nox pathway for PDGF-induced Myc induction and DNA synthesis. PMID: 16076903 [PubMed - in process] --------------------------------------------------------------- 16: J Virol. 2005 Aug;79(16):10308-29. ERK1/2 and MEK1/2 induced by Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) early during infection of target cells are essential for expression of viral genes and for establishment of infection. Sharma-Walia N, Krishnan HH, Naranatt PP, Zeng L, Smith MS, Chandran B. Department of Microbiology, Molecular Genetics and Immunology, Mail Stop 3029, The University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, KS 66160, USA. Kaposi's sarcoma-associated herpesvirus (KSHV) in vitro target cell infection is characterized by the expression of the latency-associated genes ORF 73 (LANA-1), ORF 72, and K13 and by the transient expression of a very limited number of lytic genes such as lytic cycle switch gene ORF 50 (RTA) and the immediate early (IE) lytic K5, K8, and v-IRF2 genes. During the early stages of infection, several overlapping multistep complex events precede the initiation of viral gene expression. KSHV envelope glycoprotein gB induces the FAK-Src-PI3K-RhoGTPase (where FAK is focal adhesion kinase) signaling pathway. As early as 5 min postinfection (p.i.), KSHV induced the extracellular signal-regulated kinase 1 and 2 (ERK1/2) via the PI3K-PKCzeta-MEK pathway. In addition, KSHV modulated the transcription of several host genes of primary human dermal microvascular endothelial cells (HMVEC-d) and fibroblast (HFF) cells by 2 h and 4 h p.i. Neutralization of virus entry and infection by PI-3K and other cellular tyrosine kinase inhibitors suggested a critical role for signaling molecules in KSHV infection of target cells. Here we investigated the induction of ERK1/2 by KSHV and KSHV envelope glycoproteins gB and gpK8.1A and the role of induced ERK in viral and host gene expression. Early during infection, significant ERK1/2 induction was observed even with low multiplicity of infection of live and UV-inactivated KSHV in serum-starved cells as well as in the presence of serum. Entry of UV-inactivated virus and the absence of viral gene expression suggested that ERK1/2 induction is mediated by the initial signal cascade induced by KSHV binding and entry. Purified soluble gpK8.1A induced the MEK1/2 dependent ERK1/2 but not ERK5 and p38 mitogen-activated protein kinase (MAPK) in HMVEC-d and HFF. Moderate ERK induction with soluble gB was seen only in HMVEC-d. Preincubation of gpK8.1A with heparin or anti-gpK8.1A antibodies inhibited the ERK induction. U0126, a selective inhibitor for MEK/ERK blocked the gpK8.1A- and KSHV-induced ERK activation. ERK1/2 inhibition did not block viral DNA internalization and had no significant effect on nuclear delivery of KSHV DNA during de novo infection. Analyses of viral gene expression by quantitative real-time reverse transcriptase PCR revealed that pretreatment of cells with U0126 for 1 h and during the 2-h infection with KSHV significantly inhibited the expression of ORF 73, ORF 50 (RTA), and the IE-K8 and v-IRF2 genes. However, the expression of lytic IE-K5 gene was not affected significantly. Expression of ORF 73 in BCBL-1 cells was also significantly inhibited by preincubation with U0126. Inhibition of ERK1/2 also inhibited the transcription of some of the vital host genes such as DUSP5 (dual specificity phosphatase 5), ICAM-1 (intercellular adhesion molecule 1), heparin binding epidermal growth factor, and vascular endothelial growth factor that were up-regulated early during KSHV infection. Several MAPK-regulated host transcription factors such as c-Jun, STAT1alpha, MEF2, c-Myc, ATF-2 and c-Fos were induced early during infection, and ERK inhibition significantly blocked the c-Fos, c-Jun, c-Myc, and STAT1alpha activation in the infected cells. AP1 transcription factors binding to the RTA promoter in electrophoretic mobility shift assays were readily detected in the infected cell nuclear extracts which were significantly reduced by ERK inhibition. Together, these results suggest that very early during de novo infection, KSHV induces the ERK1/2 to modulate the initiation of viral gene expression and host cell genes, which further supports our hypothesis that beside the conduit for viral DNA delivery into the cytoplasm, KSHV interactions with host cell receptor(s) create an appropriate intracellular environment facilitating infection. PMID: 16051824 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 17: Cancer Cell. 2005 Jul;8(1):25-33. The v-Jun point mutation allows c-Jun to escape GSK3-dependent recognition and destruction by the Fbw7 ubiquitin ligase. Wei W, Jin J, Schlisio S, Harper JW, Kaelin WG Jr. Department of Medical Oncology, Dana-Farber Cancer Institute and Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA. The c-Jun and c-Myc oncogenic transcription factors are highly unstable proteins due to polyubiquitination. Similar to c-Myc, we report here that phosphorylation of c-Jun by GSK3 creates a high-affinity binding site for the E3 ligase Fbw7, which targets c-Jun for polyubiquitination and proteasomal degradation. In keeping with this, we found that c-Jun levels were inversely related to GSK3 activity in mammalian cells that had entered the cell cycle. Importantly, phosphorylation of c-Jun by GSK3 requires a priming phosphorylation event at Ser-243. Ser-243 is mutated to phenylalanine in v-Jun and allows it to escape recognition by Fbw7. These findings explain the enhanced stability and oncogenicity of v-Jun relative to its cellular counterpart and reveal that GSK3 and Fbw7 coordinately regulate c-Jun and c-Myc. PMID: 16023596 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 18: Biochem Biophys Res Commun. 2005 Aug 26;334(2):425-32. Depletion of 4-hydroxynonenal in hGSTA4-transfected HLE B-3 cells results in profound changes in gene expression. Patrick B, Li J, Jeyabal PV, Reddy PM, Yang Y, Sharma R, Sinha M, Luxon B, Zimniak P, Awasthi S, Awasthi YC. Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX 77555, USA. Previously, we have shown that overexpression of 4-hydroxy-2-nonenal (HNE)-detoxifying enzyme glutathione S-transferase A4-4 (hGSTA4-4) in human lens epithelial cells (HLE B-3) leads to pro-carcinogenic phenotypic transformation of these cells [R. Sharma, et al. Eur. J. Biochem. 271 (2004) 1960-1701]. We now demonstrate that hGSTA4-transfection also causes a profound change in the expression of genes involved in cell adhesion, cell cycle control, proliferation, cell growth, and apoptosis, which is consistent with phenotypic changes of the transformed cells. The expression of p53, p21, p16, fibronectin 1, laminin gamma1, connexin 43, Fas, integrin alpha6, TGFalpha, and c-jun was down-regulated, while the expression of protein kinase C beta II (PKCbetaII), c-myc, cyclin-dependent kinase 2 (CDK2), and TGFbeta was up-regulated in transfected cells. These results demonstrate that HNE serves as a crucial signaling molecule and, by modulating the expression of genes, can influence cellular functions. PMID: 16005854 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 19: EMBO J. 2005 Jul 20;24(14):2590-601. Epub 2005 Jun 30. Stress response gene ATF3 is a target of c-myc in serum-induced cell proliferation. Tamura K, Hua B, Adachi S, Guney I, Kawauchi J, Morioka M, Tamamori-Adachi M, Tanaka Y, Nakabeppu Y, Sunamori M, Sedivy JM, Kitajima S. Department of Biochemical Genetics, Medical Research Institute and Laboratory of Genome Structure and Regulation, School of Biomedical Science, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo, Japan. The c-myc proto-oncogene encodes a transcription factor that promotes cell cycle progression and cell proliferation, and its deficiency results in severely retarded proliferation rates. The ATF3 stress response gene encodes a transcription factor that plays a role in determining cell fate under stress conditions. Its biological significance in the control of cell proliferation and its crosstalk regulation, however, are not well understood. Here, we report that the serum response of the ATF3 gene expression depends on c-myc gene and that the c-Myc complex at ATF/CREB site of the gene promoter plays a role in mediating the serum response. Intriguingly, ectopic expression of ATF3 promotes proliferation of c-myc-deficient cells, mostly by alleviating the impeded G1-phase progression observed in these cells, whereas ATF3 knockdown significantly suppresses proliferation of wild-type cells. Our study demonstrates that ATF3 is downstream of the c-Myc signaling pathway and plays a role in mediating the cell proliferation function of c-Myc. Our results provide a novel insight into the functional link of the stress response gene ATF3 and the proto-oncogene c-myc. PMID: 15990869 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 20: Ann N Y Acad Sci. 2005 May;1042:481-7. IL-5 Inhibits Apoptosis by Upregulation of c-myc Expression in Human Hematopoietic Cells. Juan SH, Yen JJ, Lee HM, Huang HM. Graduate Institute of Cell and Molecular Biology, Taipei Medical University, No. 250 Wu-Shing Street, Hsinyi District, Taipei 110, Taiwan. cmbhhm@tmu.edu.tw. Interleukin 5 (IL-5) inhibition of apoptosis is required throughout many hematopoietic lineages to regulate proliferation and differentiation. It is not clear how IL-5 mediates the intracellular molecular events regulating the anti-apoptotic effect. The cell lines TF-1 and JYTF-1 expressed different amounts of the IL-5 receptor alpha (IL-5Ralpha) subunit, which caused contrasting effects in response to IL-5. IL-5 supported the survival but not the anti-apoptotic activities of TF-1 cells, which have a low expression of IL-5Ralpha. In contrast, IL-5 supported both the survival and the anti-apoptotic activities of JYTF-1 cells, which overexpressed IL-5Ralpha compared to TF-1 cells. In this study, IL-5 stimulation increased Annexin V binding (indicative of apoptosis) in TF-1 cells and decreased apoptosis in JYTF-1 cells. The proto-oncogenes c-fos, fosB, and c-jun were not detected, whereas junB was induced by IL-5 stimulation in both types of cells. Most importantly, IL-5 significantly induced c-myc expression in JYTF-1 cells, but did not in TF-1 cells. These results are consistent with the possibility that IL-5 inhibits apoptosis in JYTF-1 cells via the upregulation of c-myc expression. PMID: 15965094 [PubMed - in process] --------------------------------------------------------------- 21: Exp Cell Res. 2005 Sep 10;309(1):12-23. Neuregulin-regulated gene expression in mammary carcinoma cells. Amin DN, Tuck D, Stern DF. Department of Pathology, Yale University School of Medicine, BML-342, 310 Cedar Street, P.O. Box 208023, New Haven, CT 06520-8023, USA. Recent studies have suggested that autocrine production of Neuregulin (NRG), a growth factor that activates members of the Epidermal Growth Factor Receptor/ErbB family of proto-oncogenes, is sufficient for breast tumor initiation and progression. To elucidate the molecular mechanisms regulating these events, we undertook a global analysis of genes regulated by NRG in luminal mammary epithelial cell lines. Gene expression profiling of estrogen receptor-positive T47D cells exposed to NRG-1 revealed both previously identified and novel targets of NRG activation. Profiling of other estrogen receptor-positive breast cancer cell lines, MCF7 and SUM44, yielded a group of twenty-one genes whose transcripts are upregulated by NRG in all three lines tested. The NRG targets are FBJ murine osteosarcoma viral oncogene homolog B, Early growth response 1, v-jun avian sarcoma virus 17 oncogene homolog, Activating transcription factor 3, Homo sapiens cDNA FLJ31636 fis, Jun B proto-oncogene, Forkhead box C1, Platelet/endothelial cell adhesion molecule 1, NADPH-dependent retinol dehydrogenase/reductase, Dual specificity phosphatase 5, NGF inducible protein TIS21, Connective tissue growth factor, Jun D proto-oncogene, Serum response factor, Cullin 1, v-myc avian myelocytomatosis viral oncogene, Transient receptor potential channel 1, Low density lipoprotein receptor, Transforming growth factor beta 1, Nucleoporin 88 kDa, and Pleckstrin homology-like domain A1. Since NRG activation of these cells induces resistance to anti-hormonal therapy, the identified genes may provide clues to molecular events regulating mammary tumor progression and hormone independence. PMID: 15963498 [PubMed - in process] --------------------------------------------------------------- 22: Clin Cancer Res. 2005 Jun 15;11(12):4295-304. Elevated expression of Wnt antagonists is a common event in hepatoblastomas. Koch A, Waha A, Hartmann W, Hrychyk A, Schuller U, Waha A, Wharton KA Jr, Fuchs SY, von Schweinitz D, Pietsch T. Department of Neuropathology, University of Bonn Medical Center, Bonn, Germany. Hepatoblastomas are the most frequent malignant liver tumors of childhood. A high frequency of activating beta-catenin mutations in hepatoblastomas indicates that the Wnt signaling pathway plays an important role in the development of this embryonic neoplasm. Stabilization of beta-catenin leads to an increased formation of nuclear beta-catenin-T-cell factor complexes and altered expression of Wnt-inducible target genes. In this study, we analyzed the mRNA expression levels of nine Wnt genes, including c-JUN, c-MYC, CYCLIN D1, FRA-1, NKD-1, ITF-2, MMP-7, uPAR, and beta-TRCP, by competitive reverse transcription-PCR. We analyzed 23 hepatoblastoma biopsies for which matching liver tissue was available, 6 hepatoblastoma cell lines, and 3 human fetal liver samples. beta-TRCP and NKD-1 were highly expressed in all hepatoblastoma samples, independent of the beta-catenin mutational status, in comparison with their nontumorous counterparts. beta-TRCP mRNA overexpression was associated with accumulation of intracytoplasmic and nuclear beta-TrCP protein. In human liver tumor cells without beta-catenin mutations, Nkd-1 inhibited the Wnt-3a-activated Tcf-responsive-luciferase reporter activity, whereas Nkd-1 in hepatoblastomas with beta-catenin mutations had no antagonistic effect. Our data emphasize the inhibitory effect of beta-TrCP and Nkd-1 on the Wnt signaling pathway in a manner analogous to Conductin (AXIN2) and Dkk-1, inhibitors shown previously to be up-regulated in hepatoblastomas. Our findings indicate that overexpression of the Wnt antagonists Nkd-1 and beta-TrCP reveals an activation of the Wnt signaling pathway as a common event in hepatoblastomas. We propose that Nkd-1 and beta-TrCP may be used as possible diagnostic markers for the activated Wnt signaling pathway in hepatoblastomas. PMID: 15958610 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 23: Biochim Biophys Acta. 2005 May 30;1740(2):189-201. Epub 2005 Jan 4. Different effect of beta-carotene on proliferation of prostate cancer cells. Dulinska J, Gil D, Zagajewski J, Hartwich J, Bodzioch M, Dembinska-Kiec A, Langmann T, Schmitz G, Laidler P. Institute of Medical Biochemistry, Jagiellonian University Medical College, ul. M. Kopernika 7, 31-034 Krakow, Poland. mblitewk@cyf-kr.edu.pl It was shown that high doses of beta-carotene (>30 microM) decrease proliferation of prostate cancer cells in vitro. However, it is rather doubtful whether such concentration of beta-carotene is really accessible at cellular level. We studied the effect of 3 and 10 microM beta-carotene on proliferation and gene expression in LNCaP and PC-3 prostate cancer cell lines. Beta-carotene--more efficiently absorbed from medium by androgen-sensitive LNCaP cells--increased proliferation of LNCaP cells whereas it had weaker effect on PC-3 cells. Initial global analysis of expression of genes in both cell lines treated with 10 microM beta-carotene (Affymetrix HG-U133A) showed remarkable differences in number of responsive genes. Their recognition allows for conclusion that differences between prostate cancer cell lines in response to beta-carotene treatment are due to various androgen sensitivities of LNCaP and PC-3 cells. Detailed analysis of expression of selected genes in beta-carotene treated LNCaP cells at the level of mRNA and protein indicated that the observed increase of proliferation could have been the result of slight induction of a few genes affecting proliferation (c-myc, c-jun) and apoptosis (bcl-2) with no significant effect on major cell cycle control genes (cdk2, RB, E2F-1). PMID: 15949686 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 24: Exp Toxicol Pathol. 2005 Apr;56(6):333-9. Nitrofurazone-induced gene expressions in rat hepatocytes and their modification by N-acetylcysteine. Ito K, Kajikawa S, Nii A, Doi K. Safety Research Laboratories, Yamanouchi Pharmaceutical Co., Ltd., 1-8 Azusawa 1-Chome, Itabashi-ku, Tokyo 174-8511, Japan. itou@yamanouchi.co.jp The antibiotic nitrofurazone (NF) at a subtoxic dose has been shown to increase hepatocyte DNA synthesis with no preceding cell damage or necrosis. This was suppressed by concomitant administration of the antioxidant N-acetylcysteine (NAC), which suggests that free radical production is involved in the process. In this study, male F344 rats were given a single oral subtoxic dose of NF to investigate the changes in genes implicated in hepatocyte proliferation between 1 and 20h postdose by real-time PCR. Some rats were also given NAC to examine the involvement of free radicals. There were transient and sequential increases in mRNA levels of c-myc and c-jun shortly after the administration, followed by tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-alpha (TGF-alpha), c-Ha-ras, and cyclin E. The increases were blocked by concomitant administration of NAC. In contrast, there were no NF-specific increases in c-fos, hepatocyte growth factor, epidermal growth factor or cyclin D1 mRNAs. These results indicate that the induction of hepatocyte proliferation by NF is triggered by free radicals, with a pathway involving increases in c-jun, c-myc, TNF-alpha, TGF-alpha, c-Ha-ras, and cyclin E. The results also indicate that NF-induced proliferation resembles that of other mitogens. PMID: 15945272 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 25: Biochim Biophys Acta. 2005 Jun 15;1735(1):30-40. Epub 2005 Feb 16. Molecular and cellular effects of cis-9, trans-11-conjugated linoleic acid in enterocytes: effects on proliferation, differentiation, and gene expression. Lampen A, Leifheit M, Voss J, Nau H. Institut fur Lebensmitteltoxikologie, Stiftung Tierarztliche Hochschule Hannover, Germany. Alfonso.Lampen@tiho-hannover.de It has been hypothesized that dietary conjugated linoleic acids (CLA) may inhibit colon tumorigenesis. The aim of our study was to investigate the cellular and molecular effects of cis-9 (9Z), trans-11 (11E)-CLA on the proliferation, differentiation, interaction with peroxisome proliferator-activated receptors (PPARs), and expression of genes relevant in the APC-beta-catenin-TCF4 signalling pathway in human HT-29 and Caco-2 colon cells. We found that 9Z,11E-CLA inhibited the proliferation of HT-29 and Caco-2 cells. Trans-vaccenic acid (VA) showed no antiproliferative effects at all. We determined that 9Z,11E-CLA induced cell differentiation as measured by intestinal alkaline phosphatase (IAP) enzyme activity in Caco-2 cells, mRNA expression of IAP, and activation of a 5' flanking region of IAP. The 9Z,11E-CLA activated human PPARdelta as measured in a reporter gene assay. Treatment of HT29 cells in the poliferation phase with 9Z,11E-CLA repressed mRNA-expression of proliferation genes such as c-myc, cyclin D1 and c-jun in a concentration dependent manner. The promoter activities of c-myc and AP1 were also inhibited after incubation with 9Z,11E-CLA. beta-Catenin mRNA and protein expression was also repressed by the treatment with 9Z,11E-CLA. In addition, the mRNA expression of PPARdelta was repressed by treatment of the HT-29 cells with 9Z,11E-CLA. We conclude that 9Z,11E-CLA has an antiproliferative effect at the cellular and molecular levels in human colon cells. The results indicate that the preventive effects of CLA in the development of colon cancer may be due to their downregulation of some target genes of the APC-beta-catenin-TCF-4- and PPARdelta signalling pathway. PMID: 15935729 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 26: Exp Cell Res. 2005 Jul 1;307(1):231-46. Epub 2005 Apr 19. Smad7 induces tumorigenicity by blocking TGF-beta-induced growth inhibition and apoptosis. Halder SK, Beauchamp RD, Datta PK. Department of Surgery, Vanderbilt University School of Medicine, 1161 21st Avenue South, D5230 MCN, Nashville, TN 37232, USA. Smad proteins play a key role in the intracellular signaling of the transforming growth factor beta (TGF-beta) superfamily of extracellular polypeptides that initiate signaling to regulate a wide variety of biological processes. The inhibitory Smad, Smad7, has been shown to function as intracellular antagonists of TGF-beta family signaling and is upregulated in several cancers. To determine the effect of Smad7-mediated blockade of TGF-beta signaling, we have stably expressed Smad7 in a TGF-beta-sensitive, well-differentiated, and non-tumorigenic cell line, FET, that was derived from human colon adenocarcinoma. Smad7 inhibits TGF-beta-induced transcriptional responses by blocking complex formation between Smad 2/3 and Smad4. While Smad7 has no effect on TGF-beta-induced activation of p38 MAPK and ERK, it blocks the phosphorylation of Akt by TGF-beta and enhances TGF-beta-induced phosphorylation of c-Jun. FET cells expressing Smad7 show anchorage-independent growth and enhance tumorigenicity in athymic nude mice. Smad7 blocks TGF-beta-induced growth inhibition by preventing TGF-beta-induced G1 arrest. Smad7 inhibits TGF-beta-mediated downregulation of c-Myc, CDK4, and Cyclin D1, and suppresses the expression of p21(Cip1). As a result, Smad7 inhibits TGF-beta-mediated downregulation of Rb phosphorylation. Furthermore, Smad7 inhibits the apoptosis of these cells. Together, Smad7 may increase the tumorigenicity of FET cells by blocking TGF-beta-induced growth inhibition and by inhibiting apoptosis. Thus, this study provides a mechanism by which a portion of human colorectal tumors may become refractory to tumor-suppressive actions of TGF-beta that might result in increased tumorigenicity. PMID: 15922743 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 27: Apoptosis. 2005 May;10(3):525-35. Sustained Akt/PKB activation and transient attenuation of c-jun N-terminal kinase in the inhibition of apoptosis by IGF-1 in vascular smooth muscle cells. Allen RT, Krueger KD, Dhume A, Agrawal DK. Departments of Medical Microbiology and Immunology, Creighton University School of Medicine, Omaha, NE 68178, USA. Characteristics of hVSMC apoptosis and its inhibition by insulin-like growth factor-1 (IGF-1) remain unclear. Also unclear is whether a balance in hVSMCs exists whereby c-Jun N-terminal stress kinases (JNK) promote apoptosis while extracellular signal-regulated (ERK1/2) MAP kinases inhibit cell death. In this study, we examined the involvement of Akt/PKB and its upstream kinase, PDK1 and whether JNK activation correlated with human and rat VSMC apoptosis induced by staurosporine and by c-myc, respectively. We observed a strong, sustained JNK activation (and c-Jun phosphorylation), which correlated with VSMC apoptosis. IGF-1 (13.3 nM), during apoptosis inhibition, transiently inhibited JNK activity at 1 h in a phosphatidylinositol 3-kinase (PI3-K)- and MEK-ERK-dependent manner, as wortmannin (100 nM) or PD98059 (30 muM) partially attenuated the IGF-1 effect. PKC down-regulation had no effect on JNK inhibition by IGF-1. While IGF-1 alone produced a strong phosphorylation of Akt/PKB in hVSMCs up to 6 h, it was notably stronger and more sustained during ratmyc and hVSMCs apoptosis inhibition. Further, whereas transient expression of phosphorylated Akt protected VSMCs from apoptosis by nearly 50%, expression of dominant interfering alleles of Akt or PDK1 strongly inhibited IGF-1-mediated VSMC survival. These results demonstrate for the first time that transient inhibition of a pro-apoptotic stimulus in VSMCs may be sufficient to inhibit a programmed cell death and that sustained anti-apoptotic signals (Akt) elicited by IGF-1 are augmented during a death stimulus. Furthermore, PI3-K and ERK-MAPK pathways may cooperate to protect VSMCs from cell death. PMID: 15909115 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 28: Apoptosis. 2005 May;10(3):503-12. Disease-associated fibronectin matrix fragments trigger anoikis of human primary ligament cells: p53 and c-myc are suppressed. Dai R, Iwama A, Wang S, Kapila YL. Department of Stomatology, School of Dentistry, University of California, San Francisco, CA 94143-0512, USA. Inflammation in periodontal disease is characterized by the breakdown of the extracellular matrix. This study shows that an inflammation-associated matrix breakdown fragment of fibronectin (FN) induces anoikis of human periodontal ligament (PDL) cells. This 40 kDa fragment was identified in human inflammatory crevicular fluid and is associated with disease status. Previously, we reported that a similar recombinant FN fragment triggered apoptosis of PDL cells by an alternate apoptotic signaling pathway that requires transcriptional downregulation of p53 and c-myc. Thus, to determine whether the physiologically relevant 40 kDa fragment triggers apoptosis in these cells, the 40 kDa fragment was generated and studied for its apoptotic properties. The 40 kDa fragment induces apoptosis of PDL cells, and preincubation of cells with intact vitronectin, FN, and to a limited extent collagen I, rescue this apoptotic phenotype. These data suggest that the 40 kDa fragment prevents PDL cell spreading, thereby inducing anoikis. The signaling pathway also involves a downregulation in p53 and c-myc, as determined by Western blotting and real time quantitative PCR. These data indicate that an altered FN matrix as is elaborated in inflammation induces anoikis of resident cells and thus may contribute to disease progression. PMID: 15909113 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 29: Cancer Biol Ther. 2005 May;4(5):506-8. Epub 2005 May 5. Tumor suppressor activities of the fbw7 e3 ubiquitin ligase receptor. Fuchs SY. The F-box protein Fbw7/Sel-10/hCdc4/Ago, which is known to regulate ubiquitination and degradation of numerous important regulators of cell division and death including Notch, cyclin E, c-Jun and c-Myc, has been recently rediscovered as a p53-dependent tumor suppressor. Delineation of the mechanisms of Fbw7 anti-oncogenic activities and of its inactivation in human cancers is expected to gain an important insight into tumorigenesis. PMID: 15908780 [PubMed - in process] --------------------------------------------------------------- 30: Blood. 2005 Aug 15;106(4):1382-91. Epub 2005 May 12. c-Jun N-terminal kinase (JNK) is required for survival and proliferation of B-lymphoma cells. Gururajan M, Chui R, Karuppannan AK, Ke J, Jennings CD, Bondada S. Department of Microbiology, Immunology, & Molecular Genetics, University of Kentucky, Lexington, KY, USA. Several primary murine and human B lymphomas and cell lines were found to constitutively express high levels of the activated form of c-jun N-terminal kinase (JNK), a member of the mitogen-activated protein (MAP) kinase family. Proliferation of murine B lymphomas CH31, CH12.Lx, BKS-2, and WEHI-231 and the human B lymphomas BJAB, RAMOS, RAJI, OCI-Ly7, and OCI-Ly10 was strongly inhibited by SP600125, an anthrapyrazolone inhibitor of JNK, in a dose-dependent manner. The lymphoma cells underwent apoptosis and arrested at the G2/M phase of cell cycle. Furthermore, JNK-specific small interfering RNA (siRNA) inhibited the growth of both murine and human B lymphomas. Thus in the B-lymphoma model, JNK appears to have a unique prosurvival role. Survival signals provided by CD40 and interleukin-10 (IL-10) together reversed the growth inhibition induced by the JNK inhibitor. c-Myc protein levels were reduced in the presence of both SP600125 and JNK-specific siRNA, and CD40 ligation restored c-Myc levels. Moreover, Bcl-xL rescued WEHI-231 cells from apoptosis induced by the JNK inhibitor. The JNK inhibitor also reduced levels of early growth response gene-1 (Egr-1) protein, and overexpressing Egr-1 partially rescued lymphoma cells from apoptosis. Thus, JNK may act via c-Myc and Egr-1, which were shown to be important for B-lymphoma survival and growth. PMID: 15890690 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 31: Exp Cell Res. 2005 May 15;306(1):168-79. The Ras suppressor Rsu-1 binds to the LIM 5 domain of the adaptor protein PINCH1 and participates in adhesion-related functions. Dougherty GW, Chopp T, Qi SM, Cutler ML. Department of Pathology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Road, B3122, Bethesda, MD 20814, USA. Rsu-1 is a highly conserved leucine rich repeat (LRR) protein that is expressed ubiquitously in mammalian cells. Rsu-1 was identified based on its ability to inhibit transformation by Ras, and previous studies demonstrated that ectopic expression of Rsu-1 inhibited anchorage-independent growth of Ras-transformed cells and human tumor cell lines. Using GAL4-based yeast two-hybrid screening, the LIM domain protein, PINCH1, was identified as the binding partner of Rsu-1. PINCH1 is an adaptor protein that localizes to focal adhesions and it has been implicated in the regulation of adhesion functions. Subdomain mapping in yeast revealed that Rsu-1 binds to the LIM 5 domain of PINCH1, a region not previously identified as a specific binding domain for any other protein. Additional testing demonstrated that PINCH2, which is highly homologous to PINCH1, except in the LIM 5 domain, does not interact with Rsu-1. Glutathione transferase fusion protein binding studies determined that the LRR region of Rsu-1 interacts with PINCH1. Transient expression studies using epitope-tagged Rsu-1 and PINCH1 revealed that Rsu-1 co-immunoprecipitated with PINCH1 and colocalized with vinculin at sites of focal adhesions in mammalian cells. In addition, endogenous P33 Rsu-1 from 293T cells co-immunoprecipitated with transiently expressed myc-tagged PINCH1. Furthermore, RNAi-induced reduction in Rsu-1 RNA and protein inhibited cell attachment, and while previous studies demonstrated that ectopic expression of Rsu-1 inhibited Jun kinase activation, the depletion of Rsu-1 resulted in activation of Jun and p38 stress kinases. These studies demonstrate that Rsu-1 interacts with PINCH1 in mammalian cells and functions, in part, by altering cell adhesion. PMID: 15878342 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 32: Toxicol Lett. 2005 Jul 28;158(1):20-9. A novel method for generation of signature networks as biomarkers from complex high throughput data. Nikolsky Y, Ekins S, Nikolskaya T, Bugrim A. Computational Biology Genego Inc., 500 Renaissance Drive, Suite 106, St. Joseph, MI 49085, USA. Traditionally, gene signatures are statistically deduced from large gene expression and proteomics datasets and have been applied as an experimental molecular diagnostic technique that is sensitive to experimental design and statistical treatment. We have developed and applied the approach of "signature networks" which overcomes some of the drawbacks of clustering methods. We have demonstrated signature network assembly, functional analysis and logical operations on the networks that can be generated. In addition, we have used this technique in a proof of concept study to compare the effect of differential drug treatment using 4-hydroxytamoxifen and estrogen on the MCF-7 breast cancer cell line from a previously published study. We have shown that the two compounds can be differentiated by the networks of interacting genes. Both networks consist of a core module of genes including c-Fos as part of c-Fos/c-Jun heterodimer and c-Myc which is clearly visible. Using algorithms in our MetaCore software we are able to subtract the 4-hydroxytamoxifen and estrogen networks to further understand differences between these two treatments and show that the estrogen network is assembled around the core with other modules essential for all phases of the cell cycle. For example, Cyclin D1 is present in networks for the estrogen treated cells from two separate studies. These signature networks represent an approach to identify biomarkers and a general approach for discovering new relationships in complex high throughput toxicology data. PMID: 15871913 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 33: Ultrasound Med Biol. 2005 May;31(5):703-8. Early gene response to low-intensity pulsed ultrasound in rat osteoblastic cells. Sena K, Leven RM, Mazhar K, Sumner DR, Virdi AS. Department of Anatomy and Cell Biology, Rush University Medical Center, 600 S. Paulina Street, Chicago, IL 60612, USA. The aim of the current research was to quantify the changes in gene expression in rat bone marrow derived stromal cells (BMSC) to low intensity pulsed ultrasound (LIPUS) during early time points after the ultrasound application. LIPUS at 1.5 MHz, 30 mW/cm(2) was applied to BMSC for a single 20 min treatment. Real-time PCR was carried out to quantify the expression of early response genes and bone differentiation marker genes 0.5, 1, 3, 6 and 12 h after the end of the LIPUS treatment. Compared with the controls, LIPUS treatment resulted in elevated transient expression of early response genes (c-jun, c-myc, COX-2, Egr-1, TSC-22) as well as the bone differentiation marker genes, osteonectin and osteopontin, at 3 h. This induction of early response genes as well as extracellular matrix genes associated with cell proliferation and differentiation may represent the effect of LIPUS to cells of osteoblastic lineage. PMID: 15866420 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 34: J Allergy Clin Immunol. 2005 Apr;115(4):856-63. Jun N-terminal kinase is essential for CD40-mediated IgE class switching in B cells. Jabara HH, Geha RS. Division of Immunology, Children's Hospital, KARP Building #10126, 1 Blackfan Circle, Boston, MA 02115, USA. Haifa.jubara@childrens.harvard.edu BACKGROUND: CD40 ligation activates nuclear factor kappaB (NF-kappaB) and the mitogen-activated protein kinases p38 and C-Jun N-terminal kinase (JNK) and causes immunoglobulin class-switch recombination (CSR) in B cells. Both NF-kappaB and p38 are important for CD40-mediated CSR. The role of JNK activation in CD40-mediated isotype switching is unknown. OBJECTIVE: We sought to determine the role of JNK activation in CD40-mediated isotype switching. METHODS: Splenic B cells from BALB/c mice were stimulated with anti-CD40 mAb and IL-4 or with soluble CD40 ligand in the presence or absence of SP600125, an anthrapyrazolone inhibitor of JNK. The following events were examined: IgE production by means of ELISA; S(mu)-S(epsilon) deletional switch recombination by means of digestion circularization PCR; Cepsilon germline, mature epsilon, and activation-induced deaminase (AID) transcription by means of RT-PCR; and proliferation by tritiated thymidine incorporation and surface expression of CD23, CD54, and CD86 by means of FACS analysis. RESULTS: SP600125 at 10 microM drastically inhibited JNK phosphorylation but had little effect on CD40-mediated p38 phosphorylation and expression of the NF-kappaB dependent genes c-Myc and bcl-xL. SP600125 inhibited IgE synthesis by approximately 88% but had no effect on B-cell proliferation and survival in response to anti-CD40 + IL-4 or on upregulation of CD23, CD54, and CD86 in response to CD40 ligation. Analysis of molecular events involved in IgE class switching revealed that SP600125 had no effect on the expression of C(epsilon) germline and AID transcripts. In contrast, SP600125 severely reduced S(mu)-S(epsilon) switch recombination and expression of mature epsilon transcripts. CONCLUSION: These results demonstrate that JNK activation is essential for CD40-mediated CSR to IgE and suggest that JNK is important for AID activity in B cells. PMID: 15806010 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 35: Pathol Oncol Res. 2005;11(1):32-9. Epub 2005 Mar 31. Immunohistochemical analysis of c-myc, c-jun and estrogen receptor in normal, hyperplastic and neoplastic endometrium. Bircan S, Ensari A, Ozturk S, Erdogan N, Dundar I, Ortac F. Department of Pathology, School of Medicine, University of Ankara, Ankara, Turkey. bircans2000@yahoo.com To evaluate the role of c-jun and c-myc proto-oncogenes in normal, hyperplastic and neoplastic endometrium in relation to estrogen receptor (ER) status and to investigate whether these genes can be related to other histopathological features of endometrial carcinoma, 32 endometrial carcinomas, 38 endometrial hyperplasias and 22 cyclic endometria (10 proliferative and 12 secretory) were evaluated histologically. Endometrial hyperplasia cases were classified as simple and complex hyperplasia without atypia, and atypical hyperplasia. Endometrial carcinoma cases were subtyped according to the International Society of Gynecological Pathologists. Modified FIGO system was used for both grading and staging. Immunohistochemical examination was performed using antibodies to ER-alpha, c-myc and c-jun with streptavidin-biotin-peroxidase technique. The mean percentage of ER-alpha positive cells changed cyclically during the menstrual cycle, and it was the highest (96%) and the lowest (31.6%) in proliferative and carcinomatous endometrium, respectively. There was a statistically significant difference between proliferative and secretory phases and proliferative and carcinomatous endometrium in relation to ER-alpha staining (p<0.05). There was also a statistically significant difference with respect to ERalpha reactivity between secretory phase and each hyperplastic group, as well as between the carcinoma group and each hyperplastic group (p<0.05). Although not significant, the mean percentage of c-myc expressing cells in the carcinoma group was higher (15.3%) than that of proliferative phase and hyperplastic groups. The mean percentage of c-jun positive cells in proliferative endometrium was slightly higher than in secretory endometrium, and it was the highest in atypical hyperplastic endometrium (28.3%), but there was no statistically significant difference between the groups. In carcinoma cases, a positive correlation was observed between c-jun positivity and tumor grade (p=0.027, r=0.3908), but such a correlation with c-myc was not found. A positive correlation was detected between ER-alpha and c-myc expression (p=0.038, r=0.3686). A progressive loss of ER seems to be correlated with increasing malignant transformation. C-myc expression might play a role in the development of endometrial carcinoma via ER. The association between c-jun and ER appears to be lost in endometrial carcinoma. The relationship between c-myc, c-jun and ER appears to be altered in endometrial carcinoma compared to that of menstrual endometrium. PMID: 15800680 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 36: Cancer Chemother Pharmacol. 2005 Jul;56(1):22-8. Epub 2005 Mar 25. Effects of histone deacetylase inhibitor FR901228 on the expression level of telomerase reverse transcriptase in oral cancer. Murakami J, Asaumi J, Kawai N, Tsujigiwa H, Yanagi Y, Nagatsuka H, Inoue T, Kokeguchi S, Kawasaki S, Kuroda M, Tanaka N, Matsubara N, Kishi K. Department of Oral and Maxillofacial Radiology, Field of Tumor Biology, Graduate School of Medicine and Dentistry, Okayama University, Shikata-cho, Okayama 700-8525, Japan. jun-m@md.okayama-u.ac.jp We speculated whether or not the expression level of telomerase reverse transcriptase (hTERT) would be modulated by agents targeting epigenetics in oral cancer cell lines. Although hTERT is known to be targeted by epigenetic changes, it remains unclear how chemoagents targeting epigenetics work on hTERT transcription. In the present study, the epigenetic effects of the histone deacetylase (HDAC) inhibitor FR901228 on hTERT transcription in oral cancer cell lines were analyzed by RT-PCR. The mRNA expression of hTERT was upregulated after exposure to FR901228 in hTERT-negative Hep2 cells, and even SAS and KB cells expressed high levels of hTERT. Moreover, cotreatment of protein synthesis inhibitor cycloheximide (CHX) resulted in the induction of hTERT transcription by FR901228. This suggests that the induction of hTERT by FR901228 requires de novo protein synthesis to some extent and is more likely a direct than an indirect effect on epigenetic changes such as histone acetylation/deacetylation. We further examined the effect of FR901228 on c-myc protein, which is one of the main hTERT transcription activators. FR901228 repressed c-myc protein only in the absence of CHX, and depended on the enhancement of de novo protein synthesis. Our results indicate that c-myc protein is repressed indirectly by FR901228 but may not contribute to FR901228-induced hTERT transcription. The present study showed that the HDAC inhibitor FR901228 induced the hTERT gene by a complex mechanism that involved transcription factors other than c-myc, in addition to inhibition of histone deacetylation. PMID: 15791453 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 37: J Biol Chem. 2005 May 20;280(20):19992-9. Epub 2005 Mar 18. JNK1 and JNK2 oppositely regulate p53 in signaling linked to apoptosis triggered by an altered fibronectin matrix: JNK links FAK and p53. Tafolla E, Wang S, Wong B, Leong J, Kapila YL. Department of Stomatology, School of Dentistry, University of California, San Francisco, 94143-0512, USA. The extracellular matrix regulates many cellular processes, including survival, and alterations in the matrix or in matrix survival signals can trigger apoptosis. Previously, we showed that an altered fibronectin matrix triggers apoptosis in primary cells via a novel pathway regulated by transcriptionally mediated decreases in p53 and c-Myc levels. Here we report that this apoptotic mechanism is propagated by decreased phosphorylation of focal adhesion kinase (FAK), which is linked to increased phosphorylation of c-Jun N-terminal kinase (JNK) and to decreased levels of p53. FAK is physically and spatially linked to JNK and p53, which relocalize from the nucleus to the cell membrane to mediate this interaction. Further, p53 participates in a feedback mechanism with JNK to regulate this apoptotic process and is oppositely regulated by JNK1 and JNK2. PMID: 15778501 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 38: J Virol. 2005 Apr;79(7):4246-56. Silencing of integrated human papillomavirus type 18 oncogene transcription in cells expressing SerpinB2. Darnell GA, Antalis TM, Rose BR, Suhrbier A. Queensland Institute of Medical Research, University of Queensland, Brisbane, Queensland, Australia. The serine protease inhibitor SerpinB2 (PAI-2), a major product of differentiating squamous epithelial cells, has recently been shown to bind and protect the retinoblastoma protein (Rb) from degradation. In human papillomavirus type 18 (HPV-18)-transformed epithelial cells the expression of the E6 and E7 oncoproteins is controlled by the HPV-18 upstream regulatory region (URR). Here we illustrate that PAI-2 expression in the HPV-18-transformed cervical carcinoma line HeLa resulted in the restoration of Rb expression, which led to the functional silencing of transcription from the HPV-18 URR. This caused loss of E7 protein expression and restoration of multiple E6- and E7-targeted host proteins, including p53, c-Myc, and c-Jun. Rb expression emerged as sufficient for the transcriptional repression of the URR, with repression mediated via the C/EBPbeta-YY1 binding site (URR 7709 to 7719). In contrast to HeLa cells, where the C/EBPbeta-YY1 dimer binds this site, in PAI-2- and/or Rb-expressing cells the site was occupied by the dominant-negative C/EBPbeta isoform liver-enriched transcriptional inhibitory protein (LIP). PAI-2 expression thus has a potent suppressive effect on HPV-18 oncogene transcription mediated by Rb and LIP, a finding with potential implications for prognosis and treatment of HPV-transformed lesions. PMID: 15767426 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 39: Mol Carcinog. 2005 Apr;42(4):213-21. Activation of Wnt/beta-catenin/Tcf signaling in mouse skin carcinogenesis. Bhatia N, Spiegelman VS. Department of Dermatology, University of Wisconsin Medical School, 1300 University Avenue B-25, Madison, WI 53706, USA. Although Wnt/beta-catenin/Tcf signaling pathway has been shown to be an important factor in the development of many malignancies including colorectal, ovarian, prostate, and many other cancers, little is known about its role in non-melanoma skin cancers. Here, we report the first evidence that beta-catenin/Tcf signaling pathway is constitutively activated in non-melanocytic skin tumors induced by two stage chemical carcinogenesis protocol. Mouse skin tumors showed cytoplasmic and nuclear accumulation of beta-catenin, and upregulation of beta-catenin/Tcf target genes (c-myc and c-jun). We found high levels of skin-expressed Wnt proteins (Wnt 3, 4, and 10b) in different parts of the tumors, likely representing key upstream events in beta-catenin/Tcf activation during mouse skin carcinogenesis. Inhibition of beta-catenin/Tcf signaling by ectopic expression of dominant negative Tcf4 resulted in significant inhibition of growth in squamous cell carcinoma cells. A role of the constitutive activation of beta-catenin/Tcf signaling in skin carcinogenesis is discussed. (c) 2005 Wiley-Liss, Inc. PMID: 15765534 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 40: Mol Cancer Ther. 2005 Feb;4(2):233-41. Gene expression profiling identifies activating transcription factor 3 as a novel contributor to the proapoptotic effect of curcumin. Yan C, Jamaluddin MS, Aggarwal B, Myers J, Boyd DD. Department of Cancer Biology, University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, USA. The antitumor effect of curcumin (diferuloylmethane) is well established. However, there have been no unbiased studies to identify novel molecular targets of this compound. We therefore undertook a gene expression profiling study to identify novel targets of curcumin. A cDNA array comprised of 12,625 probes was used to compare total RNA extracted from curcumin-treated and untreated MDA-1986 cells for differential gene expression. We identified 202 up-regulated mRNAs and 505 transcripts decreased > or =2-fold. The proapoptotic activating transcription factor 3 (ATF3) was induced >4-fold. Two negative regulators of growth control [antagonizer of myc transcriptional activity (Mad) and p27kip1] were induced 68- and 3-fold, respectively. Additionally, two dual-activity phosphatases (CL 100 and MKP-5), which inactivate the c-jun-NH2-kinases, showed augmented expression, coinciding with reduced expression of the upstream activators of c-jun-NH2-kinase (MEKK and MKK4). Of the repressed genes, the expression of Frizzled-1 (Wnt receptor) was most strongly attenuated (8-fold). Additionally, two genes implicated in growth control (K-sam, encoding the keratinocyte growth factor receptor, and HER3) as well as the E2F-5 transcription factor, which regulates genes controlling cell proliferation, also showed down-regulated expression. Considering its role in apoptosis, we determined the contribution of ATF3 to the antitumor effect of curcumin. Curcumin-treated MDA-1986 cells showed a rapid, dose-dependent increase in ATF3/mRNA protein. Moreover, expression of an exogenous ATF3 cDNA synergized with curcumin in inducing apoptosis. Thus, we have identified several putative, novel molecular targets of curcumin and showed that one, (ATF3) contributes to the proapoptotic effects of this compound. PMID: 15713895 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 41: Clin Cancer Res. 2005 Jan 15;11(2 Pt 1):600-5. Epidermal growth factor receptor gene polymorphisms predict pelvic recurrence in patients with rectal cancer treated with chemoradiation. Zhang W, Park DJ, Lu B, Yang DY, Gordon M, Groshen S, Yun J, Press OA, Vallbohmer D, Rhodes K, Lenz HJ. Division of Medical Oncology and Department of Preventive Medicine, University of Southern California/Norris Comprehensive Cancer Center, Keck School of Medicine, Los Angeles, CA, USA. An association between epidermal growth factor receptor (EGFR) signaling pathway and response of cancer cells to ionizing radiation has been reported. Recently, a polymorphic variant in the EGFR gene that leads to an arginine-to-lysine substitution in the extracellular domain at codon 497 within subdomain IV of EGFR has been identified. The variant EGFR (HER-1 497K) may lead to attenuation in ligand binding, growth stimulation, tyrosine kinase activation, and induction of proto-oncogenes myc, fos, and jun. A (CA)(n) repeat polymorphism in intron 1 of the EGFR gene that alters EGFR expression in vitro and in vivo has also been described. In the current pilot study, we assessed both polymorphisms in 59 patients with locally advanced rectal cancer treated with adjuvant or neoadjuvant chemoradiation therapy using PCR-RFLP and a 5'-end [gamma-(33)P]ATP-labeled PCR protocol. We tested whether either polymorphism alone or in combination can be associated with local recurrence in the setting of chemoradiation treatment. We found that patients with HER-1 497 Arg/Arg genotype or lower number of CA repeats (both alleles <20) tended to have a higher risk of local recurrence (P = 0.24 and 0.31, respectively). Combined analysis showed the highest risk for local recurrence was seen in patients who possessed both a HER-1 497 Arg allele and <20 CA repeats (P = 0.05, log-rank test). Our data suggest that the HER-1 R497K and EGFR intron 1 (CA)(n) repeat polymorphisms may be potential indicators of radiosensitivity in patients with rectal cancer treated with chemoradiation. PMID: 15701846 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 42: J Virol. 2005 Feb;79(4):2274-86. Expression profiling of human hepatoma cells reveals global repression of genes involved in cell proliferation, growth, and apoptosis upon infection with parvovirus H-1. Li J, Werner E, Hergenhahn M, Poirey R, Luo Z, Rommelaere J, Jauniaux JC. Department of Physiology and Biophysics, Fudan University, Shanghai, People's Republic of China. Autonomous parvoviruses are characterized by their stringent dependency on host cell S phase and their cytopathic effects on neoplastic cells. To better understand the interactions between the virus and its host cell, we used oligonucleotide arrays that carry more than 19,000 unique human gene sequences to profile the gene expression of the human hepatocellular carcinoma cell line QGY-7703 at two time points after parvovirus H-1 infection. At the 6-h time point, a single gene was differentially expressed with a >2.5-fold change. At 12 h, 105 distinct genes were differentially expressed in virus-infected cells compared to mock-treated cells, with 93% of these genes being down-regulated. These repressed genes clustered mainly into classes involved in transcriptional regulation, signal transduction, immune and stress response, and apoptosis, as exemplified by genes encoding the transcription factors Myc, Jun, Fos, Ids, and CEBPs. Quantitative real-time reverse transcription-PCR analysis on selected genes validated the array data and allowed the changes in cellular gene expression to be correlated with the accumulation of viral transcripts and NS1 protein. Western blot analysis of several cellular proteins supported the array results and substantiated the evidence given by these and other data to suggest that the H-1 virus kills QGY-7703 cells by a nonapoptotic process. The promoter regions of most of the differentially expressed genes analyzed fail to harbor any motif for sequence-specific binding of NS1, suggesting that direct binding of NS1 to cellular promoters may not participate in the modulation of cellular gene expression in H-1 virus-infected cells. PMID: 15681429 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 43: Oncogene. 2005 Mar 3;24(10):1711-7. Latent membrane protein 1 of Epstein-Barr virus coordinately regulates proliferation with control of apoptosis. Dirmeier U, Hoffmann R, Kilger E, Schultheiss U, Briseno C, Gires O, Kieser A, Eick D, Sugden B, Hammerschmidt W. Department of Gene Vectors, GSF-National Research Center for Environment and Health, Marchioninistr. 25, Munich D-81377, Germany. Latent membrane protein 1 (LMP1), an oncoprotein encoded by Epstein-Barr virus (EBV), is an integral membrane protein, which acts like a constitutively active receptor. LMP1 is critical for some facet of EBV's induction and maintenance of proliferation of infected B cells. It, in part, mimics signaling by the CD40 receptor and has been implicated in regulating proliferation, survival, or both properties of EBV-infected cells. We established a conditional LMP1 allele in the context of the intact EBV genome to define the immediate-early cellular target genes regulated by LMP1 in order to assess its contributions to infected human B cells. The functional analysis of this conditional system indicated that LMP1 specifically induces mitogenic B-cell activation through c-myc and Jun/AP1 family members and confirms its direct role in upregulating expression of multiple genes with opposing activities involved in cell survival. LMP1's signals were found to be essential for the G1/S transition in human B cells; cells lacking LMP1's signals are cell cycle arrested and survive quiescently. LMP1's activities are therefore not required to maintain survival in nonproliferating cells. LMP1 does induce both pro- and antiapoptotic genes whose balance seems to permit survival during LMP1's induction and maintenance of proliferation. PMID: 15674340 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 44: Assay Drug Dev Technol. 2004 Dec;2(6):637-46. Direct measurement of multiple mRNAs in nerve growth factor-induced PC12 cells using electrophoretic tags to monitor biomarkers of neuronal differentiation in 96-well format. Roux P, Menguy I, Soubigou S, Chinn J, Ricard S, Williams S, Guitton JD, Tian T, Singh S, Grepin C. Aventis, Vitry Sur Seine, France. Pheochromocytoma-12 (PC12) cells recapitulate the program of neuronal differentiation by developing neurites after about 12 days of nerve growth factor (NGF) treatment. This model can be used to evaluate the neuroprotective/neurotrophic effect of compounds. Specific mRNAs such as cfos and c-jun are early biomarkers of the irreversible commitment into the differentiation program as they appear after only 30-40 min of NGF treatment. Monitoring the level of these mRNAs instead of the neurite outgrowth dramatically reduces the time needed to identify the drug potential of compounds. The electrophoretic tags, or eTag reporters (ACLARA Biosciences, Inc., Mountain View, CA), are a new class of fluorescent reporters that have unique migration properties in capillary electrophoresis, which allows for their separation and identification. (The eTag Multiplex Invader Assay and products incorporate Invader technology and Cleavase enzyme licensed for use from Third Wave Technologies, Inc. [Madison, WI] for multiplexed gene expression applications.) Each eTag molecule used begins as a phosphoramidite that is incorporated into a specific oligonucleotide using standard oligonucleotide synthesis procedures. A set of distinct probes labeled with different eTag molecules can then be mixed together to simultaneously quantify the levels of different mRNAs from the same sample. When compared to existing methods for measuring multiplexed gene expression from the same sample, the eTag assay allows a direct quantification of the mRNA from cells without any extraction/purification and still provides multiplexing capability, high sensitivity, miniaturization, and reproducibility compatible with medium-throughput screening methods. The eTag technology was used to simultaneously measure the level of expression of four mRNAs-c-fos, c-jun, c-myc, and gapdh-in NGF-treated PC12 cells in a standard 96-well format. The experimental data shown here demonstrate the use of eTag technology as a new screening tool, which uniquely combines robustness, sensitivity, multiplexing capability, and direct measurement of mRNA without any sample preparation steps, such as RNA extraction/purification or a reverse transcription step. PMID: 15674022 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 45: FEBS Lett. 2005 Jan 31;579(3):615-20. Differentiative pathway activated by 3-aminobenzamide, an inhibitor of PARP, in human osteosarcoma MG-63 cells. De Blasio A, Messina C, Santulli A, Mangano V, Di Leonardo E, D'Anneo A, Tesoriere G, Vento R. Dipartimento di Biologia Cellulare e dello Sviluppo, Sezione di Biochimica, Universita di Palermo, Policlinico, via del Vespro 129, 90127 Palermo, Italy. This study describes the molecular mechanism by which treatment with 3-AB, a potent inhibitor of PARP, allows human osteosarcoma MG-63 cells to restrict growth and enter differentiation. Our findings show that in MG-63 cells, aberrant gene expression keeps Rb protein constitutively inactivated through hyperphosphorylation and this promotes uncontrolled proliferation of the cells. After 3-AB-treatment, the poly(ADP-ribosyl)ation of nuclear proteins markedly decreases and this results in an increase in both the hypophosphorylated active form of Rb and pRb/E2F complexes. These effects are accompanied by G1 arrest, downregulation of gene products required for proliferation (cyclin D1, beta-catenin, c-Jun, c-Myc and Id2) and upregulation of those implicated in the osteoblastic differentiation (p21/Waf1, osteopontin, osteocalcin, type I collagen, N-cadherins and alkaline phosphatase). Our study suggests that use of PARP inhibitors may induce a remodeling of chromatin with the reprogramming of gene expression and the activation of differentiation. PMID: 15670817 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 46: Eur J Clin Invest. 2005 Feb;35(2):140-7. Molecular events associated with accelerated proliferative response in rat livers when partial hepatectomy is preceded by a sham operation. Laurent S, Starkel P, Leclercq IA, Lambotte L, Maiter D, Horsmans Y. Department of Gastroenterology, Universite Catholique de Louvain, 1200 Brussels, Belgium. BACKGROUND: When a sham operation is performed 6 h before partial hepatectomy (PH), the regenerative response is accelerated suggesting that sham operation itself contributes to cellular events leading to proliferation. MATERIALS AND METHODS: In order to examine the mechanisms implicated in this acceleration, we compared the activation of several factors associated with the progression through the cell cycle at various times after PH and after PH preceded by sham operation (S6 h + PH). The effect of a single sham (S) and two combined sham operations (S6 h + S) was also examined. Nonoperated rats were used as controls (C). RESULTS: The early factors NF-kappaB and Stat3 were activated after S6 h + PH and S6 h + S. C-jun expression was increased 0.5 h and 2 h after PH and 6 h after sham. There was no further increase in S6 h + PH and S6 h + S. In contrast, c-myc expression returned to baseline levels after S6 h and a new increase was observed 2 h after S6 h + PH but not after S6 h + S. P53 mRNA was significantly expressed 6 h after S6 h + PH, but at a level similar than that observed 6 and 12 h after PH alone. An earlier increase in c-Ha-ras mRNA and cyclin E protein was found in S6 h + PH, in comparison with PH alone. CONCLUSIONS: The first divergent response between the two combined models involved c-myc expression. However, major differences related to the accelerated liver regenerative response observed after S6 h + PH were found at late time points associating an earlier expression of c-Ha-ras and nuclear cyclin E. PMID: 15667586 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 47: Neurochem Res. 2004 Nov;29(11):2171-6. A model to induce low temperature trauma for in vitro astrogliosis study. Yu AC, Wu BY, Liu RY, Li Q, Li YX, Wong PF, Liu S, Lau LT, Fung YW. Key Laboratory of Neuroscience, Neuroscience Research Institute, Peking University, Beijing, China. achy@dnachip.com.hk Astrogliosis is an inevitable and rapid response of astrocytes to physical, chemical and pathological injuries. To study astrogliosis, we developed a reproducible in vitro model in which low temperature injury to cultured astrocytes could be induced by placing the culture dish onto a copper pipe pre-cooled by liquid nitrogen. Using this model, the relationship between the temperature decline and the severity of cellular damage was analyzed. An increase in the expression of some known injury-related proteins, such as glial fibrillary acidic protein (GFAP), immediate early response genes (IEGs), and heat shock proteins 70 (HSP70), was demonstrated in astrocytes after low temperature trauma. With the use of this low temperature trauma model, the flexibility in the temperature control and injury area may allow researchers to evaluate cryotherapy and cryosurgery, which could be applicable to future development of quality health care. PMID: 15662852 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 48: Anticancer Drugs. 2005 Feb;16(2):175-83. Compound 278E, structurally modified from tanshinone, induces monocytic differentiation and regulates proto-oncogene expression in human leukemic HL-60 cells. Liao HF, Shyu SY, Kuo YH, Yang YC, Chen YJ. Department of Medical Research, Mackay Memorial Hospital, Taipei, Taiwan. Tanshinone derivative compounds, isolated from Salvia miltiorrhiza Bunge (Labiatae), have been reported as microtubule inhibitors with antimitotic activity. In this study, we examined the growth-inhibiting and differentiation-inducing effect of these compounds on human leukemic HL-60 cells. The expression of protein kinase C (PKC) and proto-oncogenes in 278E-treated cells was also assessed. All tanshinone derivative compounds exhibited growth-inhibitory effects on HL-60 cells, but only 278E induced cell differentiation. Morphological observation of 278E-treated HL-60 cells showed a greater percentage of monocytes and macrophages (Mo/Mphi). Treatment with 5 microg/ml 278E resulted in a marked increase in the percentages of superoxide-producing (up to 95.5+/-1.8%) and non-specific esterase-positive cells (up to 80.3+/-9.1%). The differentiated cells also expressed cell surface antigens characteristic of Mo/Mphi, including CD11b, CD14 and CD68. Neither cellular changes in isozymes of PKC nor translocation of these isozymes from cytosol to cell membrane were seen in 278E-treated HL-60 cells. 278E caused a downregulation of c-myc as well as an up-regulation of c-fms, c-jun and c-fos. PMID: 15655415 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 49: Int J Mol Med. 2005 Feb;15(2):269-75. Functional analysis of the effect of forced activation of STAT3 on M1 mouse leukemia cells. Yoshida T, Iwamoto T, Adachi K, Yokota T, Miyake Y, Hamaguchi M. Department of Ophthalmology, Nagoya University School of Medicine, Showa-ku, Nagoya 466-8550, Japan. M1 mouse myeloid leukemia cells exhibit growth arrest and differentiation to monocytes/macrophages in response to leukemia inhibitory factor (LIF) stimulation. Although recent studies have demonstrated that STAT3 plays a central role in this process, it is unknown whether STAT3 activation alone is sufficient. To address this issue, we have established M1/STAT3ER cells, where STAT3 is selectively activated by 4-hydroxytamoxifen (4HT). 4HT stimulation did not have any effect on growth and morphology of M1/ STAT3ER cells, and did not induce the down-regulation of mRNA of c-myc and c-myb, which is necessary for M1 cell differentiation. On the other hand, mRNA of jun-B, IRF1 and p19 was increased by 4HT. DNA precipitation assay indicated that both stimulation of LIF and 4HT similarly activated STAT3ER. Introduction of a constitutive active MAP kinase kinase (MEK1) into M1/STAT3ER cells did not induce differentiation either. Together, our present data suggest that signaling other than the activation of STAT3 and MEK1 may be necessary for M1 cell-growth arrest and differentiation, while a set of early genes of LIF are induced by only STAT3 activation. PMID: 15647843 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 50: Cell Cycle. 2005 Jan;4(1):87-95. Epub 2005 Jan 10. HER2-targeting antibodies modulate the cyclin-dependent kinase inhibitor p27Kip1 via multiple signaling pathways. Le XF, Pruefer F, Bast RC Jr. Department of Experimental Therapeutics, Division of Cancer Medicine, University of Texas M.D. Anderson Cancer Center, Houston, Texas, USA. Anti-HER2 antibody trastuzumab is emerging as a frontline therapy for patients with metastatic breast cancers that overexpress HER2. Understanding the molecular mechanisms by which the antibody inhibits tumor growth should permit the design of even more effective trastuzumab-based protocols. Several groups including our own have demonstrated that induction of cyclin-dependent kinase (CDK) inhibitor p27Kip1 protein is one of the key mechanisms of action of HER2-targeting antibodies. In this review, we discuss currently available data regarding the multiple signaling targets and pathways by which HER2-targeting antibodies upregulate p27Kip1 protein in breast cancer cells that overexpress HER2. Anti-HER2 antibodies inhibit HER2-mediated signaling in cancer cells, ultimately upregulating the levels and activity of p27Kip1 protein. At least six signaling targets and pathways are modulated by trastuzumab. By inhibiting CDK2 and decreasing Thr187 phosphorylation of p27Kip1, trastuzumab abrogates targeting of SCF-ubiquitin E3 ligase and minimizes proteasome degradation of p27Kip1. By inhibiting AKT and human kinase interacting stathmin (hKIS), trastuzumab blocks Thr157-, Thr198- and Ser10-induced p27Kip1 translocation from the nucleus to the cytosol, which increases the inhibitory effect of p27Kip1. By inhibiting Jun activation domain-binding protein 1 (Jab1) trastuzumab increases nuclear retention of p27Kip1. By inhibiting cyclin D and c-Myc, trastuzumab releases the sequestrated p27bKip1 protein from cyclin D-CDK4/6 complexes and increase the effect of p27Kip1 on CDK2-cyclin E complexes. By stimulating minibrain related kinase (MIRK), trastuzumab stabilizes p27Kip1 in the nucleus, which increases inhibitory action of p27Kip1 on CDK2. The targets and pathways affected by trastuzumab work in concert to maximize the expression and inhibitory effect of p27Kip1, which leads to cell cycle G1 arrest and growth inhibition. PMID: 15611642 [PubMed - in process] --------------------------------------------------------------- 51: BMC Genomics. 2004 Dec 21;5(1):97. The association of Alu repeats with the generation of potential AU-rich elements (ARE) at 3' untranslated regions. An HJ, Lee D, Lee KH, Bhak J. BioSystems Dept,, Korea Advanced Institute of Science and Technology, Yuseong-gu, Daejeon 305-701, Republic of Korea. hjan@bisl.kaist.ac.kr BACKGROUND: A significant portion (about 8% in the human genome) of mammalian mRNA sequences contains AU (Adenine and Uracil) rich elements or AREs at their 3' untranslated regions (UTR). These mRNA sequences are usually stable. However, an increasing number of observations have been made of unstable species, possibly depending on certain elements such as Alu repeats. ARE motifs are repeats of the tetramer AUUU and a monomer A at the end of the repeats ((AUUU)nA). The importance of AREs in biology is that they make certain mRNA unstable. Proto-oncogene, such as c-fos, c-myc, and c-jun in humans, are associated with AREs. Although it has been known that the increased number of ARE motifs caused the decrease of the half-life of mRNA containing ARE repeats, the exact mechanism is as of yet unknown. We analyzed the occurrences of AREs and Alu and propose a possible mechanism for how human mRNA could acquire and keep AREs at its 3' UTR originating from Alu repeats. RESULTS: Interspersed in the human genome, Alu repeats occupy 5% of the 3' UTR of mRNA sequences. Alu has poly-adenine (poly-A) regions at its end, which lead to poly-thymine (poly-T) regions at the end of its complementary Alu. It has been found that AREs are present at the poly-T regions. From the 3' UTR of the NCBI's reference mRNA sequence database, we found nearly 40% (38.5%) of ARE (Class I) were associated with Alu sequences (Table 1) within one mismatch allowance in ARE sequences. Other ARE classes had statistically significant associations as well. This is far from a random occurrence given their limited quantity. At each ARE class, random distribution was simulated 1,000 times, and it was shown that there is a special relationship between ARE patterns and the Alu repeats. CONCLUSION: AREs are mediating sequence elements affecting the stabilization or degradation of mRNA at the 3' untranslated regions. However, AREs' mechanism and origins are unknown. We report that Alu is a source of ARE. We found that half of the longest AREs were derived from the poly-T regions of the complementary Alu. PMID: 15610565 [PubMed - in process] --------------------------------------------------------------- 52: Mol Cell Biol. 2005 Jan;25(1):220-32. Ras-Raf-Arf signaling critically depends on the Dmp1 transcription factor. Sreeramaneni R, Chaudhry A, McMahon M, Sherr CJ, Inoue K. Department of Pathology, Wake Forest University Health Sciences, 2102 Gray Building, Medical Center Blvd., Winston-Salem, NC 27157, USA. Dmp1 prevents tumor formation by activating the Arf-p53 pathway. In cultured primary cells, the Dmp1 promoter was efficiently activated by oncogenic Ha-Ras(V12), but not by overexpressed c-Myc or E2F-1. Dmp1 promoter activation by Ras(V12) depended on Raf-MEK-ERK signaling. Induction of p19(Arf) and p21(Cip1) by oncogenic Raf was compromised in Dmp1-null cells, which were resistant to Raf-mediated premature senescence. A Ras(V12)-responsive element was mapped to the 5' leader sequence of the murine Dmp1 promoter, where endogenous Fos and Jun family proteins bind. Dmp1 promoter activation by Ras(V12) was strikingly impaired in c-Jun as well as in JunB knock-down cells, suggesting the critical role of Jun proteins in the activation of the Dmp1 promoter. A Ras(V12)-responsive element was mapped to the unique Dmp1/Ets site on the Arf promoter, where endogenous Dmp1 proteins bind upon oncogenic Raf activation. Therefore, activation of Arf by Ras/Raf signaling is indirectly mediated by Dmp1, explaining why Dmp1-null primary cells are highly susceptible to Ras-induced transformation. Our data indicate the presence of the novel Jun-Dmp1 pathway that directly links oncogenic Ras-Raf signaling and p19(Arf), independent of the classical cyclin D1/Cdk4-Rb-E2F pathway. PMID: 15601844 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 53: Cancer Chemother Pharmacol. 2005 Mar;55(3):286-94. Epub 2004 Nov 16. DNA damage, c-myc suppression and apoptosis induced by the novel topoisomerase II inhibitor, salvicine, in human breast cancer MCF-7 cells. Lu HR, Meng LH, Huang M, Zhu H, Miao ZH, Ding J. Division of Anti-tumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 201203, People's Republic of China. Salvicine, a diterpenoid quinone compound, possesses potent in vitro and in vivo antitumor activity. Salvicine is a novel non-intercalative topoisomerase II poison. In this study salvicine induced evident DNA damage, which was further characterized as double-strand breaks mainly in MCF-7 human breast cancer cells. The degree of damage was highly correlated with growth inhibition of MCF-7. Using a PCR-stop assay we demonstrated that this damage was selective. Preferential damage occurred in the p2 promoter region, but not the 3'-end of the protooncogene c-myc. The expression of oncogenes, such as c-myc and c-jun, was additionally investigated. Salvicine induced a dose-dependent decrease in c-myc gene transcription, concomitant with an increase in c-jun expression. Furthermore, reverse-transcription PCR and Western blotting data revealed that salvicine failed to stimulate the mRNA and protein levels of p53 and its downstream targets p21 and bax. The phosphorylation degree of serine 15 of p53, which is thought to be an active form of p53 in response to cellular DNA damage, remained in a steady state. In view of these results, we propose that the downregulation of c-myc resulting from selective damage plays a role in apoptosis signaling. Moreover, salvicine-induced apoptosis in MCF-7 subsequent to DNA damage seems to be mediated through a p53-independent pathway. PMID: 15592835 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 54: J Biol Chem. 2005 Feb 11;280(6):4117-24. Epub 2004 Dec 7. Glucocorticoid receptor-induced MAPK phosphatase-1 (MPK-1) expression inhibits paclitaxel-associated MAPK activation and contributes to breast cancer cell survival. Wu W, Pew T, Zou M, Pang D, Conzen SD. Department of Medicine and the Committee on Cancer Biology, University of Chicago, Chicago, Illinois 60637, USA. Glucocorticoid receptor (GR) activation has recently been shown to inhibit apoptosis in breast epithelial cells. We have previously described a group of genes that is rapidly up-regulated in these cells following dexamethasone (Dex) treatment. In an effort to dissect the mechanisms of GR-mediated breast epithelial cell survival, we now examine the molecular events downstream of GR activation. Here we show that GR activation leads to both the rapid induction of MAPK phosphatase-1 (MKP-1) mRNA and its sustained expression. Induction of the MKP-1 protein in the MCF10A-Myc and MDA-MB-231 breast epithelial cell lines was also seen. Paclitaxel treatment resulted in MAPK activation and apoptosis of MDA-MB-231 breast cancer cells, and both processes were inhibited by Dex pretreatment. Furthermore, induction of MKP-1 correlated with the inhibition of extracellular signal-regulated kinase (ERK1/2) and c-Jun N-terminal kinase (JNK) activity, whereas p38 activity was minimally affected. Blocking Dex-induced MKP-1 induction using small interfering RNA increased ERK1/2 and JNK phosphorylation and decreased cell survival. ERK1/2 and JNK inactivation was associated with Ets-like transcription factor-1 (ELK-1) dephosphorylation. To explore the gene expression changes that occur downstream of ELK-1 dephosphorylation, we used a combination of temporal gene expression data and promoter element analyses. This approach revealed a previously unrecognized transcriptional target of ELK-1, the human tissue plasminogen activator (tPA). We verified the predicted ELK-1--> tPA transcriptional regulatory relationship using a luciferase reporter assay. We conclude that GR-mediated MAPK inactivation contributes to cell survival and that the potential transcriptional targets of this inhibition can be identified from large scale gene array analysis. PMID: 15590693 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 55: West Afr J Med. 2004 Jul-Sep;23(3):187-90. Oncogene expression in the peri-articular osteophytes. Alonge TO, Rooney P, Oni OO. Department of Surgery, College of Medicine, University of Ibadan, University College Hospital, Ibadan, Nigeria. alonget@skannet.com OBJECTIVE: The aim of this study was to ascertain the proliferative and probably reparative potentials of the periarticular osteophytes by evaluating the sites of expression of c-myc, c-jun and c-fos oncogenes in this neoplastic repair tissue. MATERIALS AND METHODS: Sections of osteophytes were obtained from knees of patients undergoing total knee arthroplasty for osteoarthritis. Decalcified sections of osteophytes were stained for c-myc and c-jun oncogenes using the avidin HRP technique. Sections of breast carcinoma were used as positive controls. Undecalcified or frozen sections of osteophytes were stained for c-fos oncogene using the avidin alkaline phosphatase technique. Sections of the human skin were used as positive control. For both techniques, sections of normal articular cartilage were used as negative controls. RESULTS: The chondrocytes of the entire cartilage mantle of the peri-articular osteophyte had positive staining for c-myc oncogene but no staining for c-jun oncogene. The basal chondrocytes of the deep layer of the cartilage mantle of the peri-articular osteophyte had positive staining for c-fos oncogene. The normal articular cartilage sections had no staining for any of the oncogenes evaluated. CONCLUSION: The expression of c-myc oncogene in the osteophytic chondrocytes suggests that these cells are actively proliferating. However, c-fos expression in the basal chondrocytes implies that these cells are capable of transformation. This result confirms the proliferative ability of the peri-articular osteophytes and this may suggest that this osteochondral repair tissue, which is apparently wrongly sited, may be a source of tissue for osteochondral grafting for full thickness articular cartilage defects. PMID: 15587825 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 56: Int J Cancer. 2005 Apr 10;114(3):346-55. Cadmium-induced malignant transformation in rat liver cells: role of aberrant oncogene expression and minimal role of oxidative stress. Qu W, Diwan BA, Reece JM, Bortner CD, Pi J, Liu J, Waalkes MP. Inorganic Carcinogenesis Section, Laboratory of Comparative Carcinogenesis, National Cancer Institute at the National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA. Our study examined the role of oxidative stress and aberrant gene expression in malignant transformation induced by chronic, low-level cadmium exposure in non-tumorigenic rat liver epithelial cell line, TRL 1215. Cells were cultured in 1.0 microM cadmium (as CdCl(2)) for up to 28 weeks and compared to passage-matched control cells. The level of cadmium used for transformation produced no evidence of increased superoxide (O(2) (-*.)) or hydrogen peroxide (H(2)O(2)) levels in the early stages of exposure (0.05) in the levels of expression ratio at both the transcriptional and translational levels at 30 min for c-fos and c-jun and at 180 min for c-myc after serum stimulation. In addition, exposure of T98G cells to linearly (vertical and horizontal) and/or circularly polarized MFs (500 microTr) produced no significant change (P>0.05) in the expression ratio at both transcriptional and post-transcriptional levels. Thus, there was no evidence that linearly or rotating polarized MFs enhanced early response gene expression in these studies. These results suggest that environmental MFs at 1-500 microT flux density are unlikely to induce carcinogenesis through a mechanism involving altered expression of the immediate early response genes. Copyright 2002 Wiley-Liss, Inc. PMID: 11835255 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 162: J Biol Chem. 2002 Apr 12;277(15):13059-66. Epub 2002 Jan 30. Identification of Myc-mediated death response pathways by microarray analysis. Yu Q, He M, Lee NH, Liu ET. Advanced Technology Center, Center for Cancer Research, NCI/National Institutes of Health, Gaithersburg, MD 20877, USA. To understand the mechanisms of Myc-mediated apoptosis induced by DNA damage, we have characterized the death kinetics of three Rat-1 fibroblast cell lines that either overexpress Myc or lack Myc and their parental wild-type cells following exposure to the DNA-damaging agent VP-16, and we monitored the changes in gene expression using microarray. We have identified three groups of genes whose expressions are distinctly regulated during this process. One cluster (Cluster A) revealed a VP-16-dependent but Myc-independent induction of a set of genes that is not linked to the apoptotic response. Two other gene clusters, however, were associated with VP-16-induced apoptosis. Cluster B, which includes p53-responsive genes, was associated with the temporal onset of apoptosis but accounted for only the basal apoptosis. However, Cluster C, which includes c-jun, was highly regulated by Myc and appeared to be critical to mounting the maximal apoptotic response in Myc-expressing cells. Furthermore, the Myc level dropped sharply following VP-16 exposure, which varied inversely with the induction of Cluster C genes, suggesting Myc normally represses their transcription. Thus, we have proposed that removal of Myc-mediated repression of apoptotic signals, combined with Myc-associated acceleration of the p53 responsive pathway, results in complete and rapid cell death following DNA damage. PMID: 11821411 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 163: Folia Histochem Cytobiol. 2001;39 Suppl 2:93-5. The expression of oncogenes in tumour tissues of breast carcinoma patients. Rzymowska J, Grzybowska-Szatkowska L. Department of Human Genetics, Medical University, Lublin, Poland. jolarzym@interia.pl The balance between Bcl-2 and c-Myc and c-Jun seems to be an important determinant of cellular sensitivity to the induction of apoptosis. High expression of Bcl-2 was noticed to be strongly related to low rates of apoptotic cell death. The mean value of the apoptotic index was 45.0% in Bcl-2-negative tumours and 7.5% in Bcl-2-positive tumours. C-Myc and c-Jun accumulation were associated with the absence of Bcl-2 expression and with increased apoptotic activity. The loss of Bd-2 expression was strongly correlated with increased apoptotic cell death. The inverse correlation is between apoptotic and mitotic index. A high mitotic index exists in most patients with a low apoptotic index. Bcl-2, c-Myc and c-Jun does not only take part in cell death, but also in cell division in breast carcinoma cells in which the regulation of cell division and cell death are strictly connected. PMID: 11820644 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 164: Heart Fail Rev. 2002 Jan;7(1):63-9. Acute and chronic adaptation to hemodynamic overload and ischemia in the aged heart. Isoyama S, Nitta-Komatsubara Y. Tohoku Bunka Gakuen University, Faculty of Medical Science and Welfare, 6-45-16 Kunimi, Aoba-ku, Sendai 981-8551, Japan. shogen@rehab.tbgu.ac.jp Congestive heart failure occurs more frequently in older individuals. This higher incidence of heart failure may be caused by the diminished capacity of aged hearts to adapt to increased hemodynamic overload and ischemia which are the most important triggers for heart failure in the aged. In the immediate early phase after the imposition of ascending aortic banding, the mRNA expression of proto-oncogenes (c-fos, c-jun and c-myc) was diminished in aged rat hearts compared with young adult hearts. In the later phase, the pattern of expression of contractile protein genes in aged hearts differed quantitatively from that in adult hearts. The hypertrophic responses to the imposition of not only pressure-overload but also volume-overload were also diminished at the organ and cellular levels. In addition, this diminution was observed both in the left and right ventricles. Against ischemic insults, aged hearts responded with a diminished expression of proto-oncogenes and heat shock proteins. Thus, aged hearts are characterized by poor adaptation to hemodynamic overload and by a poor self-protective mechanism against cell death through necrosis and apoptosis. Of interest, more severe hemodynamic overload elevated the diminished responses to a level similar to that in adult hearts, suggesting that the threshold for the heart to respond to hemodynamic overload or ischemia is elevated in aged hearts. In addition, even in aged hearts ischemic preconditioning upregulated the diminished gene expression in a gene-dependent manner. Thus, the capacity for adaptation to hemodynamic overload and ischemia is diminished in aged hearts, but aged hearts preserve the ability to respond to these under some conditions. Publication Types: Review Review, Tutorial PMID: 11790923 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 165: Int J Oncol. 2002 Feb;20(2):391-5. Expression profile of proteins involved in the xenotransplantability of non-small cell lung cancers into athymic nude mice. Efferth T, Koomagi R, Mattern J, Volm M. Virtual Campus Rhineland-Palatinate, 55033 Mainz, Germany. efferth@vcrp.de The aim of this study was to evaluate the expression profile of proteins involved in growing of human non-small cell lung cancer (NSCLC) in athymic nude mice. The expressions of 20 gene products in primary NSCLC of 170 patients were analyzed and the proteins were correlated with the transplantability of the carcinomas in nude mice. There was no relationship between xenotransplantability of human non-small cell lung cancer in nude mice and histology, stage or lymph node involvement. Of the analyzed proliferative factors PCNA, cyclin A, cyclin D, cdk2, cdk4 and cell cycle phases only cyclin D, cdk4 and the cell cycle phases were up-regulated in growing carcinomas. There was also a correlation between the apoptotic indices and the take rate in nude mice. Concerning microvessel density and angiogenic factors only VEGF showed a relation to xenotransplantability. Of the proto-oncogenes and suppressor gene products N-RAS, P53, FOS and JUN revealed a relationship to the take rate of NSCLC, while such a relationship was not found with MYC, ERBB-1 and ERBB-2. In a second step, a hierarchical cluster analysis was carried out. The resulting clusters were correlated with the take rate of the carcinomas in nude mice. The expression of JUN, N-RAS, FOS, cyclin D, and cdk4 were significantly different in both groups with non- overlapping confidence intervals. Thus, the up-regulation of the proteins JUN, N-RAS, FOS, cyclin D and cdk4 predicts the growth of NSCLC in nude mice. PMID: 11788907 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 166: Zhonghua Yu Fang Yi Xue Za Zhi. 2001 Sep;35(5):305-8. [Effects of selenium on hepatocellular protooncogene c-myc, c-fos and c-jun expression induced by cadmium in rats] [Article in Chinese] Yu R, Chen X. Department of Environmental Health, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China. OBJECTIVES: This study was conducted to explore effects of selenium on hepatocellular protooncogene c-myc, c-fos and c-jun expression induced by cadmium in rats. METHOD: Both cadmium and selenium were given to rats by i.p. and there were 5 SD rats in each group. Protooncogene c-myc, c-fos and c-jun expression in rat liver cells was measured with Northern Dot Hybridization. RESULTS: The results showed that cadmium chloride at doses of 5, 10 or 20 mumol/kg, significantly induced proto-oncogene c-myc, c-fos and c-jun expression, and when sodium selenite at the dose of 5 mumol/kg was given at the time, the effect of cadmium chloride on hepatocellular protooncogene c-myc, c-fos and c-jun expression was inhibited. CONCLUSION: Selenium at certain doses could inhibit hepatocellular protooncogene c-myc, c-fos and c-jun expression induced by cadmium in rats. Publication Types: News PMID: 11769627 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 167: Ann Oncol. 2001;12 Suppl 2:S13-7. Oncogenes, growth factors, receptor expression and proliferation markers in digestive neuroendocrine tumours. A critical reappraisal. Delle Fave G, Corleto VD. Department of Gastrointestinal and Liver Diseases, II School of Medicine, La Sapienza University, Rome, Italy. dddhgi@tin.it BACKGROUND: The main characteristic of the digestive neuroendocrine tumours (dNETs) is the low proliferating activity, even in the presence of malignant, metastatic behavior. PATIENTS AND METHODS: Considering that dNETs are rare diseases, relatively numerous studies, often including a conspicuous number of patients, have recently investigated the molecular mechanisms of neuroendocrine tumour genesis. RESULTS: In contrast to non-endocrine tumours of the digestive system such as carcinoma of the pancreas, colon and stomach, dNETs do not show alterations in oncogenes (ras, Myc, fos jun, Src) or in common tumor suppressor genes [p53, retinoblastoma suspectibility gene (Rb)]. MEN-1 gene alterations will likely be important in a proportion of sporadic dNETs. The role of various growth factors, novel oncogenes and tumour suppressor genes have also been investigated. However, results from these studies are non-conclusive and to date the molecular pathogenesis of these tumours has not been clarified. Studies on somatostatin receptor expression and synthetic analogues, as growth inhibitors in dNETs, although promising, have not reproduced in vivo all the antiproliferative effects showed in in vitro models. CONCLUSION: Although various functional genes and molecular mechanisms have been investigated in dNETs, to date the molecular pathogenesis of these tumours remains to be elucidated. Publication Types: Review Review, Tutorial PMID: 11762339 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 168: Oncogene. 2001 Nov 8;20(51):7524-35. TOJ3, a target of the v-Jun transcription factor, encodes a protein with transforming activity related to human microspherule protein 1 (MCRS1). Bader AG, Schneider ML, Bister K, Hartl M. Institute of Biochemistry, University of Innsbruck, Peter-Mayr-Str. 1a, A-6020 Innsbruck, Austria. Using the established quail cell line Q/d3 conditionally transformed by the v-jun oncogene, cDNA clones (TOJ2, TOJ3, TOJ5, TOJ6) were isolated by representational difference analysis (RDA) that correspond to genes which were induced immediately upon conditional activation of v-jun. One of these genes, TOJ3, is immediately and specifically activated after doxycycline-mediated v-jun induction, with kinetics similar to the induction of well characterized direct AP-1 target genes. TOJ3 is neither activated upon conditional activation of v-myc, nor in cells or cell lines non-conditionally transformed by oncogenes other than v-jun. Sequence analysis revealed that the TOJ3-specific cDNA encodes a 530-amino acid protein with significant sequence similarities to the murine or human microspherule protein 1 (MCRS1, MSP58), a nucleolar protein that directly interacts with the ICP22 regulatory protein from herpes simplex virus 1 or with p120, a proliferation-related protein expressed at high levels in most human malignant tumor cells. Similar to its mammalian counterparts, the TOJ3 protein contains a bipartite nuclear localization motif and a forkhead associated domain (FHA). Using polyclonal antibodies directed against a recombinant amino-terminal TOJ3 protein segment, the activation of TOJ3 in jun-transformed fibroblasts was also demonstrated at the protein level by specific detection of a polypeptide with an apparent M(r) of 65 000. Retroviral expression of the TOJ3 gene in quail or chicken embryo fibroblasts induces anchorage-independent growth, indicating that the immediate activation of TOJ3 in fibroblasts transformed by the v-jun oncogene contributes to cell transformation. PMID: 11709724 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 169: Int J Oncol. 2002 Jan;20(1):77-9. Human chorionic gonadotropin and inhibin induce histone acetylation in human breast epithelial cells. Jiang X, Russo IH, Russo J. Breast Cancer Research Laboratory, Fox Chase Cancer Center, Philadelphia, PA 19111, USA. We have previously demonstrated that human chorionic gonadotropin or hCG-induces differentiation of the mammary gland. The effect of hCG was accompanied by the synthesis of inhibin, a heterodimeric protein that is structurally related to the transforming growth factor-beta (TGF-beta) family and activation of c-myc, c-jun, testosterone repressed prostate message 2 (TRPM2) and interleukin-l-beta-converting enzyme (ICE) transcripts. In the present work we aim to demonstrate that hCG and inhibin may control the transcription of these genes by acetylation of histones in the breast epithelial cells. For this purpose we have utilized MCF-10F cells in culture to detect the levels of acetylated histone H3 and H4 after treatment with different concentrations of hCG as well as inhibin beta-subunit at various time points. In the Western blot analysis, both acetylated histone H3 and H4 were significantly increased by hCG treatment over 12 h in MCF-10F cells at all the doses tested. Inhibin induced the accumulation of acetylated histone H3 after 4-h-treatment at the concentration of 1 ng/ml and at all the time points with higher concentration (10-1000 ng/ml). Slight induction of acetylated histone H4 was detected only in the cells treated with inhibin at 100 ng/ml for 12 and 24 h. This study is the first one to demonstrate that these hormones increase the acetylation of histones. PMID: 11743645 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 170: Mol Cell Biol. 2002 Jan;22(1):257-69. v-Src-induced modulation of the calpain-calpastatin proteolytic system regulates transformation. Carragher NO, Westhoff MA, Riley D, Potter DA, Dutt P, Elce JS, Greer PA, Frame MC. The Beatson Institute for Cancer Research, Cancer Research Campaign Beatson Laboratories, Glasgow, Scotland, United Kingdom. n.carragher@beatson.gla.ac.uk v-Src-induced oncogenic transformation is characterized by alterations in cell morphology, adhesion, motility, survival, and proliferation. To further elucidate some of the signaling pathways downstream of v-Src that are responsible for the transformed cell phenotype, we have investigated the role that the calpain-calpastatin proteolytic system plays during oncogenic transformation induced by v-Src. We recently reported that v-Src-induced transformation of chicken embryo fibroblasts is accompanied by calpain-mediated proteolytic cleavage of the focal adhesion kinase (FAK) and disassembly of the focal adhesion complex. In this study we have characterized a positive feedback loop whereby activation of v-Src increases protein synthesis of calpain II, resulting in degradation of its endogenous inhibitor calpastatin. Reconstitution of calpastatin levels by overexpression of exogenous calpastatin suppresses proteolytic cleavage of FAK, morphological transformation, and anchorage-independent growth. Furthermore, calpastatin overexpression represses progression of v-Src-transformed cells through the G(1) stage of the cell cycle, which correlates with decreased pRb phosphorylation and decreased levels of cyclins A and D and cyclin-dependent kinase 2. Calpain 4 knockout fibroblasts also exhibit impaired v-Src-induced morphological transformation and anchorage-independent growth. Thus, modulation of the calpain-calpastatin proteolytic system plays an important role in focal adhesion disassembly, morphological transformation, and cell cycle progression during v-Src-induced cell transformation. PMID: 11739739 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 171: Toxicol Lett. 2002 Jan 5;126(1):69-80. Transient induction of metallothionein isoform 3 (MT-3), c-fos, c-jun and c-myc in human proximal tubule cells exposed to cadmium. Garrett SH, Phillips V, Somji S, Sens MA, Dutta R, Park S, Kim D, Sens DA. Program in Genetics and Developmental Biology, Department of Urology, Robert C. Byrd Health Sciences Center, West Virginia University, PO Box 9251, Morgantown, WV 26506-9251, USA. Cadmium (Cd(+2)) has been shown to transiently increase the expression of mRNA for the third isoform of the metallothionein (MT-3) gene family in cultured human proximal tubule (HPT) cells. The goal of the present study was to further define the expression of MT-3 in mortal (HPT) and immortal (HK-2) cultures of HPT cells when exposed to lethal and sub-lethal concentrations of Cd(+2) under both acute and chronic time periods of exposure. Expression of MT-3 mRNA and protein was determined in cultured HPT cells and HK-2 cells using reverse-transcription-polymerase chain reaction (RT-PCR) and immuno-blotting, and expression of c-fos, c-jun and c-myc mRNA by RT-PCR. The results confirmed that exposure of the HPT cells to Cd(+2) induced a transient increase in MT-3 mRNA and extended the induction to include a subsequent transient increase in the level of the MT-3 protein. The induction of MT-3 was rapid and returned to control values within 48 h of exposure despite the continued presence of lethal and sublethal concentrations of Cd(+2). It was also demonstrated that the pattern of expression of MT-3 mRNA was similar to that of the early response genes, c-fos, c-jun and c-myc. It was shown that the HK-2 cells did not express MT-3 when exposed to Cd(+2), but had similar expression of the c-fos, c-jun and c-myc genes. The results demonstrate that MT-3 expression is metal responsive in HPT cells. PMID: 11738272 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 172: Biochem Biophys Res Commun. 2001 Nov 30;289(2):414-20. Isoform-dependent interaction of BRDG1 with Tec kinase. Yokohari K, Yamashita Y, Okada S, Ohya K, Oda S, Hatano M, Mano H, Hirasawa H, Tokuhisa T. Department of Developmental Genetics, Chiba University Graduate School of Medicine, Inohana 1-8-1, Chuo-ku, Chiba 260-8670, Japan. Tec is the prototype of an emerging family of protein-tyrosine kinases. Tec and Btk, another member of this family, together participate in the development of B-cell immune system. We previously identified one of the downstream messengers for human Tec kinase, BRDG1. BRDG1 is associated with Tec and becomes tyrosine-phosphorylated in B-cells by the engagement of B-cell antigen receptor (BCR). Here we show that overexpression of BRDG1 strongly augments BCR-mediated activation of cAMP-response element binding protein (CREB) but not that of c-Jun and the promoters of c-MYC and BCL-xL genes. Furthermore, we isolated the murine orthologue of BRDG1. Three isoforms of BRDG1 are generated by alternative splicing of the message. Two of them have a deletion of 33 amino acids in a Pleckstrin homology (PH) domain of BRDG1. Both the tyrosine-phosphorylation and CREB-activating ability of BRDG1 were isoform-dependent, suggesting a role of the PH domain of BRDG1. These data have identified a novel regulatory mechanism of CREB family of transcriptional factors. PMID: 11716489 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 173: J Biol Chem. 2002 Feb 1;277(5):3576-84. Epub 2001 Nov 19. Inducible expression of a constitutively active mutant of mitogen-activated protein kinase kinase 7 specifically activates c-JUN NH2-terminal protein kinase, alters expression of at least nine genes, and inhibits cell proliferation. Wolter S, Mushinski JF, Saboori AM, Resch K, Kracht M. Institute of Pharmacology, Medical School Hannover, Carl-Neuberg Strasse 1, D-30625 Hannover, Germany. MKK7 is a recently discovered mitogen-activated protein kinase (MAPK) kinase that is unique in that it specifically activates only the c-JUN NH(2)-terminal protein kinase (JNK) family of enzymes. Very little is known about the biological role of MKK7. We generated inducible cell lines from the human embryonal kidney carcinoma cell line, HEK293, by stable transfection with a constitutively active mutant of MKK7, MKK7(3E), fused to green fluorescent protein (GFP), under the control of an ecdysone-inducible promoter. Treatment of cells with the synthetic ecdysone analog ponasterone A induced expression of GFP-MKK7(3E) and resulted in sustained activation of endogenous JNK, but neither of the other endogenous MAPKs, ERK or p38. Red and green fluorescing cDNA copies of mRNA extracted from cells obtained before and after induction of GFP-MKK7(3E) were hybridized to microarrays containing more than 6,000 cDNAs in eight independent experiments. By selection criteria, 23 genes were differentially regulated after 24 h of induction of GFP-MKK7(3E) and 16 after 48 h. The expression of 9 genes was consistently changed after both 24 and 48 h of induction. These changes included down-regulation of three genes, c-myc, angiopoietin-2, and glucose-regulated protein 58, and up-regulation of 6 genes, tissue factor pathway inhibitor-2, GRP78, autotaxin, PPP1R7, the DKFZ cDNA p434D0818, and 1 unknown gene. Consistent with previously described roles of several of the altered genes, MKK7(3E) inhibited cell proliferation. These data implicate active MKK7 in the negative regulation of cell proliferation and provide evidence for a new role for this kinase in the regulation of a distinct, hitherto unrecognized set of genes. PMID: 11714698 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 174: J Immunol. 2001 Nov 15;167(10):6021-30. Microenvironment produced by acute myeloid leukemia cells prevents T cell activation and proliferation by inhibition of NF-kappaB, c-Myc, and pRb pathways. Buggins AG, Milojkovic D, Arno MJ, Lea NC, Mufti GJ, Thomas NS, Hirst WJ. Department of Haematological Medicine, Leukaemia Sciences, Guy's, King's and St. Thomas' School of Medicine, Rayne Institute, London, United Kingdom. andrea.buggins@kcl.ac.uk Tumors produce a variety of immunosuppressive factors which can prevent the proliferation and maturation of a number of normal hemopoietic cell types. We have investigated whether primary acute myeloid leukemia (AML) cells have an effect on normal T cell function and signaling. Tumor cell supernatant (TSN) from AML cells inhibited T cell activation and Th1 cytokine production and also prevented activated T cells from entering the cell cycle. These effects occurred in the absence of AML cell-T cell contact. We have demonstrated that AML TSN contained none of the immunosuppressors described to date, namely gangliosides, nitric oxide, TGF-beta, IL-10, vascular endothelial growth factor, or PGs. Furthermore, IL-2 did not overcome the block, despite normal IL-2R expression. However, the effect was overcome by preincubation with inhibitors of protein secretion and abolished by trypsinization, indicating that the active substance includes one or more proteins. To determine the mechanism of inhibition, we have studied many of the major pathways involved in T cell activation and proliferation. We show that nuclear translocation of NFATc and NF-kappaB are markedly reduced in T cells activated in the presence of primary AML cells. In contrast, calcium mobilization and activation of other signal transduction pathways, namely extracellular signal-regulated kinase1/2, p38, and STAT5 were unaffected, but activation of c-Jun N-terminal kinase 1/2 was delayed. Phosphorylation of pRb by cyclin-dependent kinase 6/4-cyclin D and of p130 did not occur and c-Myc, cyclin D3, and p107 were not induced, consistent with cell cycle inhibition early during the transition from G(0) to G(1). Our data indicate that TSN generated by AML cells induces T cell immunosuppression and provides a mechanism by which the leukemic clone could evade T cell-mediated killing. PMID: 11698483 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 175: Mol Cell Biochem. 2001 Jun;222(1-2):69-76. Carcinogenic potential and genomic instability of beryllium sulphate in BALB/c-3T3 cells. Keshava N, Zhou G, Spruill M, Ensell M, Ong TM. Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, West Virginia 26505-2845, USA. Occupational exposure to beryllium (Be) and Be compounds occurs in a wide range of industrial processes. A large number of workers are potentially exposed to this metal during manufacturing and processing, so there is a concern regarding the potential carcinogenic hazard of Be. Studies were performed to determine the carcinogenic potential of beryllium sulfate (BeSO4) in cultured mammalian cells. BALB/c-3T3 cells were treated with varying concentrations of BeSO4 for 72 h and the transformation frequency was determined after 4 weeks of culturing. Concentrations from 50-200 microg BeSO4/ml, caused a concentration-dependent increase (9-41 fold) in transformation frequency. Non-transformed BALB/c-3T3 cells and cells from transformed foci induced by BeSO4 were injected into both axillary regions of nude mice. All ten Be-induced transformed cell lines injected into nude mice produced fibrosarcomas within 50 days after cell injection. No tumors were found in nude mice receiving non-transformed BALB/c-3T3 cells 90 days post-injection. Gene amplification was investigated in K-ras, c-myc, c-fos, c-jun, c-sis, erb-B2 and p53 using differential PCR while random amplified polymorphic DNA fingerprinting was employed to detect genomic instability. Gene amplification was found in K-ras and c-jun, however no change in gene expression or protein level was observed in any of the genes by Western blotting. Five of the 10 transformed cell lines showed genetic instability using different random primers. In conclusion, these results indicate that BeSO4 is capable of inducing morphological cell transformation in mammalian cells and that transformed cells induced by BeSO4 are potentially tumorigenic. Also, cell transformation induced by BeSO4 may be attributed, in part, to the gene amplification of K-ras and c-jun and some BeSO4-induced transformed cells possess neoplastic potential resulting from genomic instability. PMID: 11678613 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 176: Hypertens Res. 2001 Sep;24(5):579-88. All-trans retinoic acid inhibits vascular smooth muscle cell proliferation targeting multiple genes for cyclins and cyclin-dependent kinases. Kosaka C, Sasaguri T, Komiyama Y, Takahashi H. Department of Clinical Sciences and Laboratory Medicine, Kansai Medical University, Osaka, Japan. chiya@muf.biglobe.ne.jp Retinoids have been shown to promote vascular smooth muscle cell differentiation, although the underlying mechanism is unclear. In fact, treatment of rat aortic smooth muscle cells with all-trans retinoic acid (ATRA) has been shown to markedly elevate the mRNA and protein levels of smooth muscle alpha-actin. Considering that an exit from the cell cycle is a prerequisite for cell differentiation, we examined the effect of ATRA on cellular events during the progression from Go to S phase. Pretreatment with ATRA dose-dependently inhibited DNA synthesis induced by basic fibroblast growth factor. However, ATRA did not inhibit transient activation of mitogen-activated protein kinase (MAPK) in response to mitogenic stimulation. And ATRA consistently failed to influence the phosphorylation of MAPK kinase (MEK) and the expression of MAPK-specific dual phosphatase (MKP-1). ATRA did not interfere with other early mitogenic signals either, such as the phosphorylation of FGF-1 receptor or the induction of immediate early genes c-fos, c-jun, and c-myc. In contrast, ATRA strongly suppressed the pRb kinase activities of the cyclin-dependent kinases (Cdks) Cdk4, Cdk6, and Cdk2. ATRA did not influence the expressions of Cip/Kip family Cdk inhibitors or those of cyclins D1 and D2, whereas it strongly inhibited the expressions of cyclins D3 and E, Cdk4, Cdk6, and Cdk2. These results suggest that ATRA targets multiple genes essential for entry into the cell cycle and for the subsequent progression to G1 phase, but without interrupting early mitogenic signals upstream of MAPK. PMID: 11675954 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 177: Am J Physiol Gastrointest Liver Physiol. 2001 Nov;281(5):G1279-89. Dominant-negative TAK1 induces c-Myc and G(0) exit in liver. Bradham CA, Hatano E, Brenner DA. Department of Medicine, University of North Carolina at Chapel Hill, 27707, USA. Transforming growth factor-beta (TGF-beta)-activated kinase 1 (TAK1), a serine/threonine kinase, is reported to function in the signaling pathways of TGF-beta, interleukin 1, and ceramide. However, the physiological role of TAK1 in vivo is largely unknown. To assess the function of TAK1 in vivo, dominant-negative TAK1 (dnTAK1) was expressed in the rat liver by adenoviral gene transfer. dnTAK1 expression abrogated c-Jun NH(2)-terminal kinase and c-Jun but not nuclear factor (NF)-kappaB or SMAD activation after partial hepatectomy (PH). Expression of dnTAK1 or TAM-67, a dominant-negative c-Jun, induced G(0) exit in quiescent liver and accelerated cell cycle progression after PH. Finally, dnTAK1 and TAM-67 induced c-myc expression in the liver before and after PH, suggesting that G(0) exit induced by dnTAK1 and TAM-67 is mediated by c-myc induction. PMID: 11668037 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 178: J Biol Chem. 2001 Dec 21;276(51):47958-65. Epub 2001 Oct 15. HuA and tristetraprolin are induced following T cell activation and display distinct but overlapping RNA binding specificities. Raghavan A, Robison RL, McNabb J, Miller CR, Williams DA, Bohjanen PR. Department of Microbiology, University of Minnesota, Minneapolis, Minnesota 55455, USA. AU-rich elements found in the 3'-untranslated regions of cytokine and proto-oncogene transcripts regulate mRNA degradation and function as binding sites for the mRNA-stabilizing protein HuA and the mRNA-destabilizing protein tristetraprolin. Experiments were performed to evaluate the expression of HuA and tristetraprolin in purified human T lymphocytes and to evaluate the ability of these proteins to recognize specific AU-rich sequences. HuA is a predominantly nuclear protein that can also be found in the cytoplasm of resting T lymphocytes. Within 1 h after stimulation of T lymphocytes with anti-T cell receptor antibodies or a combination of a phorbol myristate acetate and ionomycin, an increase in cytoplasmic HuA RNA-binding activity was observed. Although absent in resting cells, cytoplasmic tristetraprolin protein was detected 3-6 h following activation. HuA recognized specific AU-rich sequences found in c-jun or c-myc mRNA that were poorly recognized by tristetraprolin. In contrast, tristetraprolin recognized an AU-rich sequence in interleukin-2 mRNA that was poorly recognized by HuA. Both HuA and tristetraprolin, however, recognized AU-rich sequences from c-fos, interleukin-3, tumor necrosis factor-alpha, and granulocyte/macrophage colony-stimulating factor mRNA. HuA may transiently stabilize a subset of AU-rich element-containing transcripts following T lymphocyte activation, and tristetraprolin may subsequently mediate their degradation. PMID: 11602610 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 179: Brain Res Mol Brain Res. 2001 Oct 19;94(1-2):48-58. Differential expression of active, phosphorylation-dependent MAP kinases, MAPK/ERK, SAPK/JNK and p38, and specific transcription factor substrates following quinolinic acid excitotoxicity in the rat. Ferrer I, Blanco R, Carmona M. Unitat de Neuropatologia, Servei d'Anatomia Patologica, Hospital Princeps d'Espanya (Bellvitge), c/ Feixa Llarga sn, 08907, Hospitalet de Llobregat, Spain. iferrer@sakma.es Excitotoxicity is considered a major cell death inductor in neurodegeneration. Yet mechanisms involved in cell death and cell survival following excitotoxic insults are poorly understood. Expression of active, phosphorylation-dependent mitogen-activated extracellular signal-regulated kinases (MAPK/ERKs), stress activated c-Jun N-terminal kinases (SAPK/JNKs) and p38 kinases, as well as their putative active specific transcriptional factor substrates CREB, Elk-1, ATF-2, c-Myc and c-Jun, have been examined following intracortical injection of the glutamate analogue quinolinic acid (QA). Increased JNK(P) and p38(P) immunoreactivity has been found in the core at 1 h following QA injection, whereas increased MAPK(P) immunoreactivity occurs in neurons and glial cells localised around the lesion and in neurons in remote cortical regions. This is accompanied by strong phosphorylated Ser63 c-Jun (c-Jun(P)) immunoreactivity in the core at 3 h, and by strong phosphorylated CREB, Elk-1 and ATF-2 (CREB(P), Elk-1(P) and ATF-2(P)) immunoreactivity mainly in neurons around the core at 24 h following QA injection. Examination with the method of in situ end-labelling of nuclear DNA fragmentation has revealed large numbers of positive cells with no apoptotic morphology in the core at 24 h, thus indicating that JNK(P), p38(P) and c-Jun(P) over-expression precedes cell death. In contrast, MAPK(P), CREB(P), Elk-1(P) and ATF-2(P), but not phosphorylated c-Myc (c-Myc(P)), over-expression correlates with cell survival. Examination of cleaved, active caspase-3 has shown specific immunoreactivity restricted to a few hematogenous cells in the area of injection. Since cleaved caspase-3 is not expressed by dying cells in the present paradigm, JNK(P), p38(P) and c-Jun(P) expression is not associated with caspase-3 activation. The present results demonstrate selective activation of specific MAPK signals which are involved either in cell death or cell survival triggered by excitotoxic insult. PMID: 11597764 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 180: Apoptosis. 2001 Dec;6(6):469-77. Mechanisms of tamoxifen-induced apoptosis. Mandlekar S, Kong AN. Department of Drug Metabolism and Pharmacokinetics, DuPont Pharmaceuticals Company, Newark, DE 19711, USA. sandhya.mandlekar@dupontpharma.com Tamoxifen (TAM) has been used in the treatment of breast cancer for over a decade. The observed clinical efficacy of TAM has been attributed to both growth arrest and induction of apoptosis within the breast cancer cells. Although the primary mechanism of action of TAM is believed to be through the inhibition of estrogen receptor (ER), research over the years has indicated that additional, non-ER-mediated mechanisms exist. These include modulation of signaling proteins such as protein kinase C (PKC), calmodulin, transforming growth factor-beta (TGFbeta), and the protooncogene c-myc. Recent studies, including those from our laboratory, have implicated the role of caspases and mitogen-activated protein kinases (MAPK), including c-Jun N-terminal kinase (JNK) and p38 in TAM-induced apoptotic signaling. Oxidative stress, mitochondrial permeability transition (MPT), ceramide generation as well as changes in cell membrane fluidity may also play important roles in TAM-induced apoptosis. These various signaling pathways underlying TAM-induced apoptosis will be reviewed in this article. Publication Types: Review PMID: 11595837 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 181: Mol Carcinog. 2001 Sep;32(1):28-35. Gene expression profile in BALB/c-3T3 cells transformed with beryllium sulfate. Joseph P, Muchnok T, Ong T. Molecular Epidemiology Laboratory, Toxicology and Molecular Biology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, West Virginia 26505, USA. Differential gene expression was studied to understand the potential molecular mechanism responsible for cell transformation and tumorigenesis induced by beryllium. Cell lines were derived from tumors developed in nude mice injected subcutaneously with BALB/c-3T3 cells morphologically transformed with beryllium sulfate. Using the Atlas mouse 1.2 cDNA expression microarray, the expression profiles of 1176 genes, belonging to several different functional categories, were studied in the tumor cells as well as in the nontransformed control cells. Expression of 18 genes belonging to two functional groups was found to be consistently and reproducibly different (at least twofold) in the tumor cells compared with the control cells. The functional groups and the differentially expressed genes are as follows: The cancer-related genes (nine genes) were the ets-related transcription factor activated by ras, colony-stimulating factor, A-myb, sky, cot1, c-fos, c-jun, c-myc, and R-ras proto-oncogenes. The DNA synthesis, repair, and recombination genes (nine genes) were the DNA replication licensing factor MCM4, the DNA replication licensing factor MCM5, the DNA mismatch repair gene PMS2, the DNA excision repair gene, the DNA mismatch repair gene MSH2, the ultraviolet excision repair gene Rad23 DNA ligase 1, Rad51, and Rad52. The differential gene expression profile was confirmed with reverse transcription-polymerase chain reaction using primers specific for the differentially expressed genes. In general, expression of the cancer-related genes was upregulated, while expression of genes involved in DNA synthesis, repair, and recombination was downregulated in the tumor cells compared with the control cells. Using c-fos and c-jun, two of the differentially expressed genes, as model genes, we have found that in the nontransformed BALB/c-3T3 cells, the beryllium-induced transcriptional activation of these genes was dependent on pathways of protein kinase C and mitogen-activated protein kinase and independent of reactive oxygen species. These results indicate that beryllium-induced cell transformation and tumorigenesis are accompanied by and are possibly a product of alterations in expression of genes related to cancer and to DNA synthesis, repair, and recombination. Copyright 2001 Wiley-Liss, Inc. PMID: 11568973 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 182: Toxicol Pathol. 2001 Jul-Aug;29(4):483-91. Differential effects of transforming growth factor-beta1, a fibrogenic factor, on macrophage-like cells (HS-P) and myofibroblastic cells (MT-9) in vitro. Yamate J, Maeda M, Benn SJ, Laithwaite JE, Allan A, Ide M, Kuwamura M, Kotani T, Sakuma S, Lamarre J. Department of Veterinary Pathology, Graduate School of Agriculture and Biological Science, Osaka Prefecture University, Sakai, Japan. yamate@vet.osakafu-u.ac.jp Transforming growth factor-beta1 (TGF-beta1) produced by infiltrating macrophages plays a role in fibrotic disorders through the induction of myofibroblasts. To explore possible mechanisms by which TGF-beta1 may act in this context, we investigated effects of TGF-beta1 on macrophage-like (HS-P) and myofibroblastic (MT-9) cells, two novel cell lines developed by us. Immunocytochemically, the addition of TGF-beta1 (0, 0.1, 0.5, and 1.0 ng/ml) dose-dependently suppressed the expressions of antigens recognized by macrophage/histiocyte-specific antibodies (ED1 and ED2) in HS-P cells, whereas the addition concomitantly increased the number of anti-alpha-smooth muscle actin antibody-positive myofibroblastic cells, suggesting a possible phenotypical modulation of macrophages into myofibroblasts in the fibrotic lesions. By contrast, MT-9 cells did not show such immunophenotypical changes following TGF-beta1 addition. DNA synthesis, measured by tritiated thymidine-incorporation, was inhibited in a dose-dependent manner in MT-9 cells by TGF-beta1 addition (0, 0.1, 0.2, 0.5, 1.0, 5, and 10 ng/ml), but that in HS-P cells was unchanged. Northern blot analysis revealed that expressions of cell cycle-related early genes, c-jun and c-myc, were increased in HS-P cells after TGF-beta1 (1 ng/ml) addition, with c-jun showing peak expression prior to c-myc. By contrast, the peak expressions of c-jun and c-myc were delayed in TGF-beta1 (1 ng/ml)-added MT-9 cells, and their levels were less in MT-9 cells than in HS-P cells. Furthermore, TGF-beta1 (1 and 10 ng/ml) induced DNA laddering in MT-9 cells, but did not in HS-P cells. Based on these findings, it was speculated that TGF-beta1 could have induced G1 arrest in cell cycle and apoptosis in MT-9 cells. The present study showed that there were significant differences in the effects of TGF-beta1 between macrophage-like HS-P cells and myofibroblastic MT-9 cells, presumably depending on divergent susceptibilities to TGF-beta1 between both cell types. Because such cell types are key cells in the fibrogenesis, HS-P and MT-9 might be useful models for investigating the pathogenesis of fibrosis in vitro. PMID: 11560254 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 183: Gut. 2001 Oct;49(4):565-76. Identification of novel molecules and pathogenic pathways in primary biliary cirrhosis: cDNA array analysis of intrahepatic differential gene expression. Shackel NA, McGuinness PH, Abbott CA, Gorrell MD, McCaughan GW. AW Morrow Gastroenterology and Liver Centre, Centenary Institute of Cancer Medicine and Cell Biology, Royal Prince Alfred Hospital and the University of Sydney, Sydney, Australia. BACKGROUND: Primary biliary cirrhosis (PBC) is an autoimmune disease in which the pathogenesis of progressive liver injury is poorly understood. AIM: To provide novel insights into the pathogenesis of PBC related liver injury using cDNA array analysis, which simultaneously examines expression of many genes. METHODS: Utilising cDNA arrays of 874 genes, PBC was compared with primary sclerosing cholangitis (PSC) associated cirrhosis and non-diseased liver. Differential expression of 10 genes was confirmed by real time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Array analysis identified many differentially expressed genes that are important in inflammation, fibrosis, proliferation, signalling, apoptosis, and oxidative stress. PBC was associated with increased expression of both Th1 and Th2 type molecules of the immune response. Fibrosis related gene expression featured upregulation of connective tissue growth factor and transforming growth factor beta3. Many more apoptosis associated molecules exhibited increased expression, consistent with apoptosis being a more active and regulated process, in PSC associated cirrhosis than in PBC. Increased expression of many genes of the Wnt and notch pathways implicated these highly conserved and linked pathways in PBC pathogenesis. The observed increases in expression of c-jun, c-myc, and c-fos related antigen 1 are consistent with increased Wnt pathway activity in PBC. Differential expression of four components of the Wnt pathway, Wnt-5a, Wnt-13, FRITZ, and beta-catenin, was confirmed by quantitative RT-PCR. CONCLUSION: Many genes implicated in intrahepatic inflammation, fibrosis, and regeneration were upregulated in PBC cirrhosis. In particular, increased expression of a number of Drosophila homologues was seen in PBC. PMID: 11559656 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 184: Lab Invest. 2001 Sep;81(9):1299-307. Expression of presumed specific early and late factors associated with liver regeneration in different rat surgical models. Laurent S, Otsuka M, De Saeger C, Maiter D, Lambotte L, Horsmans Y. Gastroenterology Laboratories, Universite Catholique de Louvain, Brussels, Belgium. Experiments performed on the portal branch ligation (PBL) model indicate that early changes observed after surgery are not related to the regenerative process because they also occur in atrophying lobes. To further confirm the lack of specificity of the early events and to exclude the influence of circulatory factors released by proliferating lobes on their occurrence, we investigated this response after sham operation (SO) and portacaval shunt (PCS), a model characterized by liver atrophy. We also attempted to determine expression of later events associated specifically with regeneration, ie, expression of p53 or c-Ha-ras, or inhibition of proliferation, ie, interleukin-1beta (IL-1beta) and transforming growth factor-beta1 (TGF-beta1) after partial (PH) and temporary partial (TPH) hepatectomy, SO and PCS. Nuclear factor-kappaB (NF-kappaB) and signal transducer and activator of transcription 3 (STAT3) DNA binding were assessed by electrophoretic mobility shift assay (EMSA), interleukin-6 (IL-6) mRNA by reverse transcription-polymerase chain reaction (RT-PCR), c-myc and c-jun mRNAs by Northern blot analysis at 0.5 and 2 hours, p53 and c-Ha-ras mRNAs by Northern blot analysis at 8 and 24 hours, and IL-1beta and TGF-beta1 by RT-PCR at 24 hours. The early response including an increase of NF-kappaB, STAT3, IL-6, and immediate-early genes expression was present after PH, PCS, and SO. In SO, slight differences were observed in comparison with PH: no NF-kappaB p65/p50 DNA binding was observed, only three of six SO rats were positive for IL-6, and immediate-early genes induction showed differences in the intensity of the response. At later times, p53 mRNA increased at 8 hours after PH and TPH, c-Ha-ras mRNA at 24 hours after PH, and IL-1beta mRNA at 24 hours after PCS. Early events are not specifically associated with the reduction of liver mass or with the regenerative process, are not predictive of future cell fate, and are most likely related to surgical stress. p53 and c-Ha-ras induction is closely associated with cell cycle progression whereas IL-1beta, but not TGF-beta1, appears to be one of the negative growth regulators that might play an important role in atrophy. PMID: 11555677 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 185: Biol Sci Space. 1994 Jun;8(2):94-102. [Induction of gene expression of cancer-related genes by environmental stresses] [Article in Japanese] Matsumoto H, Ohnishi T. Department of Anatomy, Nara Medical University. Many environmental elements induce the stress response in organisms. In order to examine whether the space condition brings cancer causing on mammals, we propose the importance of the study about the effects of various environmental stresses on the gene expression of oncogenes and tumor-suppressor genes. Then we reviewed numerous findings about the induction of gene expression by environmental stresses. Many investigators have reported that three oncogenes of c-fos, c-jun and c-myc, so called "Early Response Genes", were induced at transcriptional level by a diverse set of stresses such as heat, UV and ionizing radiation, and that the accumulation of the product of a tumor-suppressor gene, p53 was induced at post-translational level by the same stresses. Publication Types: Review Review, Tutorial PMID: 11542736 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 186: Mutat Res. 2001 Sep 1;480-481:201-7. Cell proliferation in cancer prevention; effects of preventive agents on estrogen-related endometrial carcinogenesis model and on an in vitro model in human colorectal cells. Mori H, Niwa K, Zheng Q, Yamada Y, Sakata K, Yoshimi N. Department of Pathology, Gifu University School of Medicine, 40 Tsukasa-machi, Gifu 500-8705, Japan. hidmori@cc.gifu-u.ac.jp Proto-oncogenes such as c-fos, c-jun and c-myc are known to relate to cell proliferation and differentiation. Some oriental herbal medicines like Glycyrrhizae radix or Juzen-taiho-to were found to suppress estradiol-17 beta (E2)-induced expression of c-fos/jun in uterine corpus and inhibited N-methyl-N-nitrosourea and E2-induced endometrial carcinogenesis in mice. It is suggested that the effects of such oriental drugs are exerted probably through suppression of estrogen-induced c-fos/jun expression and they are promising preventing agents for endometrial cancers. In the combined in vitro assay for cell proliferation (MTS assay) and apoptosis (DNA fragmentation) in human colorectal cancer cells (Colo 320), a number of naturally occurring chemopreventive agents such as curcumin, quercetin, auraptene, 1'-acetoxychavicol acetate (ACA) and indole-3-carbinol were shown to generate apoptosis as well as to inhibit cell proliferation. The results suggest a mode of action of these chemopreventive agents and also imply that such in vitro short term assay is useful for detection of new agents for cancer prevention. PMID: 11506814 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 187: Oncogene. 2001 Aug 2;20(34):4629-39. Constitutive expression of ectopic c-Myc delays glucocorticoid-evoked apoptosis of human leukemic CEM-C7 cells. Medh RD, Wang A, Zhou F, Thompson EB. Department of Human Biological Chemistry and Genetics, The University of Texas Medical Branch, Galveston, Texas, TX-77555-0645, USA. rmedh@utmb.edu Sensitivity to glucocorticoid (GC)-evoked apoptosis in lymphoid cell lines correlates closely with GC-mediated suppression of c-Myc expression. To establish a functional role for c-Myc in GC-mediated apoptosis, we have stably expressed MycER(TM), the human c-Myc protein fused to the modified ligand-binding domain of the murine estrogen receptor alpha, in GC-sensitive CEM-C7-14 cells. In CEM-C7-14 cells, MycER(TM) constitutively imparts c-Myc functions. Cells expressing MycER(TM) (C7-MycER(TM)) exhibited a marked reduction in cell death after 72 h in 100 nM dexamethasone (Dex), with 10-20-fold more viable cells when compared to the parental CEM-C7-14 clone. General GC responsiveness was not compromised, as evidenced by Dex-mediated suppression of endogenous c-Myc and cyclin D3, and induction of c-Jun and the glucocorticoid receptor. MycER(TM) also blunted Dex-mediated upregulation of p27(kipI) and suppression of the Myc target p53. In comparison to parental CEM-C7-14 cells, Dex-evoked DNA strand breaks were negligible and caspase activation was delayed, but the extent of G1 cell cycle arrest was similar in C7-MycER(TM) cells. Myc-ER(TM) did not result in permanent, complete resistance to GC however, and the GC-treated cells eventually died, indicative of redundant or interactive mechanisms in the GC-evoked lytic response of lymphoid cells. Our results emphasize the importance of c-Myc suppression in GC-evoked apoptosis of CEM-C7-14 cells. PMID: 11498786 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 188: Biol Signals Recept. 2001 Sep-Oct;10(5):294-8. The anti-tumour effect of Klebsiella pneumoniae capsular polysaccharides. Kwok TT, Chen P, Liu PY, Tang YC, Kong SK, Fung KP, Choy YM. Department of Biochemistry, The Chinese University of Hong Kong. Hong Kong. K24 capsular polysaccharide (K24-CPS), with a known structure of a repeating unit, was isolated from the capsule of Klebsiella pneumoniae serotype K24. The polysaccharide was found to suppress the proliferation of Ehrlich ascites tumour (EAT) cells in vitro, but did not alter the cell cycle distribution of cells. K24-CPS treatment reduced the tyrosine phosphorylation of some proteins in EAT cells. Furthermore, the treatment also decreased the expression of c-JUN, but had no effect on the levels of c-FOS and c-MYC. It is speculated that the growth suppression effect of K24-CPS may be related to its effect in down-regulating c-JUN expression. Copyright 2001 S. Karger AG, Basel PMID: 11490094 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 189: FEBS Lett. 2001 Jul 13;501(1):59-64. Induction of bulk and c-myc P2 promoter-specific DNA damage by an anti-topoisomerase II agent salvicine is an early event leading to apoptosis in HL-60 cells. Meng L, Ding J. Division of Antitumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 200031, Shanghai, PR China. Salvicine is a novel diterpenoid quinone derivative possessing strong antitumor activities and was demonstrated to stabilize the DNA topoisomerase II (Topo II) cleavage complex in vitro and in vivo. In the present work we investigated the possible mechanism through which disturbance of Topo II by salvicine led to cell death. We found that salvicine induced DNA strand breaks in human promyelocytic leukemia HL-60 cells and DNA damage correlated with cell growth inhibition. DNA damage induced by brief exposure to salvicine could be partially reversed, but early DNA breaks triggered the process of apoptosis. Preferential damage in the P2 promoter region of the oncogene c-myc was detected, whereas no obvious DNA damage was found in the 3' region of the same gene. Furthermore, the expression of some protooncogenes such as c-myc, c-fos and c-jun was examined, showing that salvicine produced a reduction in the transcription rate of c-myc in a dose-dependent manner and a marked induction of c-fos and c-jun expression was observed. It appears possible that DNA damage within such genomic regions is an early event, which could lead to growth inhibition mediated by alterations of the expression of selected proliferation regulatory genes, such as c-myc, c-fos and c-jun, and ultimately cell death. PMID: 11457456 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 190: Mol Biol Cell. 2001 Jul;12(7):2171-83. Src family kinases are required for prolactin induction of cell proliferation. Fresno Vara JA, Caceres MA, Silva A, Martin-Perez J. Instituto de Investigaciones Biomedicas, Consejo Superior de Investigaciones Cientificas, Madrid 28029, Spain. Prolactin (PRL) is a pleiotropic cytokine promoting cellular proliferation and differentiation. Because PRL activates the Src family of tyrosine kinases (SFK), we have studied the role of these kinases in PRL cell proliferation signaling. PRL induced [(3)H]thymidine incorporation upon transient transfection of BaF-3 cells with the PRL receptor. This effect was inhibited by cotransfection with the dominant negative mutant of c-Src (K>A295/Y>F527, SrcDM). The role of SFK in PRL-induced proliferation was confirmed in the BaF-3 PRL receptor-stable transfectant, W53 cells, where PRL induced Fyn and Lyn activation. The SFK-selective inhibitors PP1/PP2 and herbimycin A blocked PRL-dependent cell proliferation by arresting the W53 cells in G1, with no evident apoptosis. In parallel, PP1/PP2 inhibited PRL induction of cell growth-related genes c-fos, c-jun, c-myc, and odc. These inhibitors have no effect on PRL-mediated activation of Ras/Mapk and Jak/Start pathways. In contrast, they inhibited the PRL-dependent stimulation of the SFKs substrate Sam68, the phosphorylation of the tyrosine phosphatase Shp2, and the PI3K-dependent Akt and p70S6k serine kinases. Consistently, transient expression of SrcDM in W53 cells also blocked PRL activation of Akt. These results demonstrate that activation of SFKs is required for cell proliferation induced by PRL. PMID: 11452011 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 191: News Physiol Sci. 2001 Jun;16:106-9. Polyamines: new cues in cellular signal transduction. Bachrach U, Wang YC, Tabib A. Department of Molecular Biology, Hebrew University-Hadassah Medical School, 91120 Jerusalem, Israel. The naturally occurring polyamines putrescine, spermidine, and spermine are involved in signal transduction. This has been demonstrated by using inhibitors for polyamine biosynthesis (such as alpha-difluoromethylornithine) or adding polyamines to cultured cells. Different polyamines, preferentially activated protein kinases (tyrosine kinases and MAP kinases), stimulated the expression of nuclear protooncogenes (myc, jun, and fos). Publication Types: Review Review, Tutorial PMID: 11443226 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 192: J Comp Pathol. 2001 Jul;125(1):15-24. Effects of lipopolysaccharide on a macrophage-like cell line (HS-P) from a rat histiocytic sarcoma. Yamate J, Maeda M, Benn SJ, Laithwaite JE, Allan A, Ide M, Kuwamura M, Kotani T, Sakuma S, LaMarre J. Department of Veterinary Pathology, Graduate School of Agriculture and Biological Science, Osaka Prefecture University, Gakuencho 1-1, Sakai, Osaka, 599-8531, Japan. Lipopolysaccharide (LPS) is a major modulator of macrophage functions. To characterize a newly established rat histiocytic sarcoma-derived cell line (HS-P), immunophenotypic changes and cellular growth responses of HS-P cells exposed to LPS were investigated and compared with those of MT-9 cells isolated from a rat malignant fibrous histiocytoma. MT-9 cells have somewhat histiocytic features, because occasional cells react to rat macrophage-specific antibodies. Addition of LPS to cultured HS-P cells increased the numbers of cells immunopositive to ED1 (rat macrophage-specific antibody) and ED2 (rat histiocyte-specific antibody) and stimulated the phagocytosis of latex beads, whereas LPS-treated MT-9 cells did not show such immunophenotypic changes. LPS-treated HS-P cells showed enhanced immunolabelling of alpha-smooth muscle actin, suggesting a possible modulation of macrophages towards myofibroblastic cells. To evaluate cellular growth after the addition of LPS or fetal bovine serum, DNA synthesis was examined by measuring tritiated thymidine incorporation, and the mRNA expression of c- jun and c- myc (immediate early genes in the cell cycle) was examined by Northern blot analysis. In HS-P cells, the addition of serum greatly increased DNA synthesis and induced high expression of c- jun and c- myc; in contrast, LPS markedly depressed DNA synthesis and reduced the expression of c- jun and c- myc. HS-P cells were more sensitive than MT-9 cells to the growth-promoting effect of serum and the growth-inhibiting effect of LPS. The study demonstrated that HS-P cells are highly LPS-responsive, indicating that they would be useful for studies of macrophage functions. Copyright Harcourt Publishers Ltd. PMID: 11437512 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 193: Proc Natl Acad Sci U S A. 2001 Jun 5;98(12):6674-9. Stat1-independent regulation of gene expression in response to IFN-gamma. Ramana CV, Gil MP, Han Y, Ransohoff RM, Schreiber RD, Stark GR. Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA. Although Stat1 is essential for cells to respond fully to IFN-gamma, there is substantial evidence that, in the absence of Stat1, IFN-gamma can still regulate the expression of some genes, induce an antiviral state and affect cell growth. We have now identified many genes that are regulated by IFN-gamma in serum-starved Stat1-null mouse fibroblasts. The proteins induced by IFN-gamma in Stat1-null cells can account for the substantial biological responses that remain. Some genes are induced in both wild-type and Stat1-null cells and thus are truly Stat1-independent. Others are subject to more complex regulation in response to IFN-gamma, repressed by Stat1 in wild-type cells and activated in Stat1-null cells. Many genes induced by IFN-gamma in Stat1-null fibroblasts also are induced by platelet-derived growth factor in wild-type cells and thus are likely to be involved in cell proliferation. In mouse cells expressing the docking site mutant Y440F of human IFN-gamma receptor subunit 1, the mouse Stat1 is not phosphorylated in response to human IFN-gamma, but c-myc and c-jun are still induced, showing that the Stat1 docking site is not required for Stat1-independent signaling. PMID: 11390994 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 194: Proc Natl Sci Counc Repub China B. 2001 Apr;25(2):59-66. Mechanisms of cancer chemoprevention by curcumin. Lin JK, Lin-Shiau SY. Institute of Biochemistry, College of Medicine, National Taiwan University, Taipei, ROC. Curcumin is a major component of the Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animals as indicated by its ability to block colon tumor initiation by azoxymethane and skin tumor promotion induced by phorbol ester TPA. Recently, curcumin has been considered by oncologists as a potential third generation cancer chemopreventive agent, and clinical trials using it have been carried out in several laboratories. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes, such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase. Curcumin is also a potent inhibitor of protein kinase C, EGF-receptor tyrosine kinase and IkappaB kinase. In addition, curcumin inhibits the activation of NFkappaB and the expression of c-jun, c-fos, c-myc and iNOS. It is proposed that curcumin may suppress tumor promotion by blocking signal transduction pathways in the target cells. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin, and these compounds were subsequently convened into monoglucuronide conjugates. The experimental results suggest that curcumin-glucuronide, dihydrocurcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are major metabolites of curcumin in mice. Publication Types: Review Review, Tutorial PMID: 11370761 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 195: Arch Toxicol. 2001 Mar;75(1):28-35. Increased expression of iNOS and c-fos via regulation of protein tyrosine phosphorylation and MEK1/ERK2 proteins in terminal bronchiole lesions in the lungs of rats exposed to cigarette smoke. Chang WC, Lee YC, Liu CL, Hsu JD, Wang HC, Chen CC, Wang CJ. Department of Medical Technology, Chung Hwa Institute of Technology, Tainan, Taiwan. Epidemiological evidence suggests that smoking is a major cause of human lung cancer. However, the mechanism by which cigarette smoke induces the cancer remains unestablished. To evaluate the effects of cigarette smoke on the expression of inducible nitric oxide synthase (iNOS), nuclear protooncogenes and related mitogen-activated protein kinases (MAPKs) in rat lung tissue, a histopathological study of the effects of gas-phase cigarette smoke on rat lung tissue were carried out. The terminal bronchioles were found to be infiltrated predominantly by lymphocytes in the peribronchiolar region and a mild to moderate degree of emphysema was noted in the alveolar spaces. The terminal bronchioles also showed marked lipid peroxidation, dilatation, and peribronchiolar fibrosis. Immunohistochemical evaluation showed that the expression of iNOS, NF-kappa B, MAPKs (MEK1, ERK2), phosphotyrosine protein and c-fos was increased in the terminal bronchioles but protein kinase C (PKC), MEKK-1, c-jun, p38 and c-myc showed no change. These results provide evidence to suggest that exposure to cigarette smoke results in oxidant stress which leads to the stimulation of iNOS and c-fos together with the induction of protein tyrosine phosphorylation and MEK1/ERK2 which in turn may promote lung pathogenesis. PMID: 11357518 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 196: Toxicol Sci. 2001 Jun;61(2):295-303. Cadmium-induced cell transformation and tumorigenesis are associated with transcriptional activation of c-fos, c-jun, and c-myc proto-oncogenes: role of cellular calcium and reactive oxygen species. Joseph P, Muchnok TK, Klishis ML, Roberts JR, Antonini JM, Whong WZ, Ong T. Toxicology and Molecular Biology Branch, Pathology and Physiology Research Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, West Virginia 26505, USA. pcj5@cdc.gov The molecular mechanisms of carcinogenesis by cadmium were studied using BALB/c-3T3 cell transformation and nude mouse tumorigenesis models. BALB/c-3T3 cells transformed with cadmium chloride were subcutaneously injected into nude mice to develop tumors and the cell lines derived from these tumors were used in the present study. The proto-oncogenes c-fos and c-jun were overexpressed in 100% (10 out of 10) of the cell lines, while a statistically significant overexpression of c-myc was observed in 40% (4 out of 10) of the cell lines. Analysis of tumor cells stained with fluorescent dyes specific for reactive oxygen species revealed that these cells possessed markedly higher levels of superoxide anion and hydrogen peroxide compared with the nontransformed cells. Similarly, the intracellular calcium level was higher in the tumor cells compared with the nontransformed cells. Overexpression of the proto-oncogenes in these cells was blocked by treating the cells with superoxide dismutase, catalase, and 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetra acetoxy methyl ester (BAPTA/AM), which are scavengers of superoxide anion, hydrogen peroxide, and calcium, respectively. This confirmed that the overexpression of the proto-oncogenes in the tumor cells required elevated intracellular levels of reactive oxygen species and calcium. In addition to the scavengers of reactive oxygen species and calcium, inhibitors specific for transcription (actinomycin D), protein kinase C (RO-31-8220), and MAP kinase (PD 98059) also blocked the cadmium-induced overexpression of the proto-oncogenes in the tumor cells. Exposure of the nontransformed BALB/c-3T3 cells to 20 microM cadmium chloride for 1 h caused elevated intracellular levels of superoxide anion, hydrogen peroxide, and calcium, with corresponding increases in the expression levels of c-fos, c-jun, and c-myc. As in the case of the tumor cells, treating the nontransformed cells with the various modulators prior to their exposure to cadmium chloride resulted in inhibition in the expression of the proto-oncogenes. Based on these data, we conclude that the cadmium-induced overexpression of cellular proto-oncogenes is mediated by the elevation of intracellular levels of superoxide anion, hydrogen peroxide, and calcium. Further, the cadmium-induced overexpression of the proto-oncogenes is dependent on transcriptional activation as well as on pathways involving protein kinase C and MAP kinase. PMID: 11353138 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 197: Crit Rev Oral Biol Med. 2001;12(2):152-65. A possible role for the WNT-1 pathway in oral carcinogenesis. Lo Muzio L. Institute of Dental Sciences, University of Ancona, Italy. llomuzio@tin.it Reductions in cell-cell adhesion and stromal and vascular invasion are essential steps in the progression from localized malignancy to metastatic disease for all cancers. Proteins involved in intercellular adhesion, such as E-cadherin and catenin, probably play an important role in metastatic processes and cellular differentiation. While E-cadherin and beta-catenin expression has been extensively studied in many forms of human cancers, less is known about the role of the Wingless-Type-1 (WNT-1) pathway in human tumors. A large body of genetic and biochemical evidence has identified beta-catenin as a key downstream component of the WNT signaling pathway, and recent studies of colorectal tumors have shown a functional link among beta-catenin, adenomatous polyposis coli gene product (APC), and other components of the WNT-1 pathway. WNT-1 pathway signaling is thought to be mediated via interactions between beta-catenin and members of the LEF-1/TCF family of transcription factors. The WNT signal stabilizes beta-catenin protein and promotes its accumulation in the cytoplasm and nucleus. In the nucleus, beta-catenin associates with TCF to form a functional transcription factor which mediates the transactivation of target genes involved in the promotion of tumor progression, invasion, and metastasis, such as C-Myc, cyclin D1, c-jun, fra-1, and u-PAR. There is a strong correlation between the ability of the WNT-1 gene to induce beta-catenin accumulation and its transforming potential in vivo, suggesting that the WNT-1 gene activates an intracellular signaling pathway that can induce the morphological transformation of cells. For these reasons, data obtained from the study of the WNT-1 pathway could be important in our understanding of the mechanisms of epithelial tumors, in general, and probably also of oral squamous cell carcinoma, in particular. Publication Types: Review PMID: 11345525 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 198: Dig Dis Sci. 2001 Apr;46(4):907-14. Immediate-early protooncogene expression and liver function following various extents of partial hepatectomy in the rat. Moser MJ, Gong Y, Zhang MN, Johnston J, Lipschitz J, Minuk GY. Liver Disease Unit, Department of Medicine, University of Manitoba, Winnipeg, Canada. Immediate-early protooncogenes (IEP) are thought to play an important role in hepatocyte replication. Whether the extent of their expression correlates with the strength of the proliferative stimulus and subsequent regenerative activity has yet to be documented in vivo. Data are also lacking with respect to the level at which liver disease is associated with biochemical evidence of hepatic dysfunction. Thus, the objectives of this study were to determine whether a correlation exists between IEP gene mRNA expression and varying extents of partial hepatectomy (PHx) and to document the extent of resection required to result in increases in serum bilirubin levels. Eighty-nine adult, male Sprague-Dawley rats underwent either sham surgery or 20%, 35%, 55%, 70% or 90% PHx. Postoperatively, rats were killed (N = 3-6/group) at 15 and 30 mins and 8 and 24 hrs for c-fos, c-jun, and c-myc mRNA expression by northern blot analyses. Rats killed at 24 hrs also had hepatic regenerative activity documented by [3H]thymidine incorporation into hepatic DNA and serum bilirubin determinations. While c-fos mRNA expression at 15 mins and c-myc mRNA expression at 8 hrs after PHx did not correlate with the extent of PHx (r2 = 0.478 and 0.018, respectively), a weak correlation existed between c-jun mRNA expression at 30 mins and the extent of PHx (r2 = 0.662, P < 0.05). In terms of IEP mRNA expression and hepatic regenerative activity, a strong correlation existed between c-fos mRNA expression and [3H]thymidine incorporation (r2 = 0.851, P < 0.01) but not c-jun or c-myc mRNA expression. Compared to sham operated controls, [3H]thymidine incorporation was 2.0x, 3.4x, 3.2x, 7.8x, and 2.2x increased following 20%, 35%, 55%, 70%, and 90% PHx, respectively. Serum bilirubin levels remained unchanged until 70% PHx, when they increased from baseline values of 0.54+/-0.05 mg/dl to 1.02+/-0.15 mg/dl (P < 0.05). A further increase occurred following 90% PHx (1.83+/-0.30 mg/dl, P < 0.01). In conclusion these findings indicate that c-fos mRNA expression 15 mins after PHx correlates with hepatic regenerative activity but not the strength of the regenerative stimulus and that hepatic parenchymal loss of 55-70% must occur prior to the detection of elevated serum bilirubin levels. The results also indicate that relative to a 70% PHx, 90% PHx is associated with decreased rather than increased hepatic regenerative activity. PMID: 11330432 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 199: Immunology. 2001 Mar;102(3):289-300. Functional dissection of the cytoplasmic subregions of the interleukin-5 receptor alpha chain in growth and immunoglobulin G1 switch recombination of B cells. Moon BG, Yoshida T, Shiiba M, Nakao K, Katsuki M, Takaki S, Takatsu K. Department of Immunology, Division of DNA Biology and Embryo Engineering, Centre for Experimental Medicine, Institute of Medical Science, University of Tokyo, Japan. The interleukin-5 receptor alpha chain (IL-5Ralpha) is known to regulate the development and function of B cells and eosinophils. Although the functions of IL-5Ralpha cytoplasmic domain subregions have been studied extensively using cultured cell lines, this approach has limitations when studying the functions of distinct primary B-cell subpopulations and their responsiveness to IL-5. In the present study, we generated mice on an IL-5Ralpha null background, each expressing a mutant form of an IL-5Ralpha transgene ligated to a mu enhancer and VH promoter, either lacking the cytoplasmic DC3 region or substituting two proline residues for alanine (ApvA) in the membrane-proximal ppvp motif of the cytoplasmic domain. The ppvp motif, which mediates activation of JAK2/STAT5 and Btk, also contributes to c-fos, c-jun and c-myc expression. IL-5Ralpha null mutant mice showed impaired B-1-cell development, reduced serum immunoglobulin G3 (IgG3) and IgM, no IL-5-induced enhancement of B-cell proliferation and IL-5-induced switch recombination from the mu gene to gamma1 gene; these were not recovered following the expression of the ApvA mutant. In contrast, absence of the DC3 region affected the IL-5-induced switch recombination from the mu to the gamma1 gene and B-1-cell development, while IL-5-induced proliferation and IgM production were at levels similar to those of B cells expressing wild-type IL-5Ralpha transgene. The results clearly indicated that the ppvp motif and the DC3 region of IL-5Ralpha played distinct roles in B-cell proliferation and differentiation. Thus, this present approach offers new insights into the functions of the cytoplasmic subregions of IL-5Ralpha, in particular its carboxy-terminal region. PMID: 11298827 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 200: Proc Natl Acad Sci U S A. 2001 Apr 10;98(8):4646-51. Antisense-mediated depletion of p300 in human cells leads to premature G1 exit and up-regulation of c-MYC. Kolli S, Buchmann AM, Williams J, Weitzman S, Thimmapaya B. Department of Microbiology and Immunology and Robert H. Lurie Cancer Center, and Department of Medicine, Northwestern University Medical School, Chicago, IL 60611, USA. The cAMP-response element-binding protein (CREB)-binding protein and p300 are two highly conserved transcriptional coactivators and histone acetyltransferases that integrate signals from diverse signal transduction pathways in the nucleus and also link chromatin remodeling with transcription. In this report, we have examined the role of p300 in the control of the G(1) phase of the cell cycle in nontransformed immortalized human breast epithelial cells (MCF10A) and fibroblasts (MSU) by using adenovirus vectors expressing p300-specific antisense sequences. Quiescent MCF10A and MSU cells expressing p300-specific antisense sequences synthesized p300 at much reduced levels and exited G(1) phase without serum stimulation. These cells also showed an increase in cyclin A and cyclin A- and E-associated kinase activities characteristic of S phase induction. Further analysis of the p300-depleted quiescent MCF10A cells revealed a 5-fold induction of c-MYC and a 2-fold induction of c-JUN. A direct target of c-MYC, CAD, which is required for DNA synthesis, was also found to be up-regulated, indicating that up-regulation of c-MYC functionally contributed to DNA synthesis. Furthermore, S phase induction in p300-depleted cells was reversed when antisense c-MYC was expressed in these cells, indicating that up-regulation of c-MYC may directly contribute to S phase induction. Adenovirus E1A also induced DNA synthesis and increased the levels of c-MYC and c-JUN in serum-starved MCF10A cells in a p300-dependent manner. Our results suggest an important role of p300 in cell cycle regulation at G(1) and raise the possibility that p300 may negatively regulate early response genes, including c-MYC and c-JUN, thereby preventing DNA synthesis in quiescent cells. PMID: 11296295 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 201: Mutat Res. 2001 Apr 18;475(1-2):69-87. Vitamin D and genomic stability. Chatterjee M. Department of Pharmaceutical Technology, Division of Biochemistry, Jadavpur University, 700032, Calcutta, India. m.chatterjee@mailcity.com 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] has been shown to act on novel target tissues not related to calcium homeostasis. There have been reports characterizing 1,25(OH)(2)D(3) receptors and activities in diverse tissues such as brain, pancreas, pituitary, skin, muscle, placenta, immune cells and parathyroid. The receptor hormone complex becomes localized in the nucleus, and undergoes phosphorylation by reacting with a kinase. This form of the receptor then interacts with the Vitamin D responsive element of target gene and modifies the transcription of those genes to develop the action. The modulation of gene transcription results in either the induction or repression of specific messenger RNAs (m-RNAs), ultimately resulting in changes in protein expression needed to produce biological responses. Genes for carbonic anhydrase that are expressed at high levels in osteoclast are known to be involved in bone resorption and Id genes role in osteoblast-osteoclast differentiation reflects the genomic effect of Vitamin D on bones. Genomic action of Vitamin D also explains the biosynthesis of oncogenes, polyamines, lymphokines and calcium binding proteins. However, there is a possibility that some of the actions of 1,25(OH)(2)D(3) may be mediated by non-genomic mechanisms and may not require the binding to Vitamin D receptor (VDR).Vitamin D offers a protection from genotoxic effects of Vitamin D deficiency by increasing the insulin receptor gene expression and BSP (bone sialoprotein), bone-remodeling by decreasing the osteopontin (OPN) m-RNAs, maintaining the normal epidermal structure and enamel matrix. Gonadal insufficiency in Vitamin D deficiency was corrected by vitamin mediated direct regulation of the expression of aramotase gene. The supportive role of Vitamin D in placental function is also evident by its influence on human placental lactogen (hpl) gene transcription accompanied by increase hpl m-RNA levels. Further role of Vitamin D is envisaged in identifying cyclin C as an important target for Vitamin D in cell-cycle regulation.Vitamin D at physiological concentration has been found to protect cell proteins and membranes against oxidative stress by inhibiting the peroxidative attack on membrane lipids. Vitamin D, at a concentration range of 2x10(-8)-5x10(-8)M, induces apoptosis in most cancer cells, stabilizes chromosomal structure and prevents DNA double-strand breaks induced either by endogenous or exogenous factors. Vitamin D is also effective in stimulating DNA synthesis in adult alveolar II cells and provides a novel mechanism of modulation of epithelial cell proliferation in the context of lung development and repair against injury. The regulation of various proto-oncogenes (c-myc, c-fos, c-jun), differentiation inducing properties, antiproliferative effects on keratinocytes and inhibitory effects in several human malignancy ranks Vitamin D as a novel hormone that may have physiological and clinical implication in the carcinogenic process. Publication Types: Review Review, Tutorial PMID: 11295155 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 202: FASEB J. 2001 Apr;15(6):1006-13. Cyclin D1 is an early target in hepatocyte proliferation induced by thyroid hormone (T3). Pibiri M, Ledda-Columbano GM, Cossu C, Simbula G, Menegazzi M, Shinozuka H, Columbano A. Department of Toxicology, Oncology and Molecular Pathology Unit, University of Cagliari, Italy. The thyroid hormone (T3) affects cell growth, differentiation, and regulates metabolic functions via its interaction with the thyroid hormone nuclear receptors (TRs). The mechanism by which TRs mediate cell growth is unknown. To investigate the mechanisms responsible for the mitogenic effect of T3, we have determined changes in activation of transcription factors, mRNA levels of immediate early genes, and levels of proteins involved in the progression from G1 to S phase of the cell cycle. We show that hepatocyte proliferation induced by a single administration of T3 to Wistar rats occurred in the absence of activation of AP-1, NF-kappa B, and STAT3 or changes in the mRNA levels of the immediate early genes c-fos, c-jun, and c-myc. These genes are considered to be essential for liver regeneration after partial hepatectomy (PH). On the other hand, T3 treatment caused an increase in cyclin D1 mRNA and protein levels that occurred much more rapidly compared to liver regeneration after 2/3 PH. The early increase in cyclin D1 expression was associated with accelerated onset of DNA synthesis, as demonstrated by a 20-fold increase of bromodeoxyuridine-positive hepatocytes at 12 h after T3 treatment and by a 20-fold increase in mitotic activity at 18 h. An early increase of cyclin D1 expression was also observed after treatment with nafenopin, a ligand of a nuclear receptor (peroxisome proliferator-activated receptor alpha) of the same superfamily of steroid/thyroid receptors. T3 treatment also resulted in increased expression of cyclin E, E2F, and p107 and enhanced phosphorylation of pRb, the ultimate substrate in the pathway leading to transition from G1 to S phase. The results demonstrate that cyclin D1 induction is one of the earlier events in hepatocyte proliferation induced by T3 and suggest that this cyclin might be a common target responsible for the mitogenic activity of ligands of nuclear receptors. PMID: 11292661 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 203: J Biol Chem. 2001 May 25;276(21):17800-7. Epub 2001 Feb 15. Bcl-XL expression correlates with primary macrophage differentiation, activation of functional competence, and survival and results from synergistic transcriptional activation by Ets2 and PU.1. Sevilla L, Zaldumbide A, Carlotti F, Dayem MA, Pognonec P, Boulukos KE. Institute of Signalisation, Developmental Biology and Cancer, INSERM 470, Centre de Biochimie, Universite de Nice, Faculte des Sciences, 06108 Nice, France. Depriving primary bone marrow-derived macrophages of colony-stimulating factor-1 (CSF-1) induces programmed cell death by apoptosis. We show that cell death is accompanied by decreases in the expression of anti-apoptotic Bcl-x(L) protein and the Ets2 and PU.1 proteins of the Ets transcription factor family. Macrophages require both priming and triggering signals independent of CSF-1 to kill neoplastic cells or microorganisms, and this activation of macrophage competence is accompanied by increased expression of bcl-x(L), ets2, and PU.1. Furthermore, we show that only Ets2 and PU.1, but not Ets1, function in a synergistic manner to transactivate the bcl-x promoter. The synergy observed between PU.1 and Ets2 is dependent on the transactivation domains of both proteins. Although other transcription factors like Fos, c-Jun, Myc, STAT3, and STAT5a are implicated in the activation of macrophage competence or in CSF-1 signaling, no synergy was observed between Ets2 and these transcription factors on the bcl-x promoter. We demonstrate that the exogenous expression of both Ets2 and PU.1 in macrophages increases the number of viable cells upon CSF-1 depletion and that Ets2 and PU.1 can functionally replace Bcl-x(L) in inhibiting Bax-induced apoptosis. Together, these results demonstrate that PU.1 and Ets2 dramatically increase bcl-x activation, which is necessary for the cytocidal function and survival of macrophages. PMID: 11278399 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 204: J Surg Res. 2001 Apr;96(2):289-95. Activation of interleukin-6/STAT3 and liver regeneration following transplantation. Debonera F, Aldeguer X, Shen X, Gelman AE, Gao F, Que X, Greenbaum LE, Furth EE, Taub R, Olthoff KM. Department of Surgery, University of Pennsylvania, Philadelphia 19104, USA. BACKGROUND: Every liver that is procured, stored, and transplanted experiences injury from cold ischemia and reperfusion. Most recover quickly, but some grafts sustain enough injury to result in prolonged organ dysfunction or require retransplantation. The molecular mechanisms involved in early graft function and recovery following cold ischemia and reperfusion (I/R) after liver transplantation have not been well defined. Interleukin (IL)-6 is a critical factor in the mitogenic response within the liver, and is important for cell cycle progression and protection from injury. Activation of the latent transcription factor, STAT3, is dependent on IL-6 release. The role of the IL-6/STAT3 pathway and hepatocellular regeneration in graft recovery and cell cycle progression following cold ischemia and reperfusion was studied in a rat liver transplant orthotopic (OLT) model. Methods. Rat OLT was performed in a syngeneic model. The presence, time course, and magnitude of expression of IL-6, STAT3 activation, and upregulation of target immediate early genes were determined in liver grafts with minimal (<1 h) and prolonged (12 h) cold preservation times followed by transplantation. Progression of the cell cycle and replication was confirmed by BrdU uptake. RESULTS: Prolonged cold ischemia resulted in increased IL-6 expression and STAT3 activation. This correlated with upregulation of junB, c-fos, c-myc, and c-jun, immediate early genes associated with hepatic regeneration. Extensive DNA replication was present in livers with 12-h ischemia, demonstrating successful completion of the cell cycle. CONCLUSIONS: The participation of the IL-6/STAT3 pathway leading to cell cycle progression and regeneration is an important component in the recovery of organs immediately following cold preservation and transplantation. Copyright 2001 Academic Press. PMID: 11266286 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 205: Zhonghua Bing Li Xue Za Zhi. 1998 Apr;27(2):105-8. [Influence of compensatory hepatocyte proliferation on the carcinogenesis of N-nitrosodimethylamine] [Article in Chinese] Chen D, Yan R, Ye Y. Cancer Institute, Sun Yat-sen University of Medical Sciences, Guangzhou 510060. OBJECTIVE: To study the influence of compensatory hepatocyte proliferation on the N-nitrosodimethylamine (NDMA) carcinogenesis in rats. METHODS: NDMA was given to animals of the experimental group 24 hours after partial hepatectomy, and the control group was only treated with NDMA. Expression of gamma-glutamyltransferase (GGT), glutathione S-transferase placental form (GSTP), proliferating cell nuclear antigen (PCNA), insulin-like growth factor-II (IGF-II) and oncogenes was detected. RESULTS: The numbers and areas of GGT- and GSTP-foci in the experimental group were significantly increased in comparing with the control groups. The expression of GSTP was higher than that of GGT. The total tumor incidence of the experimental group was higher than that of the control by the end of the 56th week. Up to week 71, the incidences of liver and other tumors were higher respectively in the experimental group. The amount of PCNA positive cells were corresponding with proliferative condition of the hepatic lesions. The expression of IGF-II, c-myc and H-ras mRNA increased in the altered hepatocyte foci and nodules, but markedly decreased in hepatocellular carcinoma and adenoma. No c-jun mRNA expression was detected in all the normal and abnormal tissues of liver. CONCLUSIONS: The results suggest that compensatory hepatocyte proliferation enhances the carcinogenesis induced by multiple doses of NDMA, and the over expression of IGF-II, c-myc, H-ras may play a synergetic role in NDMA-induced hepatocarcinogenesis. PMID: 11244958 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 206: Biofactors. 2000;13(1-4):153-8. Recent studies on the biofunctions and biotransformations of curcumin. Lin JK, Pan MH, Lin-Shiau SY. Institutes of Biochemistry, College of Medicine, National Taiwan University, Taipei. jklin@ha.mc.ntu.edu.tw Curcumin is a major component of Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animals as indicated by its ability to block colon tumor initiation by azoxymethane and skin tumor promotion induced by phorbol ester TPA. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase. Curcumin is also a potent inhibitor of protein kinase C, EGF-receptor tyrosine kinase and IkappaB kinase. Subsequently, curcumin inhibits the activation of NFkappaB and the expressions of c-jun, c-fos, c-myc and iNOS. It is proposed that curcumin may suppress tumor promotion through blocking signal transduction pathways in the target cells. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin and that these compounds subsequently were converted to monoglucuronide conjugates. These results suggest that curcumin-glucuronide, dihydro-curcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are major metabolites of curcumin in mice. Publication Types: Review Review, Tutorial PMID: 11237176 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 207: BMC Cancer. 2001;1:1. Epub 2001 Jan 17. Inhibition of carcinogen induced c-Ha-ras and c-fos proto-oncogenes expression by dietary curcumin. Limtrakul P, Anuchapreeda S, Lipigorngoson S, Dunn FW. Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand 50200. plimtrak@mail.med.cmu.ac.th BACKGROUND: We investigated the chemopreventive action of dietary curcumin on 7,12-dimethylbenz(a)anthracene (DMBA)-initiated and 12,0-tetradecanoylphorbol-13-acetate (TPA)-promoted skin tumor formation in Swiss albino mice. Curcumin, a yellow coloring matter isolated from roots of Curcuma longa Linn, is a phenolic compound possessing antioxidant, free radical scavenger, and antiinflammatory properties. It has been shown by previously reported work that TPA-induced skin tumors were inhibited by topical application of curcumin, and curcumin has been shown to inhibit a variety of biological activities of TPA. Topical application of curcumin was reported to inhibit TPA-induced c-fos, c-jun and c-myc gene expression in mouse skin. This paper reports the effects of orally administered curcumin, which was consumed as a dietary component at concentrations of 0.2 % or 1 %, in ad libitum feeding. RESULTS: Animals in which tumors had been initiated with DMBA and promoted with TPA experienced significantly fewer tumors and less tumor volume if they ingested either 0.2% or 1% curcumin diets. Also, the dietary consumption of curcumin resulted in a significantly decreased expression of ras and fos proto-oncogenes in the tumorous skin, as measured by enhanced chemiluminesence Western blotting detection system (Amersham). CONCLUSIONS: Whereas earlier work demonstrated that topical application of curcumin to mouse skin inhibited TPA-induced expression of c-fos, c-jun and c-myc oncogenes, our results are the first to show that orally consumed curcumin significantly inhibited DMBA- and TPA-induced ras and fos gene expression in mouse skin. PMID: 11231886 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 208: Cryobiology. 2000 Dec;41(4):301-14. Marked difference in tumor necrosis factor-alpha expression in warm ischemia- and cold ischemia-reperfusion of the rat liver. Lutterova M, Szatmary Z, Kukan M, Kuba D, Vajdova K. Laboratory of Perfused Organs, Slovak Centre for Organ Transplantation, Institute of Preventive and Clinical Medicine, Limbova 14, 83301 Bratislava, Slovakia. Although tumor necrosis factor-alpha has been implicated in liver injury after both warm ischemia- and cold ischemia-reperfusion, it is unclear whether reactivity of the liver to these stimuli is similar with regard to cytokine expression. Here we compare the effects of warm and cold ischemia on tumor necrosis factor-alpha expression and test the hypothesis that cold ischemia preceding warm ischemia causes overexpression of this cytokine. Rat livers were flushed out with University of Wisconsin solution and subjected to varying periods of warm ischemia, cold ischemia, or cold ischemia plus warm ischemia followed by reperfusion using a blood-free perfusion model. Tumor necrosis factor-alpha and interleukin-10 release into the perfusate and bile were measured by ELISA, and expression of these cytokines and that of c-fos, c-jun, and c-myc were studied by reverse-transcriptase polymerase chain reaction. We found high levels of tumor necrosis factor-alpha in the perfusates of livers subjected to warm ischemia-reperfusion, whereas minimal or no tumor necrosis factor-alpha was detected in livers subjected to cold ischemia-reperfusion or to cold ischemia plus warm ischemia-reperfusion. Reverse-transcriptase polymerase chain reaction confirmed the above findings and showed that immediate early genes were expressed in reperfused groups of livers. Measurements of cytokine release into bile showed that neither tumor necrosis factor-alpha nor interleukin-10 were upregulated by cold ischemia-reperfusion. The results suggest that (1) warm ischemia- and cold ischemia-reperfusion of rat liver lead to very different outcomes with regard to tumor necrosis factor-alpha expression and (2) cold ischemia preceding warm ischemia prevents upregulation of tumor necrosis factor-alpha. Copyright 2000 Academic Press. PMID: 11222027 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 209: Int J Radiat Biol. 2001 Jan;77(1):31-40. Oncoprotein expression in human breast epithelial cells transformed by high-LET radiation. Calaf G, Hei TK. Center for Radiological Research, College of Physicians and Surgeons of Columbia Unviersity, New York, NY 10032, USA. gmc24@columbia.edu PURPOSE: The aim of the present work was to analyze the expression of oncoproteins that are frequently altered in breast cancer with specific phenotypic stages in the neoplastic process. MATERIALS AND METHODS: Expression of c-myc, c-jun, c-Ha-ras and the tumor suppressor gene p53 oncoproteins were examined by immunohistochemical staining coupled with confocal microscopy in transformed and tumorigenic human breast epithelial cells induced by high-LET alpha-particles (150 kcV/microm). RESULTS: MCF-10F cells, irradiated with single and double doses of 60 cGy alpha-particles and subsequently treated with cstrogen, showed gradual phenotypic changes including altered morphology, increased cell proliferation relative to control, anchorage-independent growth, invasive capabilities and tumorigenicity in nude mice. MCF-10F cells irradiated with a second dose of 60 cGy alpha-particles after estrogen treatment (60 cGy+ E/60 cGy+E) showed tumorigenicity both in SCII) and nude mice. Alterations in the protein expression of several oncogenes including c-myc, c-jun, c-Ha-ras and the tumor suppressor gene p53 were detected in alpha-particle-irradiated cells and in those cells subsequently cultured in the presence of estrogen. The expression level of these oncoproteins correlated with the progressive nature of the neoplastic process. CONCLUSION: These studies suggest that overexpression of several oncoproteins is important in the neoplastic transformation of human breast epithelial cells induced by high-LET radiation. In addition, use of endocrine factors such as estrogen allows the examination of various aspects of protein expression providing the basis for understanding the complex interactions of hormones and genes. PMID: 11213348 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 210: Cell Mol Life Sci. 1999 Dec;56(11-12):908-17. Proteasomes in apoptosis: villains or guardians? Wojcik C. Department of Histology and Embryology, Institute of Biostructure, Medical University of Warsaw, Warszawa, Poland. cwojcik@ib.amwaw.edu.pl The proteasome (multicatalytic proteinase complex, prosome) is a major cytoplasmic proteolytic enzyme, responsible for degradation of the vast majority of intracellular proteins. Proteins degraded by the proteasome are usually tagged with multiple ubiquitin moieties, conjugated to the substrates by a complicated cascade of enzymes. Over the last years, evidence has accumulated that changes in the expression and activity of the different components of the ubiquitin-proteasome system occur during apoptosis. Proteasome inhibitors have been used to induce apoptosis in various cell types, whereas in others, these compounds were able to prevent apoptosis induced by different stimuli. The proteasome mediated step(s) in apoptosis is located upstream of mitochondrial changes and caspase activation, and can involve in different systems Bcl-2, Jun N-terminal kinase, heat shock proteins, Myc, p53, polyamines and other factors. Publication Types: Review Review, Tutorial PMID: 11212325 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 211: Carcinogenesis. 2001 Feb;22(2):315-20. Erratum in: Carcinogenesis 2001 Apr;22(4):685. beta-Catenin mutation in rat colon tumors initiated by 1,2-dimethylhydrazine and 2-amino-3-methylimidazo[4,5-f]quinoline, and the effect of post-initiation treatment with chlorophyllin and indole-3-carbinol. Blum CA, Xu M, Orner GA, Fong AT, Bailey GS, Stoner GD, Horio DT, Dashwood RH. Linus Pauling Institute and Department of Environment and Molecular Toxicology, Oregon State University, Corvallis, OR 97331-6512, USA. Carcinogens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 1,2-dimethylhydrazine (DMH) induce colon tumors in the rat that contain mutations in beta-catenin, but the pattern of mutation differs from that found in human colon cancers. In both species, mutations affect the glycogen synthase kinase-3beta consensus region of beta-catenin, but whereas they directly substitute critical Ser/Thr phosphorylation sites in human colon cancers, the majority of mutations cluster around Ser33 in the rat tumors. Two dietary phytochemicals, chlorophyllin and indole-3-carbinol, given post-initiation, shifted the pattern of beta-catenin mutations in rat colon tumors induced by IQ and DMH. Specifically, 17/39 (44%) of the beta-catenin mutations in groups given carcinogen plus modulator were in codons 37, 41 and 45, and substituted critical Ser/Thr residues directly, as seen in human colon cancers. None of the tumors from groups given carcinogen alone had mutations in these codons. Interestingly, many of the mutations that substituted critical Ser/Thr residues in beta-catenin were from a single group given DMH and 0.001% chlorophyllin, in which a statistically significant increase in colon tumor multiplicity was observed compared with the group given DMH only. These tumors had marked over-expression of cyclin D1, c-myc and c-jun mRNA and c-Myc and c-Jun proteins were strongly elevated compared with tumors containing wild-type beta-catenin. The results indicate that the pattern of beta-catenin mutations in rat colon tumors can be influenced by exposure to dietary phytochemicals administered post-initiation, and that the mechanism might involve the altered expression of beta-catenin/Tcf/Lef target genes. PMID: 11181454 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 212: Horm Res. 2000;53(5):251-5. Antiproliferative effect and cell cycle modulation by melatonin on GH(3) cells. Fornas O, Mato ME, Webb SM. Experimental Endocrinology Laboratory, Department of Endocrinology, Research Institute, Hospital de Sant Pau, Autonomous University of Barcelona, Spain. We have investigated the effect of melatonin on cell proliferation and modulation, in the GH(3) experimental rat pituitary cell line; the expression of oncogenes c-myc, c-jun and the tumor suppressor gene p53 were also analyzed basally and after exposure to melatonin (10(-6), 10(-8) and 10(-10) M). Melatonin exhibited an antiproliferative effect at all the doses tested, decreasing the proliferating index by 50%. After exposure to melatonin, a decrease in Ki67 and Proliferation cell nuclear antigen occurred acute- and transiently (at 2 h) after a single dose which recovered at 4 h, as well as chronically after repeated 12-hour doses which persisted at 48 h; a similar behavior was observed both acute- and chronically for c-myc and c-jun, while it was opposite for p53, rising acute- and transiently as well as after repeated exposure. These results demonstrate that melatonin modulates the proliferation mechanisms of the GH(3) cells. Copyright 2000 S. Karger AG, Basel PMID: 11150887 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 213: Cancer. 2001 Jan 1;91(1):106-12. Expression of c-Myc, c-Fos, and c-jun in hepatocellular carcinoma. Yuen MF, Wu PC, Lai VC, Lau JY, Lai CL. Department of Medicine, The University of Hong Kong, Queen Mary Hospital, Hong Kong, People's Republic of China. BACKGROUND: Increased expression of the proto-oncogene c-myc is a common phenomenon in hepatocellular carcinoma (HCC). The proto-oncogenes c-fos and c-jun are involved in cell cycle progression and cellular proliferation. METHODS: The objective of this study was to elucidate the mechanism of hepatocarcinogenesis with regard to the expressions of c-myc, c-fos, and c-jun. One hundred fifty biopsied HCC specimens were stained immunohistochemically for the above phenotypic markers both in tumor tissue and in adjacent nontumor tissue. RESULTS: Although the expression of c-myc was high (74%) in tumor tissue, it was significantly less compared with the expression in nontumor tissue (100%; P = 0.0002). The expression of c-myc was inversely proportional to the grade of differentiation in tumor tissue (P = 0.0108; correlation coefficient [r] = -0.244); that is, tissue with poorer histologic differentiation had a lower level of c-myc expression. There were inverse associations between the expression of c-myc and the expression of mutated p53 (P = 0.0017; r = -0.285) as well as the expression of Ki67 (P = 0.057; r = -0.147). There was significantly high expression of c-fos in tumor tissue compared with the expression in nontumor tissue (91% vs. 0%; P < 0.0001). Both the tumor tissue and the nontumor tissue had high levels of expression of c-jun (96.53% and 100%, respectively). There was a trend toward a positive association between the expression of c-fos and the expression of c-jun in tumor tissue (P = 0.07; r = 0.162). CONCLUSIONS: Because c-myc is a known inducer of wild type p53, decreased c-myc expression may lead to uncontrolled cell growth because of the lack of p53 expression that normally induces apoptosis. The coordinated expression of c-fos and c-jun in HCC may reflect the coordinated tumor cell cycle of progression and proliferation; however, future studies are required to elucidate this possibility. Copyright 2001 American Cancer Society. PMID: 11148566 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 214: Toxicol Appl Pharmacol. 2000 Dec 15;169(3):205-21. Morphologic analysis correlates with gene expression changes in cultured F344 rat mesothelial cells. Crosby LM, Hyder KS, DeAngelo AB, Kepler TB, Gaskill B, Benavides GR, Yoon L, Morgan KT. Curriculum in Toxicology, University of North Carolina at Chapel Hill, USA. The gene expression pattern of mesothelial cells in vitro was determined after 4 or 12 h exposure to the rat mesothelial, kidney, and thyroid carcinogen and oxidative stressor potassium bromate (KBrO(3)). Gene expression changes observed using cDNA arrays indicated oxidative stress, mitotic arrest, and apoptosis in treated immortalized rat peritoneal mesothelial cells. Increases occurred in oxidative stress responsive genes HO-1, QR, HSP70, GADD45, GADD153, p21(WAF1/CIP16), GST's, GAPDH, TPX, and GPX-1(0); transcriptional regulators c-jun, c-fos, jun B, c-myc, and IkappaB; protein repair components Rdelta, RC10-II, C3, RC-7, HR6B ubiquitin-conjugating enzyme and ubiquitin; DNA repair components PCNA, msh2, and O-6 methylguanine DNA methyltransferase; lipid peroxide excision enzyme PLA2; and apoptogenic components TNFalpha, iNOS1 and FasL. Decreases occurred in bcl-2 (antiapoptotic), bax alpha, bad, and bok (proapoptotic) and cell cycle control elements (cyclins). Cyclin G and p14ink4b (which inhibit entry into cell cycle) were increased. Numerous signal transduction, cell membrane transport, membrane-associated receptor, and fatty acid biosynthesis and repair components were altered. Morphologic endpoints examined were number of mitotic figures, number of apoptotic cells, and antibody-specific localization of HO-1 (which demonstrated increased HO-1 protein expression). PCR analysis confirmed HO-1, p21(waf1/cip1), HSP70, GPX1, GADD45, QR, mdr1, PGHS, and cyclin D1 changes. A model for KBrO(3)-induced carcinogenicity in the F344 rat mesothelium is proposed, whereby KBrO(3) generates a redox signal that activates p53 and results in transcriptional activation of oxidative stress and repair genes, dysregulation of growth control, and imperfect DNA repair leading to carcinogenesis. Copyright 2000 Academic Press. PMID: 11133343 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 215: Anticancer Res. 2000 Sep- Oct;20(5B):3449-58. Cellular predictive factors for the drug response of lung cancer. Volm M, Rittgen W. Department of Oncological Diagnostics and Therapy, German Cancer Research Center, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany. The main objective of this study to analyze which of 31 cellular factors (resistance proteins, proliferative factors, apoptotic factors, angiogenic factors, proto-oncogenes) most accurately predict the resistance of non-small cell lung carcinomas. To this purpose, we used a short-term in vitro test that measures changes in the rate at which radioactive nucleic acid precursors are incorporated into tumor cells after the addition of doxorubicin to determine the response to doxorubicin in 94 non-small cell lung carcinomas. The results obtained by the short-term test were related to the various cellular factors which were in turn determined by immunohistochemistry and flow cytometry. A significant correlation was found between the data obtained by the short-term test and the expression of P-glycoprotein 170 (P = 0.00004), glutathione-S-transferase-pi (P = 0.0002), metallothionein (P = 0.0008), thymidylate synthase (P = 0.002), O6-methylguanine-DNA-methyltransferase (P = 0.008) and lung resistance-related protein (LRP, P = 0.03). There was only a weak correlation between heat shock proteins (HSP70) and no correlation between the expression of topoisomerase II or catalase and the short-term test results. To measure the proliferative activity, the following were determined: PCNA, cyclin A, cyclin D and cdk2. Only a weak relationship was found between the expression of cdk2 (P = 0.04) and PCNA (P = 0.05) and the doxorubicin response in vitro. Of the investigated pro-apoptotic factors (Fas/CD95, Fas ligand, caspase-3), only Fas/CD95 is significantly associated with the drug response (P = 0.007). The apoptotic index also reveals a significant correlation (P = 0.03). Angiogenesis, as measured by the microvessel density and the angiogenic factors, is inversely correlated to the resistance of non-small cell lung cancer. Platelet-derived endothelial cell growth factor (PD-ECGF) and vascular endothelial growth factor (VEGF) exhibit a significant relationship to the drug resistance (P = 0.0006 and P = 0.004, respectively). Of the investigated proto-oncogenes (Fos, Jun, ErbB-1, ErbB-2, Myc, Ras), only ErbB-2 is weakly associated with the in vitro short term test. In order to determine whether combining factors can result in improved predictive information, combinations of the factors (pairs, triplets) were analyzed. The systematic investigation of these combinations yields an improvement in the predictive information. With one factor up to 76.6% of the tumors, with two factors up to 85.4% and with three factors up to 89.5% of the tumors could be correctly diagnosed. PMID: 11131647 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 216: Oncogene. 2000 Nov 30;19(51):5906-18. v-Jun sensitizes cells to apoptosis by a mechanism involving mitochondrial cytochrome C release. MacLaren A, Clark W, Gillespie DA. Beatson Institute for Cancer Research, Cancer Research Campaign Beatson Laboratories, Glasgow, Scotland, UK. v-Jun shares the ability of the Myc, E1A, and E2F oncogenes to both sustain cell cycle progression and promote apoptosis in the absence of mitogenic stimulation. To gain an insight into the mechanism of apoptosis sensitization, we examined the possible involvement of key regulatory proteins previously implicated in oncogene-induced cell death during v-Jun-induced apoptosis triggered by serum withdrawal. We observed that ectopic expression of the anti-apoptotic Bcl-2 protein, or of two downstream effectors of growth factor signalling, v-PI 3-Kinase and v-Src, partially or completely suppressed apoptosis. Apoptosis was also observed in the presence of serum growth factors when endogenous PI3K activity was blocked using the synthetic inhibitor LY294002, further suggesting an important role for PI3-K in cell survival. Cytochrome C was released into the cytosol of apoptotic v-Jun expressing cells, and this release was inhibited by Bcl-2, suggesting an important role for mitochondrial dysfunction in v-Jun induced apoptosis. In contrast, inhibition of Fas signalling using dominant negative FADD did not inhibit apoptosis, nor was there any evidence for accumulation or activation of p53 in v-Jun transformed cells. Consistent with this latter observation, inhibition of p53 function by HPV16 E6 protein had no effect on v-Jun induced cell death. Taken together, these results suggest that mitochondrial dysfunction is an important component of the mechanism through which v-Jun sensitizes cells to apoptosis, but that the apoptotic signals elicited by v-Jun upstream of the mitochondria do not depend on increased levels of p53 activity or Fas signalling. PMID: 11127822 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 217: Exp Mol Pathol. 2000 Dec;69(3):202-10. Dexamethasone enhances mallory body formation in drug-primed mouse liver. Yuan QX, Nagao Y, French BA, Wan YJ, French SW. Department of Pathology, Harbor-UCLA Medical Center, Torrance, California 90509, USA. In a clinical study in which patients with alcoholic hepatitis were treated with prednisone for 1 month, posttreatment liver biopsies showed diminished inflammation, but Mallory bodies were not diminished. This suggested that steroid treatment may reduce inflammation by inhibiting NFkappaB activation. Sparing of Mallory bodies suggests that NFkappaB activation may not be involved mechanistically in Mallory body formation. To test this idea, we induced Mallory body formation in drug-primed mice with or without dexamethasone treatment. As predicted, dexamethasone decreased NFkappaB activation; however, Mallory body formation was increased. Surprisingly, TNFalpha and iNOS, which normally increase as a result of NFkappaB activation, were upregulated by the dexamethasone treatment. It was concluded that NFkappaB activation is not involved in Mallory body formation. Despite this, induced increases in TNFalpha, iNOS, c-jun/API and c-myc expression indicate that oxidative stress is likely involved in Mallory body formation. Copyright 2000 Academic Press. PMID: 11115361 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 218: J Exp Zool. 2000 Dec 1;287(7):477-84. Mitogen-activated protein kinases and anoxia tolerance in turtles. Greenway SC, Storey KB. Faculty of Medicine, University of Manitoba, Winnipeg, Canada. The response of two vertebrate mitogen-activated protein kinase (MAPK) family members, the extracellular signal-regulated kinases (ERKs) and c-Jun NH2-terminal kinases (JNKs), to anoxia exposure in vivo was examined in organs (liver, heart, kidney, brain, spleen) of the anoxia-tolerant adult turtle, Trachemys scripta elegans. ERKs activities rose during anoxia only in spleen (a 2.8-fold increase). JNK activity showed a significant increase only in liver (4-fold increase) after 5 hr of anoxic submergence but declined thereafter. Levels of the transcription factor c-Fos were strongly suppressed in liver, heart, and kidney of anoxia-exposed turtles, whereas levels increased 2-fold in anoxic brain. The effect of anoxia on c-Myc was organ-specific and variable with 2- and 1.5-fold increases in protein expression in kidney and brain, respectively, and a 60% decrease in anoxic spleen. These results for an anoxia-tolerant animal suggest the potential importance of the MAPKs and of the immediate-early genes (c-fos, c-myc) in mediating adaptive responses to oxygen deprivation. PMID: 11110161 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 219: Cancer Res. 2000 Nov 15;60(22):6276-80. Activation of the transcription factor Oct-1 in response to DNA damage. Zhao H, Jin S, Fan F, Fan W, Tong T, Zhan Q. Department of Radiation Oncology, Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pennsylvania 15213, USA. Mammalian cells exhibit complex cellular responses to genotoxic stress, including cell cycle checkpoint, DNA repair, and apoptosis. Inactivation of these important biological events will result in genomic instability and cell transformation. It has been demonstrated that gene activation is a critical initial step during the cellular response to DNA damage. A number of investigations have shown that transcription factors are involved in the regulation of stress-inducible genes. These transcription factors include p53, c-Myc, and AP-1 (c-fos and c-jun). However, the role for the octamer-binding transcription factor Oct-1 in the DNA damage-activated response is unknown. In this report, we have presented the novel observation that the transcription factor Oct-1 is induced after cells are exposed to multiple DNA-damaging agents and therapeutic agents, including UV radiation, methylmethane sulfonate, ionizing radiation, etoposide, cisplatin, and camptothecin. The induction of the Oct-1 protein is mediated through a posttranscriptional mechanism and does not require the normal cellular function of the tumor suppressor p53, indicating that the Oct-1 protein, as a transcription factor, may play a role in p53-independent gene activation. In addition to increased protein level, the activity of Oct-1 DNA binding to its specific consensus sequence is also enhanced by DNA damage. Therefore, these results have implicated that the transcription factor Oct-1 might participate in cellular response to DNA damage, particularly in p53-independent gene activation. PMID: 11103783 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 220: J Biol Chem. 2001 Feb 23;276(8):6030-6. Epub 2000 Nov 28. Glutamine-dependent antiapoptotic interaction of human glutaminyl-tRNA synthetase with apoptosis signal-regulating kinase 1. Ko YG, Kim EY, Kim T, Park H, Park HS, Choi EJ, Kim S. National Creative Research Initiatives Center for ARS Network, Sung Kyun Kwan University, Suwon, Kyunggido 440-746, Korea. Glutamine has been known to be an apoptosis suppressor, since it blocks apoptosis induced by heat shock, irradiation, and c-Myc overexpression. Here, we demonstrated that HeLa cells were susceptible to Fas-mediated apoptosis under the condition of glutamine deprivation. Fas ligation activated apoptosis signal-regulating kinase 1 (ASK1) and c-Jun N-terminal kinase (JNK; also known as stress-activated protein kinase (SAPK)) in Gln-deprived cells but not in normal cells, suggesting that Gln might be involved in the activity control of ASK1 and JNK/SAPK. As one of the possible mechanisms for the suppressive effect of Gln on ASK1, we investigated the molecular interaction between human glutaminyl-tRNA synthetase (QRS) and ASK1 and found the Gln-dependent association of the two molecules. While their association was enhanced by the elevation of Gln concentration, they were dissociated by Fas ligation within 5 min. The association involved the catalytic domains of the two enzymes. The ASK1 activity was inhibited by the interaction with QRS as determined by in vitro kinase and transcription assays. Finally, we have shown that QRS inhibited the cell death induced by ASK1, and this antiapoptotic function of QRS was weakened by the deprivation of Gln. Thus, the antiapoptotic interaction of QRS with ASK1 is controlled positively by the cellular concentration of Gln and negatively by Fas ligation. The results of this work provide one possible explanation for the working mechanism of the antiapoptotic activity of Gln and suggest a novel function of mammalian ARSs. PMID: 11096076 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 221: Mol Cell Neurosci. 2000 Oct;16(4):453-69. AP-1 activity during the growth, differentiation, and death of O-2A lineage cells. FitzGerald UF, Barnett SC. Department of Neurology, University of Glasgow, United Kingdom. Oligodendrocyte differentiation has been correlated with AP-1 activity, being low in progenitors and high in differentiated cells. In this study we have carried out a detailed temporal analysis of AP-1 activity in oligodendrocyte-type-2 astrocyte (O-2A) lineage cells. We show that low AP-1 activity in progenitor cells depended on the application of growth factors. Treatment of cells with B104-conditioned medium induced high AP-1 activity, increased process length, and improved growth. The role of AP-1 in proliferation and process extension was emphasized when progenitor cells overexpressing a c-Jun dominant-negative mutant had impaired growth and shortened processes. AP-1 DNA-binding activity during O-2A differentiation in vitro showed an initial down-regulation followed by up-regulation after 2 days. Increased AP-1 levels in oligodendrocytes were inhibited by overexpression of bcl-2, indicating that AP-1 in mature oligodendrocytes is involved in the regulation of apoptosis. Prevention of cell death by bcl-2 in oligodendrocytes was accompanied by progressive differentiation and expression of MOG and PLP. PMID: 11085881 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 222: J Immunol. 2000 Oct 15;165(8):4312-8. IL-2R beta agonist P1-30 acts in synergy with IL-2, IL-4, IL-9, and IL-15: biological and molecular effects. Eckenberg R, Moreau JL, Melnyk O, Theze J. Unite d'Immunogenetique Cellulaire, Institut Pasteur, Paris, France. From the sequence of human IL-2 we have recently characterized a peptide (p1-30), which is the first IL-2 mimetic described. P1-30 covers the entire alpha helix A of IL-2 and spontaneously folds into a alpha helical homotetramer mimicking the quaternary structure of a hemopoietin. This neocytokine interacts with a previously undescribed dimeric form of the human IL-2 receptor beta-chain likely to form the p1-30 receptor (p1-30R). P1-30 acts as a specific IL-2Rbeta agonist, selectively inducing activation of CD8 and NK lymphocytes. From human PBMC we have also shown that p1-30 induces the activation of lymphokine-activated killer cells and the production of IFN-gamma. Here we demonstrate the ability of p1-30 to act in synergy with IL-2, -4, -9, and -15. These synergistic effects were analyzed at the functional level by using TS1beta, a murine T cell line endogenously expressing the common cytokine gamma gene and transfected with the human IL-2Rbeta gene. At the receptor level, we show that expression of human IL-2Rbeta is absolutely required to obtain synergistic effects, whereas IL-2Ralpha specifically impedes the synergistic effects obtained with IL-2. The results suggest that overexpression of IL-2Ralpha inhibits p1-30R formation in the presence of IL-2. Finally, concerning the molecular effects, although p1-30 alone induces the antiapoptotic molecule bcl-2, we show that it does not influence mRNA expression of c-myc, c-jun, and c-fos oncogenes. In contrast, p1-30 enhances IL-2-driven expression of these oncogenes. Our data suggest that p1-30R (IL-2Rbeta)(2) and intermediate affinity IL-2R (IL-2Rbetagamma), when simultaneously expressed at the cell surface, may induce complementary signal transduction pathways and act in synergy. PMID: 11035066 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 223: EMBO J. 2000 Oct 2;19(19):5114-22. p56(dok-2) as a cytokine-inducible inhibitor of cell proliferation and signal transduction. Suzu S, Tanaka-Douzono M, Nomaguchi K, Yamada M, Hayasawa H, Kimura F, Motoyoshi K. Biochemical Research Laboratory, Morinaga Milk Industry Co. Ltd, Higashihara, Zama-city, Kanagawa 228-8583, Japan. p56(dok-2) acts as a multiple docking protein downstream of receptor or non-receptor tyrosine kinases. However, the role of p56(dok-2) in biological functions of cells is not clear. We found that transcription of the p56(dok-2) gene in macrophages was increased markedly in response to cytokines such as macrophage colony-stimulating factor (M-CSF), granulocyte/macrophage-CSF and interleukin-3 (IL-3). Forced expression of p56(dok-2) inhibited M-CSF-, granulocyte-CSF-, IL-3- and stem cell factor-induced proliferation of myeloid leukemia cells, M-NFS-60. The p56(dok-2)-overexpressing cells showed an impaired induction of c-myc but not of c-jun, junB or c-fos when stimulated with M-CSF. Consistent with these results, the peritoneal cavity of the hairless (hr/hr) strain of mutant mice, whose cells expressed less p56(dok-2) than wild-type mice, contained more macrophages than that of +/hr mice. Moreover, the inhibition of endogenous p56(dok-2) expression in macrophage-like tumor cells, J774A.1, by stable expression of antisense p56(dok-2) mRNA accelerated cell proliferation. The study identifies a novel role for p56(dok-2) as a molecule that negatively regulates signal transduction and cell proliferation mediated by cytokines in a feedback loop. PMID: 11013214 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 224: Braz J Med Biol Res. 2000 Oct;33(10):1133-40. Proliferative signaling initiated in ACTH receptors. Lotfi CF, Lepique AP, Forti FL, Schwindt TT, Eichler CB, Santos MO, Rebustini IT, Hajj GN, Juliano L, Armelin HA. Departamento de Bioquimica, Instituto de Quimica, Universidade de Sao Paulo, Brasil. This article reviews recent results of studies aiming to elucidate modes of integrating signals initiated in ACTH receptors and FGF2 receptors, within the network system of signal transduction found in Y1 adrenocortical cells. These modes of signal integration should be central to the mechanisms underlying the regulation of the G0-->G1-->S transition in the adrenal cell cycle. FGF2 elicits a strong mitogenic response in G0/G1-arrested Y1 adrenocortical cells, that includes a) rapid and transient activation of extracellular signal-regulated kinases-mitogen-activated protein kinases (ERK-MAPK) (2 to 10 min), b) transcription activation of c-fos, c-jun and c-myc genes (10 to 30 min), c) induction of c-Fos and c-Myc proteins by 1 h and cyclin D1 protein by 5 h, and d) onset of DNA synthesis stimulation within 8 h. ACTH, itself a weak mitogen, interacts with FGF2 in a complex manner, blocking the FGF2 mitogenic response during the early and middle G1 phase, keeping ERK-MAPK activation and c-Fos and cyclin D1 induction at maximal levels, but post-transcriptionally inhibiting c-Myc expression. c-Fos and c-Jun proteins are mediators in both the strong and the weak mitogenic responses respectively triggered by FGF2 and ACTH. Induction of c-Fos and stimulation of DNA synthesis by ACTH are independent of PKA and are inhibited by the PKC inhibitor GF109203X. In addition, ACTH is a poor activator of ERK-MAPK, but c-Fos induction and DNA synthesis stimulation by ACTH are strongly inhibited by the inhibitor of MEK1 PD98059. Publication Types: Review Review, Tutorial PMID: 11004713 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 225: Oncogene. 2000 Sep 14;19(39):4513-22. Implication of multiple mechanisms in apoptosis induced by the synthetic retinoid CD437 in human prostate carcinoma cells. Sun SY, Yue P, Lotan R. Department of Thoracic/Head and Neck Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas, TX 77030, USA. The synthetic retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) induces apoptosis in several types of cancer cell. CD437 inhibited the growth of both androgen-dependent and -independent human prostate carcinoma (HPC) cells in a concentration-dependent manner by rapid induction of apoptosis. CD437 was more effective in killing androgen-independent HPC cells such as DU145 and PC-3 than the androgen-dependent LNCaP cells. The caspase inhibitors Z-VAD-FMK and Z-DEVD-FMK blocked apoptosis induced by CD437 in DU145 and LNCaP cells, in which increased caspase-3 activity and PARP cleavage were observed, but not in PC-3 cells, in which CD437 did not induce caspase-3 activation and PARP cleavage. Thus, CD437 can induce either caspase-dependent or caspase-independent apoptosis in HPC cells. CD437 increased the expression of c-Myc, c-Jun, c-Fos, and death receptors DR4, DR5 and Fas. CD437's potency in apoptosis induction in the different cell lines was correlated with its effects on the expression of oncogenes and death receptors, thus implicating these genes in CD437-induced apoptosis in HPC cells. However, the importance and contribution of each of these genes in different HPC cell lines may vary. Because CD437 induced the expression of DR4, DR5 and Fas, we examined the effects of combining CD437 and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and Fas ligand, respectively, in HPC cells. We found synergistic induction of apoptosis, highlighting the importance of the modulation of these death receptors in CD437-induced apoptosis in HPC cells. This result also suggests a potential strategy of using CD437 with TRAIL for treatment of HPC. Oncogene (2000) 19, 4513 - 4522. PMID: 11002424 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 226: Exp Parasitol. 2000 Jul;95(3):202-8. Development of brain injury in mice by Angiostrongylus cantonensis infection is associated with the induction of transcription factor NF-kappaB, nuclear protooncogenes, and protein tyrosine phosphorylation. Lee HH, Shiow SJ, Chung HC, Huang CY, Lin CL, Hsu JD, Shyu LY, Wang CJ. Department of Parasitology. Chung SHan Medical and Dental College, Taichung, Taiwan. Eosinophilic meningitis or meningoencephalitis caused by Angiostrongylus cantonensis is endemic to the Pacific area of Asia, especially Taiwan, Thailand, and Japan. Although eosinophilia is an important clinical manifestation of A. cantonensis infection, the role of eosinophils in the progress of the infection remains to be elucidated. In this experiment, we showed that A. cantonensis-caused eosinoplia and inflammation might lead to the induction of NF-kappaB and protooncogene expression via activation of the tyrosine phosphorylation signal pathway. After mice were infected daily with 30 third-stage larvae of A. cantonensis by oral adminstration for 6 weeks, no significant differences PKC-alpha, MEK-1, ERK-2, JNK, and p38 protein expression were found between the control and infected mice. However, the protein tyrosine phosphorylation levels, NF-kappaB, and iNOS protein products were significantly increased by 3.5-, 3.3-, and 6.3-fold, respectively, after 3 weeks of A. cantonensis infection. The same pattern was found for c-Myc, c-Jun, and c-Fos proteins, which were elevated by 3.2-, 2.3-, and 3.4-fold, respectively, compared to control animals after 3 weeks. The expression potency of these proteins started increasing in week 1, reaching maximal induction in week 3, and then declining in week 5 after A. cantonensis infection. Another consistent result was noted in the pathological observations, including eosinophilia, leukocyte infiltration, granulomatous reactions, and time responses in brain tissues of infected mice. These data suggest that the development of brain injury by eosinophlia of A. cantonensis infection is associated with NF-kappaB and/or nuclear protooncogenes expression, which is activated by the tyrosine phosphorylation pathway. Copyright 2000 Academic Press. PMID: 10964648 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 227: Cancer Lett. 2000 Oct 1;158(2):185-93. Hypomethylation and overexpression of c-jun and c-myc protooncogenes and increased DNA methyltransferase activity in dichloroacetic and trichloroacetic acid-promoted mouse liver tumors. Tao L, Yang S, Xie M, Kramer PM, Pereira MA. Department of Pathology, Medical College of Ohio, Health Education Building, 3055 Arlington Ave., Toledo, OH 43614-5806, USA. ltao@mco.edu Dichloroacetic acid (DCA) and trichloroacetic acid (TCA) are mouse liver carcinogens. Methylation of the c-jun and c-myc genes, expression of both genes and DNA methyltransferase (DNA MTase) activity were determined in liver tumors initiated by N-methyl-N-nitrosourea and promoted by DCA and TCA in female B6C3F1 mice. Hypomethylated and over-expression of c-jun and c-myc genes were found in DCA- and TCA-promoted liver tumors. DNA MTase activity was increased in tumors while decreased in non-involved liver. Thus, DCA- and TCA-promoted carcinogenesis appears to include decreased methylation and increased expression of c-jun and c-myc genes in the presence of increased DNA MTase activity. PMID: 10960769 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 228: Anticancer Res. 2000 Jul-Aug;20(4):2441-8. Apoptosis induced by the sodium butyrate in human gastric cancer TMK-1 cells. Tsai LC, Hung MW, Chang GG, Chang TC. Department of Medical Research, Veterans General Hospital, Taipei, Taiwan, R.O.C. The effects of sodium butyrate on cell proliferation, gene expression, and apoptosis were investigated. Upon exposure to sodium butyrate the cells exhibited marked morphological changes, reduced cell proliferation and most cells died through apoptosis within 48 hours. In the presence of dexamethasone, however, the sodium butyrate-triggered apoptosis was markedly reduced. Studies using the glucocorticoid receptor antagonist RU486 indicated that the protective effect of dexamethasone was mediated through glucocorticoid receptor. Sodium butyrate markedly induced the c-jun proteins level, whereas the c-Myc protein was down-regulated rapidly. c-Jun protein may play an important role in the action of sodium butyrate since its induction preceded the onset of DNA fragmentation. In addition, preincubation of the cells with dexamethasone markedly delayed the induction of c-jun levels by sodium butyrate. Analysis of the expression of bel-2-related genes indicated that the Bcl-xS protein level was increased in the presence of sodium butyrate and the up-regulation of Bcl-xS by sodium butyrate was also blocked by dexamethasone. Taken together, these results indicate that c-myc, c-jun and Bcl-xS proteins may be involved in the mechanism of sodium butyrate-triggered apoptosis in these cells. PMID: 10953308 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 229: Biochem Biophys Res Commun. 2000 Aug 18;275(1):1-6. Differential regulation of phospholipase C-beta isozymes in cardiomyocyte hypertrophy. Schnabel P, Mies F, Nohr T, Geisler M, Bohm M. Klinik III fur Innere Medizin, Universitat zu Koln, Joseph-Stelzmann-Strasse 9, Cologne, 50924, Germany. Cardiac hypertrophy is a major predictor of heart failure and of morbidity and mortality in developed countries. Many hormones and growth factors induce cardiac hypertrophy via activation of members of the phospholipase C (PLC) family. The expression pattern of the PLCbeta isozyme subfamily was investigated in neonatal rat cardiomyocytes after stimulation with different hypertrophic stimuli. Under control conditions and after stimulation with norepinephrine, cardiomyocytes expressed similar amounts of PLCbeta3 mRNA. In the presence of fetal calf serum (FCS), additional expression of PLCbeta1 was induced. Growth hormone (GH) and insulin-like growth factor-I (IGF-I) both induced a substantial increase in PLCbeta3 mRNA expression. The response to GH could not be abolished by the IGF-I receptor blocker IGF-I analogue indicating an IGF-I-independent action of GH. The upregulation of PLCbeta3 by IGF-I was abolished by preincubation of cardiomyocytes with the IGF-I receptor antagonist IGF-I analogue, the tyrosine kinase inhibitor genistein, the extracellular signal-related kinase (ERK) inhibitor PD 98059, the phosphatidylinositol-3- (PI-3) kinase inhibitor wortmannin and the p70 S6 kinase inhibitor rapamycin. Induction of the immediate early genes c-myc, c-fos, and c-jun by IGF-I was abolished by preincubation with antisense oligos against PLCbeta3. It is concluded that the expression of PLCbeta isozymes in cardiomyocytes is differentially regulated by different hypertrophic stimuli. The upregulation of PLCbeta3 by IGF-I is dependent on the activity of tyrosine kinase, ERK, PI3 kinase, and p70 S6 kinase and PLCbeta3 expression seems to be required for the induction of immediate early genes by IGF-I. The involvement of the PLCbeta subfamily in signal transduction of receptors other than G-protein-coupled receptors is suggested. Copyright 2000 Academic Press. PMID: 10944430 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 230: Am J Hematol. 2000 Sep;65(1):66-71. A case of monoclonal gammopathy associated with acute myelomonocytic leukemia with eosinophilia suggested to be the result of lineage infidelity. Nagata T, Mugishima H, Yoden A, Yoshikawa K, Oguni T, Yamashiro K, Yamamori S, Harada K. Department of Pediatrics, Nihon University, School of Medicine, Tokyo, Japan. tnagata@med.nihon-u.ac.jp Acute myelomonocytic leukemia (AMMoL) accompanied by monoclonal gammopathy is a rare condition, and its pathogenesis and the cytogenetic mechanism of such leukemogenesis have not been determined in detail. A case of AMMoL with eosinophilia accompanied by immunoglobulin G kappa monoclonal gammopathy is described. Immunophenotypic studies of the peripheral blood and bone marrow mononuclear cells revealed no evidence of abnormally proliferating cells of B-lineage. DNA analyses of bone marrow mononuclear cells containing leukemic cells revealed rearrangement of the kappa-light chain (Igkappa) gene and c-myc and c-jun proto-oncogenes. The intensities of the rearranged bands for these genes on Southern blot analysis suggested the existence of a major population of leukemic cells with rearranged Igkappa gene and minor population(s) of leukemic cells with rearranged c-myc and/or c-jun proto-oncogene(s) in the patient's bone marrow and indicated the occurrence of genetic evolutionary changes in leukemic cells in this patient before starting chemotherapy. These results suggest that these leukemic cells are the most likely candidate for immunoglobulin G kappa monoclonal protein production, and structural abnormalities of c-myc and c-jun proto-oncogenes may have contributed to the evolution of leukemic cells in this patient. Copyright 2000 Wiley-Liss, Inc. Publication Types: Case Reports PMID: 10936867 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 231: Breast Cancer Res Treat. 2000 May;61(1):69-78. Ectopic expression of Rsu-1 results in elevation of p21CIP and inhibits anchorage-independent growth of MCF7 breast cancer cells. Vasaturo F, Dougherty GW, Cutler ML. Department of Pathology, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA. Signal transduction from tyrosine kinase receptors mediates growth regulation of breast cancer cells in part through the GTPase Ras and downstream kinases. Rsu-1 is a cDNA previously identified as an inhibitor of Ras-induced transformation. An HA-epitope tagged Rsu-1 cDNA was introduced into the MCF7 breast carcinoma cell line. Stable transfectants were selected and used for analysis of Rsu-1 expression on growth control and Ras-dependent kinase pathways. Assessment of biological activity of HA-Rsu-1 transfectants revealed that HA-Rsu-1 clones showed slower anchorage dependent growth rates than control MCF7 cell lines and a significant reduction in anchorage independent growth. Analysis of cell cycle regulatory proteins required for transit through G1 revealed that HA-Rsu-1 transfectant cell lines expressed elevated levels of p21CIP CDK inhibitor. Perturbations in signal transduction pathways which can be activated by Ras were detected in the Ha-Rsu-1 transfectants. Exposure of serum-starved cells to EGF revealed that expression of HA-Rsu-1 increased ERK-2 kinase activation, decreased activation of Jun kinase and inhibited Rho-dependent Rho-alpha kinase (ROK) activity compared to control cells. While serum starvation reduced AKT activity to undetectable levels in HA-Rsu-1 transfectants but not in control MCF7 cells, activation of AKT kinase by serum was unaffected by HA-Rsu-1 expression. Finally, the level of c-myc transcription in HA-Rsu-1 transfectants reached only 60% of the MCF7 control cell line following serum stimulation of starved cells while Fos RNA levels were similar to control cells. These results demonstrate that increased Rsu-1 expression critically altered cell cycle regulation and growth of MCF7 cells as well as signaling pathways in MCF7 cells required for malignant growth. PMID: 10930091 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 232: J Endocrinol. 2000 Aug;166(2):329-38. Stabilization of cyclin D1 mRNA via the phosphatidylinositol 3-kinase pathway in MCF-7 human breast cancer cells. Dufourny B, van Teeffelen HA, Hamelers IH, Sussenbach JS, Steenbergh PH. Utrecht Graduate School of Developmental Biology, Laboratory for Physiological Chemistry, Utrecht University, PO Box 80042, 3508 TA Utrecht, The Netherlands. Treatment of quiescent MCF-7 human breast cancer cells with either the polypeptide growth factors insulin-like growth factor-I (IGF-I) or epidermal growth factor (EGF), the steroid hormone estradiol (E2) or the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) results in increased steady-state levels of cyclin D1 mRNA and protein. Unexpectedly, this elevation of cyclin D1 expression by all of these agents is inhibited by the specific phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002. Since transcriptional activation of the cyclin D1 promoter by EGF, E2 and TPA is independent of PI3-K activity, these findings suggest a post-transcriptional role for PI3-K in the regulation of cyclin D1 expression. Here we show that inhibition of PI3-K by LY294002 decreases the half-life of the 4.5 kb cyclin D1 mRNA species. In contrast, the stability of the 1.5 kb cyclin D1 mRNA is not affected by PI3-K inhibition. PI3-K-mediated stabilization of mRNA is not a general phenomenon, since other rapidly regulated and unstable mRNAs, such as those encoding c-fos, c-jun and c-myc, are not stabilized upon activation of the PI3-K signaling pathway. PMID: 10927622 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 233: Kidney Int. 2000 Aug;58(2):638-46. Phospholipase A2 mediates immediate early genes in cultured renal epithelial cells: possible role of lysophospholipid. Kohjimoto Y, Honeyman TW, Jonassen J, Gravel K, Kennington L, Scheid CR. Department of Physiology, University of Massachusetts Medical School, Worcester 01655-0127, USA. BACKGROUND: Exposure to high levels of oxalate induces oxidant stress in renal epithelial cells and produces diverse changes in cell function, ranging from cell death to cellular adaptation, as evidenced by increased DNA synthesis, cellular proliferation, and induction of genes associated with remodeling and repair. These studies focused on cellular adaptation to this oxidant stress, examining the manner by which oxalate exposure leads to increased expression of immediate early genes (IEGs). Specifically, our studies assessed the possibility that oxalate-induced changes in IEG expression are mediated by phospholipase A2 (PLA2), a common pathway in cellular stress responses. METHODS: Madin-Darby canine kidney (MDCK) cells were exposed to oxalate in the presence or absence of PLA2 inhibitors: mepacrine and arachidonyl trifluoromethyl ketone (AACOCF3). Expression of IEG (c-jun, egr-1, and c-myc) mRNA was assessed by Northern blot analysis. PLA2 activity was determined by measuring the release of [3H]arachidonic acid (AA) from prelabeled cells. RESULTS: Oxalate exposure (1 to 1.5 mmol/L) induced time- and concentration-dependent increases in IEG mRNA. Treatment with mepacrine resulted in a 75 to 113% reduction of oxalate-induced c-jun, egr-1, and c-myc mRNA, while AACOCF3 caused a 41 to 46% reduction of oxalate-induced c-jun and egr-1 mRNA. Of the two major byproducts of PLA2, only lysophosphatidylcholine (20 micromol/L) increased c-jun and egr-1 mRNA. In contrast, AA (25 micromol/L) attenuated the oxalate-induced increase in c-jun and egr-1 mRNA, presumably by inhibiting PLA2 activity. CONCLUSIONS: These findings suggest that PLA2 plays a major role in oxalate-induced IEG expression in renal epithelial cells and that lysophospholipids might be a possible lipid mediator in this pathway. PMID: 10916087 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 234: Ann N Y Acad Sci. 2000 Jun;910:21-33; discussion 33-5. Control of beta-catenin signaling in tumor development. Behrens J. Max-Delbruck-Center for Molecular Medicine, Berlin, Germany. jbehren@mdc-berlin.de The wnt signal transduction pathway is involved in various differentiation events during embryonic development and leads to tumor formation when aberrantly activated. The wnt signal is transmitted to the nucleus by the cytoplasmic component beta-catenin: in the absence of wnts, beta-catenin is constitutively degraded in proteasomes, whereas in the presence of wnts beta-catenin is stabilized and can associate with HMG box transcription factors of the LEF/TCF family. The LEF/TCF/beta-catenin complexes activate specific wnt target genes. In tumors, beta-catenin degradation is blocked by mutations of beta-catenin or of the tumor suppressor gene product APC. As a consequence, beta-catenin is stabilized, constitutive complexes with LEF/TCF factors are formed, and oncogenic target genes, such as c-myc, cyclin D1, and c-jun, are activated. Thus, control of beta-catenin is a major regulatory event in normal wnt signaling and during tumor formation. It has been found that a multiprotein complex assembled by the cytoplasmic component conductin induces degradation of cytoplasmic beta-catenin. The complex includes APC, the serine/threonine kinase GSK3 beta, and beta-catenin, which bind to conductin at distinct domains. In colon carcinoma cells, forced expression of conductin downregulates beta-catenin, whereas in normal cells mutants of conductin that are deficient in complex formation stabilize beta-catenin. Fragments of APC that contain a conductin-binding domain also block beta-catenin degradation. In Xenopus embryos, conductin inhibits the wnt pathway. In situ hybridization analysis shows that conductin is expressed in various embryonal tissues known to be regulated by wnts, such as the developing brain, mesenchyme below the epidermis, lung mesenchyme, and kidney. It is suggested that conductin controls wnt signaling by assembling the essential components of the beta-catenin degradation pathway. Alterations of conductin function may lead to tumor formation. Publication Types: Review Review, Tutorial PMID: 10911903 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 235: Cell Immunol. 2000 Jun 15;202(2):124-35. STAT3 regulates the growth and immunoglobulin production of BCL(1) B cell lymphoma through control of cell cycle progression. Karras JG, McKay RA, Lu T, Pych J, Frank DA, Rothstein TL, Monia BP. Department of Molecular and Cellular Pharmacology, Isis Pharmaceuticals, Inc., 2292 Faraday Avenue, Carlsbad, California 92008, USA. STAT3 is constitutively phosphorylated on tyrosine(705) in self-renewing, CD5(+) murine B-1 lymphocytes. Nuclear extracts from untreated primary B-1 or CD5(+) BCL(1) B lymphoma cells were found to contain immunoreactive STAT3 protein that binds to a sis-inducible element present in the promoter of the p21(waf1/cip1) tumor suppressor gene and is constitutively phosphorylated on serine(727). To determine the functional significance of constitutive STAT3 activation in B lymphoma cells, a specific STAT3 antisense oligonucleotide was developed and used to examine basal BCL(1) cell growth and IgM production. Abrogating STAT3 expression in BCL(1) cells inhibited their proliferative capacity and induced a corresponding decrease in secretion of IgM. Cell cycle analysis showed a block in progression through G1 in BCL(1) cells treated with the STAT3 antisense oligonucleotide. These results indicate that STAT3 controls cell growth and immunoglobulin secretion by enhancing progression through the G1 phase of the cell cycle in BCL(1) B cell lymphoma. Copyright 2000 Academic Press. PMID: 10896772 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 236: Anat Embryol (Berl). 2000 May;201(5):349-55. Expression of proto-oncogenes in bovine preimplantation blastocysts. Tetens F, Kliem A, Tscheudschilsuren G, Navarrete Santos A, Fischer B. Department of Anatomy and Cell Biology, Martin Luther University Faculty of Medicine, Halle (Saale), Germany. Proto-oncogenes are involved in the regulation of gene expression, for example after ligand binding to growth factor receptors. Expression of the proto-oncogenes c-fos, c-jun, c-ha-ras and c-myc was studied in in vivo grown and in vitro cultured bovine preimplantation blastocysts employing RT-PCR, ribonuclease protection assay and immunohistochemistry. Thirteen- and 14- day-old preimplantation blastocysts, i.e. stages before and during trophoblast elongation, were used. In in vivo-grown blastocysts c-fos, c-jun and c-ha-ras transcripts as well as c-Fos, c-Jun and c-Myc proteins were detected in all stages studied. Cultured blastocysts were treated with 10 nM epidermal growth factor and 10 nM transforming growth factor-alpha simultaneously. Epidermal growth factor and transforming growth factor-alpha treatment induced c-fos mRNA and c-Myc protein expression. The induction of downstream targets of the epidermal growth factor receptor by epidermal growth factor and transforming growth factor-alpha indicates a functional epidermal growth factor signal transduction pathway in elongating bovine blastocysts. PMID: 10839631 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 237: J Biol Chem. 2000 Sep 1;275(35):27377-85. The interferon- and differentiation-inducible p202a protein inhibits the transcriptional activity of c-Myc by blocking its association with Max. Wang H, Liu C, Lu Y, Chatterjee G, Ma XY, Eisenman RN, Lengyel P. Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA. p202a is a murine protein that is induced during the fusion of myoblasts to myotubes and can also be induced by interferon. Even 2-3-fold overexpression of p202a in cells retards proliferation. p202a was shown to modulate transcription by binding, and inhibiting the activity of several transcription factors including c-Fos, c-Jun, AP-2, E2F1, E2F4, NF-kappaB, MyoD, and myogenin. Here we report that p202a also bound the c-Myc protein in vitro and in vivo; the C-terminal p202a b segment bound the C-terminal basic region helix-loop-helix-leucine zipper (bHLHLZ) region of c-Myc. The transfection of a p202a expression plasmid inhibited the c-Myc-dependent expression of reporter plasmids in transient assays; moreover, overexpression of p202a in stable cell lines decreased the endogenous levels of mRNAs whose expression is driven by c-Myc. These effects of p202a are consistent with our finding that the binding of p202a to c-Myc inhibited the binding of c-Myc to Max in vitro and in vivo. p202a also inhibited the c-Myc-induced anchorage-independent growth and apoptosis of Rat-1 cells. The inhibition of c-Myc-dependent transcription, proliferation, and apoptosis by p202a is in line with the involvement of p202a in differentiation. PMID: 10835425 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 238: Front Biosci. 2000 Jun 1;5:D594-601. Cell cycle implications in the pathogenesis of rheumatoid arthritis. Michael VV, Alisa KE. Department of Medicine, Northwestern University, Chicago, IL 60611, USA. Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by hyperplasia of the synovial lining cells, angiogenesis, and infiltration of mononuclear cells resulting in pannus formation, cartilage erosion and ultimately joint destruction. Synovial tissue (ST) fibroblast hyperplasia is reminiscent of tumor-like proliferation and is a major cause of cartilage destruction in the RA joint. The RA joint is replete with cytokines and growth factors which exert a synergistic mitogenic effect on ST fibroblasts. As a result, RA ST fibroblasts exhibit elevated gene expression of proto- oncogenes, such as c-Myc, c-Ras, and c-Jun and apoptosis inhibitors such as Bcl-2. At the same time, RA ST fibroblasts contain mutations in tumor suppressor genes such as p53. The altered rates of proliferation and apoptosis of RA synovial cells result in the hyperplasia of synovial tissue and in concert with the chronic inflammatory environment ultimately lead to the destruction of the RA joint. Publication Types: Review Review, Tutorial PMID: 10833466 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 239: Biochem Biophys Res Commun. 2000 Jun 7;272(2):591-5. Studies on hepatic gene expression in different liver regenerative models. Nagy P, Bisgaard HC, Schnur J, Thorgeirsson SS. First Institute of Pathology and Experimental Cancer Research, Semmelweis University of Medicine, Budapest, Hungary. peter_nagy@nih.gov We have investigated the expression of several growth-related genes in the liver after partial hepatectomy in three experimental models: normal, Dexamethasone-pretreated, and hypophysectomized rats. Dexamethasone and hypophysectomy resulted in a delay in the peak of cell replication in 6 and 18 h, respectively, when compared to the normal animals. TGFalpha mRNA expression was shifted together with the DNA synthesis, but the expression of c-myc, c-fos, c-jun, HGF, TGFbeta1, IL1beta did not delay. This result suggests that liver-derived TGFalpha but not the other factors are important in the timing of the proliferative response after partial hepatectomy. Copyright 2000 Academic Press. PMID: 10833457 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 240: Cell Mol Life Sci. 2000 Mar;57(3):421-8. Calcium and disease: molecular determinants of calcium crystal deposition diseases. Misra RP. Department of Biochemistry, Medical College of Wisconsin, Milwaukee 53226, USA. rmisra@mcw.edu Deposition of basic calcium phosphate (hydroxyapatite, octacalcium phosphate and tricalcium phosphate) (BCP) and crystalline calcium pyrophosphate dihydrate (CPPD) is associated with a variety of aging-related pathologies, including osteoarthritis, cartilage degeneration and pseudogout. These diseases of calcium deposition serve as some of the best-studied examples of how calcium-regulated changes in gene expression can directly lead to pathogenic consequences. Tissue damage can result when crystals stimulate cells to release matrix-degrading molecules or secrete cytokines that stimulate the release of matrix-degrading molecules. Exposure of cultured cells to crystals induces expression of cellular proto-oncogenes such as c-fos, c-myc and c-jun, by a calcium-dependent mechanism, and this response can be blocked by a potential therapeutic compound, phosphocitrate. Activation of the c-fos and c-jun genes is directly involved in expression of metalloproteinases such as collagenase and stromelysin, suggesting that crystal-mediated activation of these genes is directly involved in pathogenesis. In this review recent advances in the molecular mechanisms responsible for crystal-mediated cell activation are discussed. Publication Types: Review Review, Tutorial PMID: 10823243 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 241: Growth Factors. 2000;17(4):249-68. Gene response of human skin fibroblasts to urokinase- and tissue-type plasminogen activators. Copeta A, Tavian D, Marchina E, De Petro G, Barlati S. Department of Biomedical Sciences and Biotechnologies, University of Brescia, Italy. In a previous work we have reported evidences on the mitogenic activity of urokinase-type and tissue-type plasminogen activator (u-PA, t-PA) on serum-deprived human dermal fibroblasts. In this work we have studied the transcription-dependent changes of some cell-cycle related genes associated with the biological activity of PAs, as well as the possible involvement of protein tyr kinases (PTK) and/or protein kinase C (PKC) in the mitogenic signal transduction. The data obtained demonstrate that the growth factor activity of PAs is associated with: - a rapid transient activation of early response genes, c-fos, c-jun and c-myc; - the subsequent coordinated down-regulation of p53 and p21CIP1; - the constant expression of the MEK1 mRNA in every phase of the cell cycle. Quiescent (G0) cells did not express c-fos, c-jun, c-myc and cyclin A, but upon stimulation with mitogens (fetal calf serum (FCS), u-PA, t-PA) the cyclin A mRNA expression was observed in concomitance with the activation of DNA synthesis. Therefore u-PA, t-PA and FCS similarly modulate the expression of c-fos, c-jun, c-myc, p53, p21CIP1 and cyclin A with only slight differences likely related to the time required for activation of DNA synthesis. The PAs mitogenic stimulation of serum-starved cells was associated with the internalization of their molecules, as revealed by immunostaining. The biological activity of u-PA, t-PA, as well as that of limiting concentration of FCS (1%), was mediated by PTK and PKC. Conversely, PTK, but not PKC, was involved in the activation of the proliferative response of basic fibroblast growth factor in the same experimental conditions. In conclusion, u-PA and t-PA can utilize two different pathways, one depending on PTK and the other on PKC in a way similar to the mitogenic activity induced by low concentration of FCS (1%). PMID: 10801075 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 242: Toxicol Appl Pharmacol. 2000 May 1;164(3):291-300. Cadmium induces c-myc, p53, and c-jun expression in normal human prostate epithelial cells as a prelude to apoptosis. Achanzar WE, Achanzar KB, Lewis JG, Webber MM, Waalkes MP. Inorganic Carcinogenesis Section, National Cancer Institute at National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, 27709, USA. Cadmium is a suspected human prostatic carcinogen shown to induce prostatic tumors and proliferative lesions in rats. The carcinogenic mechanism of cadmium is unknown, but its poor mutagenicity points toward an epigenetic mechanism. Here we studied the effect of cadmium on genes involved in growth regulation of prostate epithelial cell using the human prostate epithelial cell line RWPE-1, which is immortalized but not transformed and is androgen-responsive. Treatment with 10 microM cadmium resulted in transient increases in c-myc and p53 mRNA levels that peaked at 2-fold and 1.4-fold, respectively, compared to control after 2 h. In contrast, c-jun mRNA levels were increased >3-fold after 2, 4, and 6 h and 20-fold after 24 h. DNA synthesis decreased after 24 h of cadmium exposure. Further study revealed a significant increase in apoptosis after 48 h of cadmium exposure. However, approximately 35% of the cells were still viable and appeared normal, indicating this subpopulation was more resistant to cadmium. Furthermore, these resistant cells had 2.5-fold more metallothionein than untreated control cells. This suggests that cadmium could act to select for apoptotic-defective cells in vivo, thereby increasing the likelihood of tumor formation. This work represents the first description of cadmium affecting oncogene expression in a human cell model of a potential in vivo target site of cadmium carcinogenesis. Copyright 2000 Academic Press. PMID: 10799339 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 243: Toxicol In Vitro. 2000 Apr;14(2):159-67. Responses of the rabbit tracheal epithelium in vitro to H(2)O(2)-induced oxidative stress. Baeza-Squiban A, Delcher L, Kukreti R, Joly AC, Guennou C, Houcine O, Marano F. Laboratoire de Cytophysiologie et Toxicologie cellulaire, Universite Paris 7, Denis Diderot, tour 53-54, 3e etage, case 7073, 2 place jussieu, 75 251, Paris, France. baeza@paris7.jussieu.fr A model of rabbit tracheal epithelial (RTE) cells in primary culture was used to characterize specific and repair responses of airway epithelial cells to oxidative stress. Two well-known reactive oxygen species (ROS) generating systems were used: H(2)O(2) alone or in combination with Fe(2+) to produce the hydroxyl radical. RTE cells exhibited lipid peroxidation when exposed to H(2)O(2) + Fe(2+). Moreover, catalase (CAT) activity decreased after a 1-hour treatment in 3-day-old cultures but increased in 7-day-old cultures which have higher antioxidant enzyme activities. Superoxide dismutase (SOD) activity was never affected. In addition, RTE cells displayed a repair response leading to squamous metaplasia. H(2)O(2) + Fe(2+) treatment resulted in a time-dependent increase in the steady-state level of c-myc mRNA while c-jun and c-fos were not activated. Moreover, a chronic exposure induced the expression of the squamous phenotype characterized by the expression of the cytokeratin 13 confirmed both at the message and protein levels. RTE cells in primary culture react early to H(2)O(2) + Fe(2+) exposure by an increase in c-myc expression and by modifications in CAT activity. Further, a lipid peroxidation occurs and the tracheal epithelium evolves to squamous metaplasia. PMID: 10793294 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 244: Radiat Res. 2000 May;153(5 Pt 2):658-62. Exposure to low-frequency electromagnetic fields does not alter HSP70 expression or HSF-HSE binding in HL60 cells. Morehouse CA, Owen RD. FDA Center for Devices and Radiological Health, Rockville, Maryland 20850, USA. Environmental exposure to extremely low-frequency electromagnetic fields (ELF EMFs) has been identified as a possible contributor to increased cancer incidence and other diseases. In vitro studies designed to probe for biological mechanisms that might explain such relationships have included several studies of gene expression. While gene expression studies have focused on MYC, effects of ELF EMFs on the expression of beta-actin, histone H2B, beta-tubulin, SRC, FOS and JUN have also been reported. In addition, some investigators have reported both an induction of HSP70 expression and an increase in HSF-HSE binding in HL60 (human promyelocytic leukemia) cells after exposure to a 60 Hz magnetic field. In this study, HL60 cells were exposed to a weak 60 Hz magnetic field (6.3 or 8.0 microT) or to a positive control heat shock (42 or 44 degrees C). While heat shock led to reproducible induction of HSP70 expression and HSF-HSE binding, no significant effect of exposure to ELF EMFs on either of these end points was observed. PMID: 10790289 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 245: Biol Chem. 2000 Mar;381(3):193-8. p38/SAPK2-dependent gene expression in Jurkat T cells. Rolli-Derkinderen M, Gaestel M. Innovationskolleg Zellspezialisierung der Martin-Luther-Universitat Halle/Wittenberg, Halle, Germany. The stress-activated protein kinase p38/SAPK2 is known to regulate the activity of transcription factors and to control expression of several genes at the transcriptional or post-transcriptional level. In order to identify genes whose expression is under the control of p38/SAPK2 activity, we have compared the mRNA levels of a pattern of 588 genes between human Jurkat T cells with anisomycin-activated p38/SAPK2 and cells in which p38/SAPK2 was inhibited by the compound SB203580. Genes strongly expressed at the transcript level as a result of p38/SAPK2 activation are the transcription factors c-jun, fos-related antigen 1 (fra-1), the growth-arrest and DNA-damage gene gadd153 and early-growth-related gene 1 (egr-1) as well as the c-srk kinase csk and the nucleotide exchange factor ras-GRF. mRNAs significantly down-regulated include the insulin receptor IR, the adapter grb2, the transcription factor c-myc and the defender against apoptotic death, dad-1. For six selected genes, p38/SAPK2-regulated expression was confirmed and further analysed by Northern blot experiments, demonstrating a complex regulation of these genes under stress conditions. PMID: 10782990 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 246: Mol Cell Biol. 2000 May;20(10):3616-25. UV-Induced stabilization of c-fos and other short-lived mRNAs. Blattner C, Kannouche P, Litfin M, Bender K, Rahmsdorf HJ, Angulo JF, Herrlich P. Forschungszentrum Karlsruhe, Institute of Toxicology and Genetics, 76021 Karlsruhe, Germany. Irradiation of cells with short-wavelength ultraviolet light (UVC) changes the program of gene expression, in part within less than 15 min. As one of the immediate-early genes in response to UV, expression of the oncogene c-fos is upregulated. This immediate induction is regulated at the transcriptional level and is transient in character, due to the autocatalyzed shutoff of transcription and the rapid turnover of c-fos mRNA. In an experiment analyzing the kinetics of c-fos mRNA expression in murine fibroblasts irradiated with UVC, we found that, in addition to the initial transient induction, c-fos mRNA accumulated in a second wave starting at 4 to 5 h after irradiation, reaching a maximum at 8 h, and persisting for several more hours. It was accompanied by an increase in Fos protein synthesis. The second peak of c-fos RNA was caused by an UV dose-dependent increase in mRNA half-life from about 10 to 60 min. With similar kinetics, the mRNAs of other UV target genes (i.e., the Kin17 gene, c-jun, IkappaB, and c-myc) were stabilized (e.g., Kin17 RNA from 80 min to more than 8 h). The delayed response was not due to autocrine cytokine secretion with subsequent autostimulation of the secreting cells or to UV-induced growth factor receptor activation. Cells unable to repair UVC-induced DNA damage responded to lower doses of UVC with an even greater accumulation of c-fos and Kin17 mRNAs than repair-proficient wild-type cells, suggesting that a process in which a repair protein is involved regulates mRNA stability. Although resembling the induction of p53, a DNA damage-dependent increase in p53 was not a necessary intermediate in the stabilization reaction, since cells derived from p53 knockout mice showed the same pattern of c-fos and Kin17 mRNA accumulation as wild-type cells. The data indicate that the signal flow induced by UV radiation addresses not only protein stability (p53) and transcription but also RNA stability, a hitherto-unrecognized level of UV-induced regulation. PMID: 10779351 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 247: Mol Cell Biol. 2000 May;20(10):3396-406. Slap negatively regulates Src mitogenic function but does not revert Src-induced cell morphology changes. Manes G, Bello P, Roche S. Centre de Recherche de Biochimie Macromoleculaire, Centre National de la Recherche Scientifique UPR-1086, 34293 Montpellier, France. Src-like adapter protein (Slap) is a recently identified protein that negatively regulates mitogenesis in murine fibroblasts (S. Roche, G. Alonso, A. Kazlausakas, V. M. Dixit, S. A. Courtneidge, and A. Pandey, Curr. Biol. 8:975-978, 1998) and comprises an SH3 and SH2 domain with striking identity to the corresponding Src domains. In light of this, we sought to investigate whether Slap could be an antagonist of all Src functions. Like Src, Slap was found to be myristylated in vivo and largely colocalized with Src when coexpressed in Cos7 cells. Microinjection of a Slap-expressing construct into quiescent NIH 3T3 cells inhibited platelet-derived growth factor (PDGF)-induced DNA synthesis, and the inhibition was rescued by the transcription factor c-Myc but not by c-Jun/c-Fos expression. Fyn (or Src) overexpression overrides the G(1)/S block induced by both SrcK- and a Slap mutant with a deletion of its C terminus (SlapDeltaC), but not the block induced by Slap or SlapDeltaSH3, implying that the C terminus is a noncompetitive inhibitor of Src mitogenic function. Furthermore, a chimeric adapter comprising SrcDeltaK fused to the Slap C terminus (Src/SlapC) also inhibited Src function during the PDGF response in a noncompetitive manner, as Src coexpression could not rescue PDGF signaling. Slap, however, did not reverse deregulated Src-induced cell transformation, as it was unable to inhibit depolymerization of actin stress fibers while still being able to inhibit SrcY527F-induced DNA synthesis. This was attributed to a distinct Slap SH3 binding specificity, since the chimeric Slap/SrcSH3 molecule, in which the Slap SH3 was replaced by the Src SH3 sequence, substantially restored stress fiber formation. Indeed, three amino acids important for ligand binding in Src SH3 were replaced in the Slap SH3 sequence; Slap SH3 did not bind to the Src SH3 partners p85alpha, Shc, and Sam68 in vitro, and the chimeric tyrosine kinase Slap/SrcK, composed of SlapDeltaC fused to the SH2 linker kinase sequence of Src, was not regulated in vivo. Furthermore, the Src SH3 domain is required for signaling during mitogenesis and since Slap/SrcK behaved as a dominant negative in the PDGF mitogenic response when microinjected into quiescent fibroblasts. We conclude that Slap is a negative regulator of Src during mitogenesis involving both the SH2 and the C terminus domains in a noncompetitive manner, but it does not regulate all Src function due to specific SH3 binding substrates. PMID: 10779329 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 248: Injury. 2000 Jun;31(5):363-6. Proto-oncogene expression during fracture repair. Oni OO. The Glenfield Hospital, Groby Road, Leicester, UK. The possibility that proto-oncogenes play a role in fracture repair has been investigated in this study. Closed fractures were created in the tibiae of NZW rabbits and treated with a plaster cast. At intervals of 1 and 2 weeks, sections of the fracture callus were obtained and stained using an immunocytochemical technique which localised c-myc and c-jun. Osteogenic cells of the periosteum, endosteum, Haversian canal and new bone trabeculae expressed proto-oncogenes while hypertrophic chondrocytes were generally immunoreactive negative. The results suggest that proto-oncogene expression may be involved in post-fracture osteogenesis but not in the chondrogenic processes which precede endochondral bone formation. This could be used as a tool for diagnosing slow healing in the early stages of fracture repair. PMID: 10775693 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 249: Toxicol Sci. 2000 Apr;54(2):399-407. Effect of trichloroethylene and its metabolites, dichloroacetic acid and trichloroacetic acid, on the methylation and expression of c-Jun and c-Myc protooncogenes in mouse liver: prevention by methionine. Tao L, Yang S, Xie M, Kramer PM, Pereira MA. Department of Pathology, Medical College of Ohio, Toledo 43614-5806, USA. ltao@mco.edu Trichloroethylene (TCE), dichloroacetic acid (DCA), and trichloroacetic acid (TCA) are environmental contaminants that are carcinogenic in mouse liver. 5-Methylcytosine (5-MeC) in DNA is a mechanism that controls the transcription of mRNA, including the protooncogenes, c-jun and c-myc. We have previously reported that TCE decreased methylation of the c-jun and c-myc genes and increased the level of their mRNAs. Decreased methylation of the protooncogenes could be a result of a deficiency in S-adenosylmethionine (SAM), so that methionine, by increasing the level of SAM, would prevent hypomethylation of the genes. For 5 days, female B6C3F1 mice were administered, daily by oral gavage, either 1000 mg/kg body weight of TCE or 500 mg/kg DCA or TCA. At 30 min after each dose of carcinogen, the mice received, by ip injection, 0-, 30-, 100-, 300-, or 450-mg/kg methionine. Mice were euthanized at 100 min after the last dose of DCA, TCA, or TCE. Decreased methylation in the promoter regions of the c-jun and c-myc genes and increased levels of their mRNA and proteins were found in livers of mice exposed to TCE, DCA, and TCA. Methionine prevented both the decreased methylation and the increased levels of the mRNA and proteins of the two pro-tooncogenes. The prevention by methionine of DCA- TCA-, and TCE-induced DNA hypomethylation supports the hypothesis that these carcinogens act by depleting the availability of SAM. Hence, methionine would prevent DNA hypomethylation by maintaining the level of SAM. Furthermore, the results suggest that the dose of DCA, TCA, or TCE must be sufficient to decrease the level of SAM in order for these carcinogens to be active. PMID: 10774822 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 250: Int Rev Cytol. 2000;197:137-202. Protooncogenes as mediators of apoptosis. Teng CS. Department of Anatomy, Physiological Sciences, and Radiology, North Carolina State University, Raleigh 27606, USA. Apoptosis has been well established as a vital biological phenomenon that is important in the maintenance of cellular homeostasis. Three major protooncogene families and their encoded proteins function as mediators of apoptosis in various cell types and are the subject of this chapter. Protooncogenic proteins such as c-Myc/Max, c-Fos/c-Jun, and Bcl-2/Bax utilize a synergetic effect to enhance their roles in the pro- or antiapoptotic action. These family members activate and repress the expression of their target genes, control cell cycle progression, and execute programmed cell death. Repression or overproduction of these protooncogenic proteins induces apoptosis, which may vary as a result of either cell type specificity or the nature of the apoptotic stimuli. The proapoptotic and antiapoptotic proteins exert their effects in the membrane of cellular organelles. Here they generate cell-type-specific signals that activate the caspase family of proteases and their regulators for the execution of apoptosis. Publication Types: Review PMID: 10761117 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 251: Kidney Int. 2000 Apr;57(4):1277-82. Pleiotropic action of aldosterone in epithelia mediated by transcription and post-transcription mechanisms. Verrey F, Pearce D, Pfeiffer R, Spindler B, Mastroberardino L, Summa V, Zecevic M. Institute of Physiology, University of Zurich, Switzerland. verrey@physiol.unizh.ch The aldosterone-induced increase in sodium reabsorption across tight epithelia can be divided schematically into two functional phases: an early regulatory phase starting after a lag period of 20 to 60 minutes, during which the pre-existing transport machinery is activated, and a late phase (>2.5 h), which can be viewed as an anabolic action leading to a further amplification/differentiation of the Na+ transport machinery. At the transcriptional level, both early and late responses are initiated during the lag period, but the functional impact of newly synthesized regulatory proteins is faster than that of the structural ones. K-Ras2 and SGK were identified as the first early aldosterone-induced regulatory proteins in A6 epithelia. Their mRNAs also were shown to be regulated in vivo by aldosterone, and their expression (constitutively active K-Ras2 and wild-type SGK) was shown to increase the function of ENaC coexpressed in Xenopus oocytes. Recently, aldosterone was also shown to act on transcription factors in A6 epithelia: It down-regulates the mRNAs of the proliferation-promoting c-Myc, c-Jun, and c-Fos by a post-transcriptional mechanism, whereas it up-regulates that of Fra-2 (c-Fos antagonist) at the transcriptional level. Together, these new data illustrate the complexity of the regulatory network controlled by aldosterone and support the view that its early action is mediated by the induction of key regulatory proteins such as K-Ras2 and SGK. These early induced proteins are sites of convergence for different regulatory inputs, and thus, their aldosterone-regulated expression level tunes the impact of other regulatory cascades on sodium transport. This suggests mechanisms for the escape from aldosterone action. Publication Types: Review Review, Tutorial PMID: 10760054 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 252: Crit Rev Biochem Mol Biol. 2000;35(1):1-33. Ku autoantigen: a multifunctional DNA-binding protein. Tuteja R, Tuteja N. International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi. renu@icgeb.res.in Ku is a heterodimeric protein composed of approximately 70- and approximately 80-kDa subunits (Ku70 and Ku80) originally identified as an autoantigen recognized by the sera of patients with autoimmune diseases. Ku has high binding affinity for DNA ends and that is why originally it was known as a DNA end binding protein, but now it is known to also bind the DNA structure at nicks, gaps, hairpins, as well as the ends of telomeres. It has been reported also to bind with sequence specificity to DNA and with weak affinity to RNA. Ku is an abundant nuclear protein and is present in vertebrates, insects, yeast, and worms. Ku contains ssDNA-dependent ATPase and ATP-dependent DNA helicase activities. It is the regulatory subunit of the DNA-dependent protein kinase that phosphorylates many proteins, including SV-40 large T antigen, p53, RNA-polymerase II, RP-A, topoisomerases, hsp90, and many transcription factors such as c-Jun, c-Fos, oct-1, sp-1, c-Myc, TFIID, and many more. It seems to be a multifunctional protein that has been implicated to be involved directly or indirectly in many important cellular metabolic processes such as DNA double-strand break repair, V(D)J recombination of immunoglobulins and T-cell receptor genes, immunoglobulin isotype switching, DNA replication, transcription regulation, regulation of heat shock-induced responses, regulation of the precise structure of telomeric termini, and it also plays a novel role in G2 and M phases of the cell cycle. The mechanism underlying the regulation of all the diverse functions of Ku is still obscure. Publication Types: Review PMID: 10755664 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 253: Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2000 Apr;17(2):87-90. [IL-4 gene transfer induces the differentiation of cells and inhibits the activity of telomerase in hepatoblastoma cells] [Article in Chinese] Fu J, Li C, Yang X, Li X. Children's Hospital of Shenzhen, Guangdong, 518026 P.R. China. OBJECTIVE: To investigate the effects of human interleukin-4(hIL-4) gene transfer on the differentiation and the activities of telomerase in hepatoblastoma cells. METHODS: Retroviral vector (PL-IL-4-SN) was employed to introduce hIL-4 gene into human hepatoblastoma cell line(Hep G2)cells. Trypan blue and Wright's stain, radioimmunoassay, in situ hybridization, flow cytometry, PCR-ELISA were used to determine the change in morphology and cell cycle, the expression of proto-oncogenes, the synthesis of AFP and the activities of telomerase in IL-4 gene transferred tumor cells. RESULTS: The shape of the hepatoblastoma cells tended to become relatively mature; the growth of the cells was significantly suppressed(P<0.05); the synthesis of AFP was reduced from 15.36+/-0. 67 ng x10(6) cells(-1); x24h(-1); to 3.26 +/- 1.43ng x10(6) cells(-1); x 24h(-1); the proliferation of the cells was significantly suppressed(P<0.05); the cell cycle was arrested at G(0)/ G(1) stage; the expression of c-fos, c-jun, c-myc and the activities of telomerase were remarkably decreased in IL-4 gene-modified Hep G2 cells. CONCLUSION: hIL-4 gene transfer could induce the differentiation of heptoblastoma cells and down-regulate the activity of tolemarase in Hep G2 cells. PMID: 10751527 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 254: Prog Cell Cycle Res. 2000;4:157-62. HTLV-I Tax and cell cycle progression. Neuveut C, Jeang KT. Laboratoire de Recombinaison et Expression Genetique, Institut Pasteur, Paris, France. Human T-cell leukemia virus type I (HTLV-I) is the etiological agent for adult T-cell leukemia (ATL) and various human myopathies/neuropathies. HTLV-I encodes a 40 kDa phosphoprotein, Tax, which has been implicated in cellular transformation. In similarity with several other oncoproteins such as Myc, Jun, and Fos, Tax is a transcriptional activator. How Tax mechanistically dysregulates the cell cycle remains unclear. Recent findings from us and others have shown that Tax targets key regulators of G1/S and M progression such as p16INK4a, cyclin D1, cyclin D3-cdk, and the mitotic spindle checkpoint apparatus. Thus, Tax influences the progression of cells in various phases of the cell cycle. In this regard, we will discuss three distinct mechanisms through which Tax affects cell-cycling: a) through direct association Tax can abrogate the inhibitory function of p16INK4a on the G1-cdks, b) Tax can also directly influence cyclin D-cdk activities by a protein-protein interaction, and c) Tax targets the HsMAD1 mitotic spindle-assembly checkpoint protein. Through these varied routes, the HTLV-I oncoprotein dysregulates cellular growth controls and engenders a proclivity of cells toward a loss of DNA-damage surveillance. Publication Types: Review Review, Tutorial PMID: 10740823 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 255: Life Sci. 2000 Feb 18;66(13):1261-70. Altered ischemic induction of immediate early gene and heat shock protein 70 mRNAs after preconditioning in rat hearts. Nitta-Komatsubara Y, Abe K, Aoki M, Isoyama S. Department of Cardiovascular Medicine, University of Tokyo Graduate School of Medicine, Japan. Immediate early genes and heat shock protein (HSP) 70s, which may play a role in adaptation and cellular protection, respectively, are induced by ischemia in hearts. We examined if the induction of immediate early gene (c-fos, c-myc, c-jun, and junB) and HSP70 mRNAs by ischemia is affected by ischemic preconditioning. Transient ischemia (5 or 10 minute) was applied to Wistar rat (n=75) hearts, by tightening a snare placed around left coronary arterial branches 7 days before applying ischemia. Rats without earlier ischemia (control group, C) and rats with 5-minute ischemia 12 or 24 hours earlier (EI12 or 24 group) were given 10-minute ischemia and sacrificed at 0, 0.5, 1, 2, and 4 hour. RNA was extracted from the ischemic region and Northern blot analysis was performed. The induction of c-fos and c-myc mRNAs was significantly increased in EI12 but not in EI24 compared with that in C. The induction of c-jun and junB mRNAs showed no change in both EI12 and EI24 compared with that in C. The induction of HSP72 and 73 mRNAs was decreased in EI12 and decreased further in EI24. Thus, ischemic preconditioning altered the induction of immediate early gene and HSP70 mRNAs by ischemia. The effect of preconditioning differed among genes studied and changed with time after preconditioning. Ischemic preconditioning alters protective and adaptive responses to ischemia at the gene level. PMID: 10737421 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 256: Food Chem Toxicol. 2000 Apr;38(4):339-49. Promotion of morphological transformation by Di-n-butyltin dichloride in C3H/10T1/2 cells: prediction by prior expression of tumour promoter-responsive genes. Parfett CL, Marquardt T, Pilon R. Mutagenesis Section, Environmental Health Directorate, Health Canada, Environmental Health Centre, Tunney's Pasture, Ottawa, Ontario, Canada. Craig_Parfett@hc-sc.gc.ca Previous studies in our laboratory have shown that chemical treatments may induce increases in proliferin gene family mRNA accumulation in cultured murine embryonic cells. Proliferin inductions are highly correlated with subsequent promotional outcomes during two-stage focus-formation assays in C3H/10T1/2 cell cultures. In work reported here, the strong affiliation between these two responses was further validated after treating cells with di-n-butyltin dichloride which is a polyvinyl chloride (PVC) plastic additive that often contaminates food and water. Increased proliferin expression and promotion of morphological transformation occurred at similar concentrations. Promotion of transformation was detected at di-n-butyltin dichloride concentrations of 80 nM (24 ng/ml) and above, if added to initiated cultures before confluent monolayers had formed. Proliferin induction and morphological transformation were both reduced in confluent cultures treated with di-n-butyltin dichloride, as compared to subconfluent cultures. Proliferin expression measured in near-confluent cultures was induced up to 10-fold during the 36-hr period following di-n-butyltin dichloride exposure and was accompanied by increased accumulation of transcripts from many genes regulated by oxidative stresses, growth-inducing agents, and/or other promoting agents (asbestos, superoxide radicals ). Di-n-butyltin dichloride-induced mRNA species included members of the fos and jun proto-oncogene families, c-myc, egr1, ribonucleotide reductase (R2 subunit), odc, macrophage chemotactic protein/je, hsp70, metallothionine IIA, c-sod and mn-sod. The observed patterns of RNA accumulation suggested that a small subset of mRNA species, including proliferin, exhibit regulatory behaviour as a response to dissimilar agents or conditions that promote focus-formation in C3H/10T1/2 cultures. Plausible predictions of promotional effects in two-stage morphological transformation assays can be made from gene-expression responses to test agents. PMID: 10722888 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 257: Thyroid. 2000 Feb;10(2):131-40. Erratum in: Thyroid 2001 Jun;11(6):following 609. Characterization of autonomous thyroid adenoma: metabolism, gene expression, and pathology. Deleu S, Allory Y, Radulescu A, Pirson I, Carrasco N, Corvilain B, Salmon I, Franc B, Dumont JE, Van Sande J, Maenhaut C. IRIBHN, Free University of Brussels, Brussels, Belgium. Fifty-one in vivo characterized autonomous single adenomas have been studied for functional parameters in vitro, for gene and protein expression and for pathology, and have been systematically compared to the corresponding extratumoral quiescent tissue. The adenomas were characterized by a high level of iodide trapping that corresponds to a high level of Na+ /iodide symporter gene expression, a high thyroperoxidase mRNA and protein content, and a low H2O2 generation. This explains the iodide metabolism characteristics demonstrated before, ie, the main cause of the "hot" character of the adenomas is their increased iodide transport. The adenomas spontaneously secreted higher amounts of thyroid hormone than the quiescent tissue and in agreement with previous in vivo data, this secretion could be further enhanced by thyrotropin (TSH). Inositol uptake was also increased but there was no spontaneous increase of the generation of inositol phosphates and this metabolism could be further activated by TSH. These positive responses to TSH are in agreement with the properties of TSH-stimulated thyroid cells in vitro and in vivo. They are compatible with the characteristics of mutated TSH receptors whose constitutive activation accounts for the majority of autonomous thyroid adenomas in Europe. The number of cycling cells, as evaluated by MIB-1 immunolabeling was low but increased in comparison with the corresponding quiescent tissue or normal tissue. The cycling cells are observed mainly at the periphery; there was very little apoptosis. Both findings account for the slow growth of these established adenomas. On the other hand, by thyroperoxidase immunohistochemistry, the whole lesion appeared hyperfunctional, which demonstrates a dissociation of mitogenic and functional stimulations. Thyroglobulin, TSH receptor, and E-cadherin mRNA accumulations were not modified in a consistent way, which confirms the near-constitutive expression of the corresponding genes in normal differentiated tissue. On the contrary, early immediate genes expressions (c-myc, NGF1B, egr 1, genes of the fos and jun families) were decreased. This may be explained by the proliferative heterogeneity of the lesion and the previously described short, biphasic expression of these genes when induced by mitogenic agents. All the characteristics of the autonomous adenomas can therefore be explained by the effect of the known activating mutations of genes coding for proteins of the TSH cyclic adenosine monophosphate (cAMP) cascade, all cells being functionally activated while only those at the periphery multiply. The reason of this heterogeneity is unknown. PMID: 10718549 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 258: Oncogene. 2000 Mar 2;19(10):1277-87. Differential roles of JNK and Smad2 signaling pathways in the inhibition of c-Myc-induced cell death by TGF-beta. Mazars A, Tournigand C, Mollat P, Prunier C, Ferrand N, Bourgeade MF, Gespach C, Atfi A. INSERM U482, Hopital Saint-Antoine, 184 Rue du Faubourg Saint-Antoine, 75571, Paris Cedex 12, France. The transforming growth factor beta (TGF-beta) plays an important role in constraining cellular proliferation, but it is also a potent inducer of programmed cell death or apoptosis. Here, we demonstrate that TGF-beta can have an opposite effect, acting as a survival factor to prevent c-Myc-induced cell death in Rat-1 fibroblasts. However, in marked contrast to TGF-beta, Smad2, which is a critical intracellular mediator of the TGF-beta signaling pathway, functions as an antagonist to induce increased cell death. The protective activity of TGF-beta was associated with the activation of c-Jun N-terminal Kinase (JNK) and was not linked to the ability of TGF-beta to promote cell cycle progression. Expression of dominant-interfering forms of various components of the JNK signaling pathway, including Rac1, Cdc42, mitogen-activated protein kinase kinase 4 (MKK4), and c-Jun, abolished TGF-beta-mediated cell survival. Furthermore, overexpression of the constitutively activated mutant RacL61F37A, which selectively stimulates JNK cascade but not G1 cell cycle progression or actin polymerization, was sufficient to prevent apoptosis induced by c-Myc. These findings describe a differential effect of two separated signaling pathways of TGF-beta and indicate for the first time that Smad2 can act as antagonist to suppress TGF-beta-dependent cell survival. Oncogene (2000) 19, 1277 - 1287. PMID: 10713669 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 259: Mol Cell Biol. 2000 Apr;20(7):2529-42. v-Jun overrides the mitogen dependence of S-phase entry by deregulating retinoblastoma protein phosphorylation and E2F-pocket protein interactions as a consequence of enhanced cyclin E-cdk2 catalytic activity. Clark W, Black EJ, MacLaren A, Kruse U, LaThangue N, Vogt PK, Gillespie DA. Beatson Institute for Cancer Research, Cancer Research Campaign Beatson Laboratories, Garscube Estate, Bearsden, Glasgow G61 1BD, Scotland, United Kingdom. v-Jun accelerates G(1) progression and shares the capacity of the Myc, E2F, and E1A oncoproteins to sustain S-phase entry in the absence of mitogens; however, how it does so is unknown. To gain insight into the mechanism, we investigated how v-Jun affects mitogen-dependent processes which control the G(1)/S transition. We show that v-Jun enables cells to express cyclin A and cyclin A-cdk2 kinase activity in the absence of growth factors and that deregulation of cdk2 is required for S-phase entry. Cyclin A expression is repressed in quiescent cells by E2F acting in conjunction with its pocket protein partners Rb, p107, and p130; however, v-Jun overrides this control, causing phosphorylated Rb and proliferation-specific E2F-p107 complexes to persist after mitogen withdrawal. Dephosphorylation of Rb and destruction of cyclin A nevertheless occur normally at mitosis, indicating that v-Jun enables cells to rephosphorylate Rb and reaccumulate cyclin A without exogenous mitogenic stimulation each time the mitotic "clock" is reset. D-cyclin-cdk activity is required for Rb phosphorylation in v-Jun-transformed cells, since ectopic expression of the cdk4- and cdk6-specific inhibitor p16(INK4A) inhibits both DNA synthesis and cell proliferation. Despite this, v-Jun does not stimulate D-cyclin-cdk activity but does induce a marked deregulation of cyclin E-cdk2. In particular, hormonal activation of a conditional v-Jun-estrogen receptor fusion protein in quiescent, growth factor-deprived cells stimulates cyclin E-cdk2 activity and triggers Rb phosphorylation and DNA synthesis. Thus, v-Jun overrides the mitogen dependence of S-phase entry by deregulating Rb phosphorylation, E2F-pocket protein interactions, and ultimately cyclin A-cdk2 activity. This is the first report, however, that cyclin E-cdk2, rather than D-cyclin-cdk, is likely to be the critical Rb kinase target of v-Jun. PMID: 10713176 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 260: Anticancer Res. 1999 Nov-Dec;19(6B):4969-76. Correlation of TGF beta 1 overexpression with down-regulation of proliferation-inducing molecules in HPV-11 transformed human tissue xenografts. Shier MK, Neely EB, Ward MG, Richards ME, Manders EC, Meyers C, Howett MK. Department of Microbiology and Immunology, Milton S. Hershey Medical Center, Pennsylvania State University College of Medicine 17033, USA. The epidemiologic association of human papillomavirus (HPV) infection with dysplasia and cervical cancer is well established. Transforming growth factor beta 1 (TGF beta 1) has regulatory effects on a broad spectrum of cell types and is a growth inhibitory protein for epithelial cells. To examine the phenotype of experimentally generated, HPV-11 transformed human tissues, we looked at expression of TGF beta 1 and a number of proliferation-enhancing molecules which are known to be regulated by TGF beta 1, including bcl-2, c-myc, c-Ha-ras, c-jun and NFkB. HPV-11 transformed xenografts showed up-regulation of TGF beta 1 expression and down-regulation of the expression levels of bcl-2, c-myc, c-Ha-ras, c-jun and NFkB. These results suggest that TGF beta 1 may exert antiproliferative effects on HPV-11 transformed papillomas by down-regulating different proliferation-enhancing molecules. PMID: 10697498 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 261: Biochem Biophys Res Commun. 2000 Feb 16;268(2):243-8. Regulation of beta-catenin signaling in the Wnt pathway. Kikuchi A. Department of Biochemistry, Hiroshima University School of Medicine, 1-2-3, Kasumi, Minami-ku, Hiroshima, 734-8551, Japan. akikuchi@mcai.med.hiroshimau.ac.jp beta-Catenin not only regulates cell to cell adhesion as a protein interacting with cadherin, but also functions as a component of the Wnt signaling pathway. The Wnt signaling pathway is conserved in various organisms from worms to mammals, and plays important roles in development, cellular proliferation, and differentiation. Wnt stabilizes cytoplasmic beta-catenin and then beta-catenin is translocated into the nucleus where it stimulates the expression of genes including c-myc, c-jun, fra-1, and cyclin D1. The amounts and functions of beta-catenin are regulated in both the cytoplasm and nucleus. Its molecular mechanisms are becoming increasingly well understood. Copyright 2000 Academic Press. Publication Types: Review Review, Tutorial PMID: 10679188 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 262: Am J Physiol Cell Physiol. 2000 Feb;278(2):C331-5. Inhibition of ornithine decarboxylase induces STAT3 tyrosine phosphorylation and DNA binding in IEC-6 cells. Pfeffer LM, Yang CH, Pfeffer SR, Murti A, McCormack SA, Johnson LR. Department of Pathology, University of Tennessee Health Science Center, Memphis, Tennessee 38163, USA. lpfeffer@utmem.edu Polyamines are required for the proliferation of the rat intestinal mucosal IEC-6 cell line. Ornithine decarboxylase (ODC) is the enzyme that catalyzes the first step in polyamine synthesis. ODC inhibition not only leads to polyamine depletion but also leads to inhibition of cell proliferation and regulates the expression of the immediate-early genes c-fos, c-myc, and c-jun. Members of the signal transducers and activators of transcription (STAT) transcription factor family bind to the sis-inducible element (SIE) present in the promoters to regulate the expression of a variety of important genes. In the present study, we tested the hypothesis that the STAT3 transcription factor, which is responsible for activation of the acute phase response genes, is activated after inhibition of ODC. We found that inhibition of ODC rapidly induces STAT3 activation as determined by STAT3 tyrosine phosphorylation, translocation of STAT3 from the cytoplasm into the nucleus, and the presence of STAT3 in SIE-dependent DNA-protein complexes. STAT3 activation upon inhibition of ODC was accompanied by the activation of a STAT3-dependent reporter construct. Moreover, prolonged polyamine depletion resulted in downregulation of cellular STAT3 levels. PMID: 10666028 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 263: Cytokine Growth Factor Rev. 1999 Sep-Dec;10(3-4):255-65. Modulation of Wnt signaling by Axin and Axil. Kikuchi A. Department of Biochemistry, Hiroshima University School of Medicine, Japan. akikuchi@mcai.med.hiroshima-u.ac.jp The Wnt signaling pathway is conserved in various species from worms to mammals, and plays important roles in development, cellular proliferation, and differentiation. The molecular mechanisms by which the Wnt signal regulates cellular functions are becoming increasingly well understood. Wnt stabilizes cytoplasmic beta-catenin, which stimulates the expression of genes including c-myc, c-jun, fra-1, and cyclin D1. Axin and its homolog Axil, newly recognized as components of the Wnt signaling pathway, negatively regulate this pathway. Other components of the Wnt signaling pathway, including Dvl, glycogen synthase kinase-3beta (GSK-3beta), beta-catenin, and adenomatous polyposis coli (APC), interact with Axin, and the phosphorylation and stability of beta-catenin are regulated in the Axin complex. Axil has similar functions to Axin. Thus, Axin and Axil act as scaffold proteins in the Wnt signaling pathway, thereby modulating the Wnt-dependent cellular functions. Publication Types: Review Review, Tutorial PMID: 10647780 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 264: Anticancer Res. 1999 Sep-Oct;19(5B):4221-7. Suppression of the TPA-induced expression of nuclear-protooncogenes in mouse epidermis by crocetin via antioxidant activity. Hsu JD, Chou FP, Lee MJ, Chiang HC, Lin YL, Shiow SJ, Wang CJ. Department of Pathology, Chung Shan Medical and Dental College Hospital, Taichung, Taiwan. Crocetin, a major component of the fruit of Gardenia jasminoides Ellis, was investigated for its antitumor promoting effect on 12-O-tetradecanoylphorbol-13-acetate-promoted mouse skin carcinogenesis. Topical application of 5 nmol TPA to CD-1 mice once daily for 5 days caused epidermal hyperplasia, and increases in the levels of c-Fos, c-Jun and c-Myc in the suprabasal layer of epidermis and the muscle layer of dermis. Immunocytolochemical examination showed that pretreatment of 1 mumol crocetin repressed the TPA-induced epidermal hyperplasia and the expressions of c-Jun, c-Fos and c-Myc to the extent of 47, 44 and 45% respectively. Crocetin of 3.0 mumol exhibited stronger inhibition on the induced hyperplasia and the oncoproteins levels (by 60, 53 and 55% respectively). Western blotting analysis confirmed this inhibitory effect of crocetin. Pretreatment of crocetin also repressed the TPA-induced H2O2 production and myeloperoxidase activity. These data indicate that crocetin suppresses the TPA-induced skin carcinogenesis maybe via its antioxidant property which, in turn, leads to a reduction in the TPA-induced expressions of c-Jun, c-Fos and c-Myc in mouse epidermis. PMID: 10628378 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 265: Cell Signal. 1999 Nov;11(11):777-88. Roles of Axin in the Wnt signalling pathway. Kikuchi A. Department of Biochemistry, Hiroshima University School of Medicine, Kasumi, Minami-ku, Japan. akikuchi@mcai.med.hiroshima-u.ac.jp The Wnt signalling pathway is conserved in various species from worms to mammals, and plays important roles in development, cellular proliferation, and differentiation. The molecular mechanisms by which the Wnt signal regulates cellular functions are becoming increasingly well understood. Wnt stabilizes cytoplasmic beta-catenin, which stimulates the expression of genes including c-myc, c-jun, fra-1, and cyclin D1. Axin, newly recognized as a component of the Wnt signalling pathway, negatively regulates this pathway. Other components of the Wnt signalling pathway, including Dvl, glycogen synthase kinase-3beta, beta-catenin, and adenomatous polyposis coli, interact with Axin, and the phosphorylation and stability of beta-catenin are regulated in the Axin complex. Thus, Axin acts as a scaffold protein in the Wnt signalling pathway, thereby regulating cellular functions. Publication Types: Review Review, Tutorial PMID: 10617280 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 266: Mol Cell Biol. 2000 Jan;20(2):684-96. Erratum in: Mol Cell Biol 2000 Apr;20(8):2949. Caspase 3 cleavage of the Ste20-related kinase SLK releases and activates an apoptosis-inducing kinase domain and an actin-disassembling region. Sabourin LA, Tamai K, Seale P, Wagner J, Rudnicki MA. Institute for Molecular Biology, McMaster University, Hamilton, Ontario, Canada. We have demonstrated that a novel Ste20-related kinase, designated SLK, mediates apoptosis and actin stress fiber dissolution through distinct domains generated by caspase 3 cleavage. Overexpression of SLK in C2C12 myoblasts stimulated the disassembly of actin stress fibers and focal adhesions and induced apoptosis, as determined by annexin V binding and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling analysis. SLK was cleaved by caspase 3 in vitro and in vivo during c-Myc-, tumor necrosis factor alpha, and UV-induced apoptosis. Furthermore, cleavage of SLK released two domains with distinct activities: an activated N-terminal kinase domain that promoted apoptosis and cytoskeletal rearrangements and a C-terminus domain that disassembled actin stress fibers. Moreover, our analysis has identified a novel conserved region (termed the AT1-46 homology domain) that efficiently promotes stress fiber disassembly. Finally, transient transfection of SLK also activated the c-Jun N-terminal kinase signaling pathway. Our results suggest that caspase-activated SLK represents a novel effector of cytoskeletal remodeling and apoptosis. PMID: 10611247 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 267: Oncogene. 1999 Dec 9;18(52):7566-75. Induction of apoptosis by SLK, a Ste20-related kinase. Sabourin LA, Rudnicki MA. MOBIX, Institute for Molecular Biology and Biotechnology, McMaster University, Hamilton, Ontario, Canada. We have cloned and characterized a novel murine Ste20-related kinase designated SLK. SLK displays high homology to the Ste20-related kinase LOK, and is more distantly related to MST1 and 2, both Ste20-like kinases. In addition, SLK displays high homology to microtubule and nuclear associated protein (M-NAP) and AT1-46, both of unknown function. SLK is ubiquitously expressed as multiple mRNAs in tissues and cell lines and is downregulated by mitogen depletion in differentiating myoblasts. Biochemical characterization showed that SLK overexpression activates c-Jun amino-terminal kinase 1 (JNK1). However, in vitro kinase assays indicated that SLK was not activated in response to various growth factors or stress-inducing agents. Immunofluorescence studies revealed that SLK colocalized to distinct cytosolic domains, preferentially at the periphery of the cells. In addition, prolonged overexpression of SLK in cultured fibroblasts resulted in apoptosis as demonstrated by annexin-V and TUNEL staining. Our results suggest that SLK belongs to a new family of protein kinases, mediating activation of the stress response pathway through a novel signaling cascade. PMID: 10602516 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 268: Oncogene. 1999 Dec 9;18(52):7552-8. Protein stabilization: a common consequence of mutations in independently derived v-Myc alleles. Gavine PR, Neil JC, Crouch DH. Biomedical Research Centre, University of Dundee, Ninewells Hospital and Medical School, Dundee DD1 9SY, UK. Myc is overexpressed in many cancers as a result of gene rearrangement or amplification, but coding sequence changes which cluster in the N-terminal transactivation domain also appear to play a role in tumour progression. The prototypic v-Myc gene of MC29 virus differs from avian c-Myc by a series of mutations, including a change at a regulatory phosphorylation site within the mutational hotspot (thr-61) which is known to potentiate transformation in vitro. We now show that the mutation at thr-61 stabilizes the v-Myc protein (turnover difference) and that this single mutation is both necessary and sufficient for the phenotype. A major involvement of the proteasome in Myc degradation was confirmed, but surprisingly, a dilysine motif adjacent to thr-61 proved not to be the ubiquitin target. Two other v-Myc genes which carry a mutation at thr-61 (avian MH2) or a large deletion encompassing this domain (feline T17) were found to be stabilized to a similar extent as MC29, showing that stabilization is a common feature of independently derived Myc oncogenes. These results suggest a common selective process in the genesis of these three viral oncoproteins and a mechanistic link with Jun, Fos and Myb oncoproteins which are also stabilized relative to their cellular counterparts. PMID: 10602514 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 269: Oncogene. 1999 Nov 25;18(50):7016-25. Induction of apoptosis in U937 human leukemia cells by suberoylanilide hydroxamic acid (SAHA) proceeds through pathways that are regulated by Bcl-2/Bcl-XL, c-Jun, and p21CIP1, but independent of p53. Vrana JA, Decker RH, Johnson CR, Wang Z, Jarvis WD, Richon VM, Ehinger M, Fisher PB, Grant S. Department of Medicine, Medical College of Virginia, Richmond 23298, USA. Determinants of differentiation and apoptosis in myelomonocytic leukemia cells (U937) exposed to the novel hybrid polar compound SAHA (suberoylanilide hydroxamic acid) have been examined. In contrast to hexamethylenbisacetamide (HMBA), SAHA-related maturation was limited and accompanied by marked cytoxicity. SAHA-mediated apoptosis occurred within the G0G1 and S phase populations, and was associated with decreased mitochondrial membrane potential, caspase-3 activation, PARP degradation, hypophosphorylation/cleavage of pRB, and down-regulation of c-Myc, c-Myb, and B-Myb. Enforced expression of Bcl-2 or Bcl-XL inhibited SAHA-induced apoptosis, but only modestly potentiated differentiation. While SAHA induced the cyclin-dependent kinase inhibitor p21CIP1, antisense ablation of this CDKI increased, rather than decreased, SAHA-related lethality. In contrast, conditional expression of wild-type p53 failed to modify SAHA actions, but markedly potentiated HMBA-induced apoptosis. Finally, SAHA modestly increased expression/activation of the stress-activated protein kinase (SAPK/JNK); moreover, SAHA-related lethality was partially attenuated by a dominant-negative c-Jun mutant protein (TAM67). SAHA did not stimulate mitogen-activated protein kinase (MAPK), nor was lethality diminished by the specific MEK/MAPK inhibitor PD98059. These findings indicate that SAHA potently induces apoptosis in human leukemia cells via a pathway that is p53-independent but at least partially regulated by Bcl-2/Bcl-XL, p21CIP1, and the c-Jun/AP-1 signaling cascade. PMID: 10597302 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 270: J Comp Physiol [B]. 1999 Oct;169(7):521-7. Discordant responses of mitogen-activated protein kinases to anoxia and freezing exposures in hatchling turtles. Greenway SC, Storey KB. Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario, Canada. The role of two vertebrate mitogen-activated protein kinases (MAPKs) in mediating responses to in vivo anoxia or freezing exposures was examined in four organs (liver, heart, kidney and brain) of hatchling red-eared turtles, Trachemys scripta elegans, which are naturally tolerant of these stresses. The extracellular signal-regulated kinases were not stress-activated except in brain of frozen turtles. The c-Jun NH2-terminal kinases (JNKs) were transiently activated by anoxia exposure in all four organs (after 1 h in brain or 5 h in other organs) but activity was suppressed during freezing except in brain which showed a transient activation of JNK after 1 h. Changes in the concentrations of the transcription factors, c-Fos and c-Myc, were also stress- and organ-specific. The patterns of MAPK activation in a stress-tolerant animal suggest the relative importance of these kinase pathways in cellular adaptation to oxygen deprivation or freezing and identify novel natural activators of MAPKs in vivo. The specificity of the signaling pathways is also emphasized here as the general whole-body stresses, anoxia and freezing, activated individual MAPKs in a tissue-, time-, and stress-dependent manner. PMID: 10595322 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 271: FASEB J. 1999 Dec;13(15):2091-104. Studies of the molecular mechanisms in the regulation of telomerase activity. Liu JP. Molecular Signaling Laboratory, Baker Medical Research Institute, Prahran, Victoria, Australia. jun-ping.liu@baker.edu.au Telomerase, a specialized RNA-directed DNA polymerase that extends telomeres of eukaryotic chromosomes, is repressed in normal human somatic cells but is activated during development and upon neoplasia. Whereas activation is involved in immortalization of neoplastic cells, repression of telomerase permits consecutive shortening of telomeres in a chromosome replication-dependent fashion. This cell cycle-dependent, unidirectional catabolism of telomeres constitutes a mechanism for cells to record the extent of DNA loss and cell division number; when telomeres become critically short, the cells terminate chromosome replication and enter cellular senescence. Although neither the telomere signaling mechanisms nor the mechanisms whereby telomerase is repressed in normal cells and activated in neoplastic cells have been established, inhibition of telomerase has been shown to compromise the growth of cancer cells in culture; conversely, forced expression of the enzyme in senescent human cells extends their life span to one typical of young cells. Thus, to switch telomerase on and off has potentially important implications in anti-aging and anti-cancer therapy. There is abundant evidence that the regulation of telomerase is multifactorial in mammalian cells, involving telomerase gene expression, post-translational protein-protein interactions, and protein phosphorylation. Several proto-oncogenes and tumor suppressor genes have been implicated in the regulation of telomerase activity, both directly and indirectly; these include c-Myc, Bcl-2, p21(WAF1), Rb, p53, PKC, Akt/PKB, and protein phosphatase 2A. These findings are evidence for the complexity of telomerase control mechanisms and constitute a point of departure for piecing together an integrated picture of telomerase structure, function, and regulation in aging and tumor development-Liu, J.-P. Studies of the molecular mechanisms in the regulation of telomerase activity. Publication Types: Review PMID: 10593857 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 272: Calcif Tissue Int. 1999 Nov;65(5):369-73. Immunohistochemical localization of selected early response genes expressed in trabecular bone of young rats given hPTH 1-34. Liang JD, Hock JM, Sandusky GE, Santerre RF, Onyia JE. Skeletal Disease Research Group, 0403 Endocrine Division, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285, USA. Intermittent administration of parathyroid hormone (PTH) increases trabecular bone mass in vivo by stimulating bone formation. To further characterize the cellular and molecular mediators of the anabolic response to PTH, we examined the effect of intermittent synthetic hPTH 1-34 on the expression and localization of selected early response genes, c-fos, c-jun, c-myc, and IL-6 protein, in bone tissue by immunohistochemistry. Young male Sprague-Dawley rats, 70-100 g, were injected s.c. with 8 microg/100 g PTH or vehicle control, once daily for 5 days. Femurs were harvested 1 and 24 hours after the fifth injection, then fixed, decalcified, processed for wax embedding, and sections were immunostained. Early response genes, c-fos, c-jun and IL-6, were strongly expressed in osteoblasts, osteocytes, and megakaryocytes in bones 1 hour after PTH, when compared with vehicle-treated controls or sections from rats, 24 hours after PTH injection. Osteoblasts, osteocytes, and megakaryocytes were also positive for c-myc but the differences in stain intensity between control and treated groups were marginal. Also, scattered islands of hematopoietic cells in the marrow stained intensely for IL-6 by 1 hour after PTH, but the stain intensity decreased to control level 24 hours after the last PTH injection. Scattered islands of hematopoietic cells in the bone marrow stained more strongly for c-fos than either c-jun or c-myc, but neither localization nor stain intensity were regulated by PTH at the time points examined. We conclude that during the immediate early phase of the anabolic response, PTH regulates c-fos, c-jun, and IL-6 expression in osteoblasts, osteocytes, megakaryocytes, and selected bone marrow hematopoietic cells in bone. PMID: 10541762 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 273: Endocrinology. 1999 Dec;140(12):5626-34. Epidermal growth factor and insulin-induced deoxyribonucleic acid synthesis in primary rat hepatocytes is phosphatidylinositol 3-kinase dependent and dissociated from protooncogene induction. Band CJ, Mounier C, Posner BI. Department of Medicine, McGill University, Montreal, Quebec, Canada. The mitogenic response to insulin and epidermal growth factor (EGF) was studied in subconfluent and confluent cultures of primary rat hepatocytes. In subconfluent cultures, wortmannin, LY294002, and rapamycin reversed insulin- and EGF-induced [3H]thymidine incorporation into DNA. The mitogen-activated protein kinase (MAPK) kinase 1 (MEK1) inhibitor PD98059 was without significant effect on either insulin- or EGF-induced [3H]thymidine incorporation. Insulin treatment did not alter levels of messenger RNAs (mRNAs) for c-fos, c-jun, and c-myc. EGF induced an increase in c-myc, but not c-fos or c-jun, mRNA levels in subconfluent hepatocyte cultures. This increase in c-myc mRNA was abolished by PD98059. In confluent cells that could not be induced to synthesize DNA, EGF treatment also promoted an increase in c-myc mRNA to levels seen in subconfluent cultures. This increase was also abrogated by PD98059. These data indicate that in primary rat hepatocyte cultures, 1) the phosphoinositol 3-kinase pathway, perhaps through p70s6k activation, regulates DNA synthesis in response to insulin and EGF; 2) the MAPKpathway is not involved in insulin- and EGF-induced DNA synthesis; and 3) p44/42 MAPKs are involved the induction of c-myc mRNA levels, although this induction is not required for DNA synthesis. These studies define two distinct signal transduction pathways that independently mediate growth-related responses in a physiologically relevant, normal cell system. PMID: 10579326 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 274: J Korean Med Sci. 1999 Oct;14(5):487-96. Angiotensin II stimulates proliferation of adventitial fibroblasts cultured from rat aortic explants. Kim DK, Huh JE, Lee SH, Hong KP, Park JE, Seo JD, Lee WR. Department of Medicine, Sungkyunkwan University School of Medicine, Samsung Cardiac and Vascular Center, Seoul, Korea. dkkim@smc.samsung.co.kr It has been proposed that the local renin-angiotensin system is activated in the adventitia after vascular injury. However, the physiological role of Angiotensin II (Ang II) in the adventitia has not been studied at a cellular level. This study was designed to assess the role of Ang II in the growth response of cultured adventitial fibroblasts (AFs). Adventitial explants of the rat thoracic aorta showed outgrowth of AFs within 5-7 days. Ang II caused hyperplastic response of AF cultures. The Ang II-induced mitogenic response of AFs was mediated primarily by the AT1 receptor. Ang II caused a rapid induction of immediate early genes (c-fos, c-myc and jun B). Induction of c-fos expression was fully blocked by an AT1 receptor antagonist but not by an AT2 receptor antagonist. Epidermal growth factor (EGF), platelet-derived growth factor-BB (PDGF-BB) and basic fibroblast growth factor (bFGF) induced DNA synthesis in AFs. Co-stimulation of AFs with the growth factors and Ang II potentiated the incorporation of 3H-thymidine into DNA. Results from this study indicate that Ang II causes mitogenesis of AFs via AT1 receptor stimulation and potentiates the responses to other mitogens. These data suggest that the Ang II may play an important role in regulating AF function during vascular remodeling following arterial injury. PMID: 10576143 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 275: J Biol Chem. 1999 Nov 12;274(46):32580-7. Regulation of c-Myc through phosphorylation at Ser-62 and Ser-71 by c-Jun N-terminal kinase. Noguchi K, Kitanaka C, Yamana H, Kokubu A, Mochizuki T, Kuchino Y. Biophysics Division, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. The expression of c-myc promotes cell proliferation and also sensitizes cells to various extracellular apoptotic stimuli. However, signal pathways regulating the function of Myc proteins during apoptosis are unknown. c-Jun N-terminal kinase (JNK) is activated by various apoptotic stimuli, but neither the target molecule(s) or the action of JNK has been identified in Myc-mediated apoptosis. Here, we found that JNK selectively interacted with, and phosphorylated, c-Myc at Ser-62 and Ser-71 as confirmed with phospho-c-Myc-specific antibodies. Interestingly, dominant negative mutant JNK(APF) impaired the c-Myc-dependent apoptosis, but not mutated c-Myc (S62A/S71A)-dependent apoptosis triggered by UV irradiation. Furthermore, c-Myc (S62A/S71A)-expressing NIH3T3 cells were not sensitized like wild type c-Myc-expressing NIH3T3 cells to JNK-activating apoptotic stimuli, such as UV and Taxol. These results indicate that the JNK pathway is selectively involved in the c-Myc-mediated apoptosis and that the apoptotic function of c-Myc is directly regulated by JNK pathway through phosphorylation at Ser-62 and Ser-71. PMID: 10551811 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 276: Toxicol Appl Pharmacol. 1999 Nov 1;160(3):262-70. Selective activation in the MAPK pathway by Hg(II) in precision-cut rabbit renal cortical slices. Turney KD, Parrish AR, Orozco J, Gandolfi AJ. College of Pharmacy, University of Arizona, Tuscon, Arizona, 85721, USA. The kidneys are the primary organ for the accumulation and toxicity of inorganic mercury. In these studies the molecular response of precision-cut rabbit renal cortical slices to low levels of inorganic mercury was examined. Cortical slices (275 microm) were obtained from 1.0 kg NZW rabbits and exposed to mercuric chloride [Hg(II)] at concentrations of 0.01-10 microM for 2-8 h. Overt cytotoxicity, as assessed by intracellular K(+) levels, was not observed following exposure to these concentrations of Hg(II). However, an induction of heme-oxygenase-1 (Hsp32) was seen following a 2-h challenge to Hg(II). A dose-dependent induction of the DNA binding activity of the AP-1 transcription factor after 4 h of Hg(II) exposure correlated with a dose-dependent enhancement of c-jun gene expression following 2 h of Hg(II) exposure. Additionally, an increase in phosphorylated c-Jun NH(2)-terminal protein kinase (JNK) was observed following 2 h of Hg(II) exposure. These results suggest activation of the mitogen-activated protein (MAP) signal transduction pathway, specifically the c-Jun NH(2)-terminal protein kinase (JNK) pathway. No changes were observed, however, in the DNA binding activity of ATF2 and Elk-1, transcription factors involved in both the JNK and p38 pathways of MAP signal transduction, nor in the gene expression of c-myc. This selectivity of alterations in molecular signaling suggests an acute response in signal transduction, specifically activation of the JNK pathway in renal tissue following exposure to nanomolar concentrations of Hg(II). Copyright 1999 Academic Press. PMID: 10544060 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 277: Proc Assoc Am Physicians. 1999 Sep-Oct;111(5):438-47. Heme oxygenase: recent advances in understanding its regulation and role. Elbirt KK, Bonkovsky HL. Department of Medicine, University of Massachusetts Medical School, Worcester, USA. Heme oxygenase (HO) is responsible for the physiological breakdown of heme into equimolar amounts of biliverdin, carbon monoxide, and iron. Three isoforms (HO-1, HO-2, and HO-3) have been identified. HO-1 is ubiquitous and its mRNA and activity can be increased several-fold by heme, other metalloporphyrins, transition metals, and stimuli that induce cellular stress. HO-1 is recognized as a major heat shock/stress response protein. Recent work from our laboratory has demonstrated several potential consensus regulatory elements in the 5'-untranslated region (UTR) of HO-1, including activator protein 1 (AP-1), metal responsive element (MRE), oncogene c-myc/max heterodimer binding site (Myc/Max), antioxidant response element (ARE), and GC box binding (Sp1) sites. Using deletion-reporter gene constructs, we have mapped sites that mediate the arsenite-dependent induction of HO-1, and we have shown that components of the extracellular signal-regulated kinase (ERK) and p38 (a homologue of the yeast HOG1 kinase), but not c-jun N-terminal kinase (JNK), mitogen-activated protein (MAP) kinase pathways are involved in arsenite-dependent upregulation. In contrast, HO-2 is present chiefly in the brain and testes and is virtually uninducible. HO-3 has very low activity; its physiological function probably involves heme binding. Products of the HO reaction have important effects: carbon monoxide is a potent vasodilator, which is thought to play a key role in the modulation of vascular tone, especially in the liver under physiological conditions, and in many organs under "stressful" conditions associated with HO-1 induction. Biliverdin and its product bilirubin, formed in most mammals, are potent antioxidants. In contrast, "free" iron increases oxidative stress and regulates the expression of many mRNAs (e.g., DCT-1, ferritin, and transferrin receptor) by affecting the conformation of iron regulatory protein (IRP)-1 and its binding to iron regulatory elements (IREs) in the 5'- or 3'-UTRs of the mRNAs. Publication Types: Review Review, Tutorial PMID: 10519165 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 278: Endocr J. 1999 Jun;46(3):383-8. Effects of PMA and transcription factors on ovine interferon-tau transactivation in various cell lines. Yamaguchi H, Ikeda Y, Taylor A, Katsumura M, Miyazawa T, Takahashi E, Imakawa K, Sakai S. Laboratory of Animal Breeding, Faculty of Agriculture, University of Tokyo, Japan. Interferon-tau (IFNtau) is produced by the trophectoderm of ruminant ungulates and its gene transactivation in vitro has so far been achieved only in human choriocarcinoma cells, JAR and JEG3. To examine if ovine IFNtau gene transactivation could be induced in cells other than JAR or JEG3 cells and its activation could be aided by the expression of a protooncogene(s), a transient transfection system was developed with the upstream region of ovine IFNtau gene that had been inserted into the chloramphenicol acetyltransferase (CAT) reporter plasmid (IFNtau-CAT). The effect of a protein kinase C (PKC) activator, phorbol 12-myristate 13-acetate (PMA), on IFNtau-CAT transcriptional activity was examined in JEG3, human embryonic kidney (293), HeLa and Vero cells. Upon transfection and PMA treatment, ovine IFNtau gene was transactivated in two unrelated cell lines, JEG3 and 293 cells. Since IFNtau-CAT was not induced in HeLa or Vero cells, HeLa and JEG3 cells were further examined for their ability to support IFNtau-CAT transactivation in a co-transfection system. While the expression of c-myc, interferon regulatory factor 1 or 2 (IRF-1 or IRF-2) was not effective, CAT activity was strongly enhanced in both JEG3 and HeLa cells with the co-transfection of c-Jun or c-Jun plus c-Fos. These data suggest that ovine IFNtau gene transcription induced by PMA is not specific for trophoblast cells and a protooncogene, c-jun, is a downstream effector of PMA activated nuclear factors in its signal transduction cascade resulting in IFNtau gene transactivaion. PMID: 10503990 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 279: J Cell Physiol. 1999 Nov;181(2):342-54. Immediate early gene expression in dog thyrocytes in response to growth, proliferation, and differentiation stimuli. Deleu S, Pirson I, Clermont F, Nakamura T, Dumont JE, Maenhaut C. Institute of Interdisciplinary Research, School of Medicine, University of Brussels, Brussels, Belgium. sdeleu@ulb.ac.be In dog thyroid cells, insulin or IGF-1 induces cell growth and is required for the mitogenic action of TSH through cyclic AMP, of EGF, and of phorbol esters. HGF per se stimulates cell proliferation and is thus the only full mitogenic agent. TSH and cAMP enhance, whereas EGF phorbol esters and HGF repress differentiation expression. In this study, we have investigated for each factor and regulatory cascade of the intermediate step of immediate early gene induction, that is, c-myc, c-jun, jun D, jun B, c-fos, fos B, fra-1, fra-2, and egr1; fra-1 and fra-2 expressions were very low. TSH or forskolin increased the levels of c-myc, jun B, jun D, c-fos, and fos B while decreasing those of c-jun and egr1. Phorbol myristate ester stimulated the expression of all the genes. EGF and HGF stimulated the expression of all the genes except jun D and for EGF fos B. All these effects were obtained in the presence and in the absence of insulin, which shows that insulin is not necessary for the effects of the mitogens on immediate early gene expression. The definition of the repertoire of early immediate genes inductible by the various growth cascades provides a framework for the analysis of gene expression in tumors. (1) Insulin was able to induce all the protooncogenes investigated except fos B. This suggests that fos B could be the factor missing for insulin to induce mitogenesis. (2) No characteristic pattern of immediate early gene expression has been observed for insulin, which induces cell hypertrophy and is permissive for the action of the other growth factors. These effects are therefore not accounted for by a specific immediate early gene expression. On the other hand, insulin clearly enhances the effects of TSH, phorbol ester, and EGF on c-myc, junB, and c-fos expression. This suggests that the effect of insulin on mitogenesis might result from quantitative differences in the transcription complexes formed. (3) c-myc, c-fos, and jun B mRNA induction by all stimulating agents, whether inducing cell hypertrophy, or growth and dedifferentiation, or growth and differentiation, suggests that, although these expressions are not sufficient, they may be necessary for the various growth responses of thyroid cells. (4) The inhibition of c-jun and egr1 mRNA expression, and the marked induction of jun D mRNA appear to be specific features of the TSH cAMP pathway. They might be related to its differentiating action. (5) fos B, which is induced by TSH, forskolin, phorbol ester, and HGF but not by insulin, could be involved in the mitogenic action of the former factors. Copyright 1999 Wiley-Liss, Inc. PMID: 10497313 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 280: Oncogene. 1999 Aug 26;18(34):4777-87. Suppression of anchorage-independent growth of human cancer cell lines by the drs gene. Yamashita A, Hakura A, Inoue H. Department of Tumor Virology, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan. We have previously reported that the drs gene, whose mRNA expression is downregulated by retroviral oncogenes such as v-src and v-K-ras, has the ability to suppress transformation by v-src in a rat cell line F2408. We have now isolated a human homolog of this gene (h-drs) and found that the expression of h-drs mRNA is markedly downregulated in a variety of human cancer cell lines including those of the colon, bladder, and ovary. To investigate the function of the drs gene as a tumor suppressor in human cancer cells, we constructed recombinant amphotropic retrovirus containing the drs gene, introduced this virus into human cancer cell lines whose drs expression was downregulated and found that drs has the ability to suppress anchorage-independent growth of these cells without disturbing cell proliferation. Analyses with deletion mutants of the drs gene revealed that both the C-terminal region inside the transmembrane domain and three consensus repeats in the N-terminal region are essential for the suppression of anchorage-independent growth of the cells. We also found that the G1-S progression of the cell cycle and expression of cyclin A mRNA were significantly suppressed in T24 cells expressing the drs gene under non-adhesion culture conditions. In contrast, the expression of cyclin D and E and the phosphorylation of Rb protein were not affected by ectopic expression of the drs gene, suggesting that an Rb-independent downregulation of cyclin A is involved in the suppression of anchorage-independent growth by means of the drs gene. PMID: 10490811 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 281: Zhonghua Gan Zang Bing Za Zhi. 1999 Jun;7(2):110-2. [Abnormal expression of several oncogenes during hepatocarcinogenesis in WHV/myc transgenic mice] [Article in Chinese] Liu P, Zhao Z, Dominique B. First Affiliated Hospital of Nanjing Medical University. OBJECTIVE: To explore at cellular level the abnormal expression of related oncogenes during hepatocarcinogenesis in the WHV/myc transgenic mice. METHODS: With specific molecular probes, in situ hybridization was used to detect the expression of c-myc transgene,IGF-II gene as well as c-jun, c-fos, and c-H-ras oncogenes in the liver sections from WHV/myc transgenic mice in different stages of the carcinogenic process. RESULTS: The overexpression of c-myc transgene and IGF-II gene transcripts were detected in liver sections from the earlier neonatal transgenic mice. Then, the expression levels of these two genes were rapidly decreased and undetectable. At the time of the liver tumor development, the expression of c-myc transgene and IGF-II gene resumed. The c-jun, c-fos and c-H-ras genes expressed more intensely in the livers from preneoplastic WHV/myc mice than from normal mice. CONCLUSION: The overexpressions of c-myc transgene and IGF-II gene in the earlier neonatal stage and tumoral stage may be of importance for the development and maintenance of a transformed phenotype. During the prenoplastic "silent" period of c-myc transgene expression in WHV/myc transgenic mice, several oncogenes are overexpressed in the livers. PMID: 10488423 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 282: Mol Carcinog. 1999 Sep;26(1):10-9. Inhibition of rat mammary tumorigenesis by human chorionic gonadotropin associated with increased expression of inhibin. Srivastava P, Russo J, Russo IH. Breast Cancer Research Laboratory, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA. This work was designed to test whether the suppression of 7,12-dimethylbenz[a]anthracene (DMBA)-induced rat mammary carcinomas by human chorionic gonadotropin (hCG) is associated with the synthesis of inhibin. For this purpose, virgin rats received 8 mg DMBA/100 g body weight; 20 d later they were injected daily with 100 IU of hCG for 40 d (DMBA+hCG group). Age-matched untreated (control), hCG-, and DMBA+saline-treated rats were used for comparisons. Mammary tissues were collected for histopathological and mRNA analyses after 5, 10, 20, and 40 d of hCG injection and 20 d after treatment. None of the animals in the control and hCG-treated groups developed mammary tumors. DMBA-treated rats developed a high incidence of both microscopic lesions, i.e., intraductal proliferations and ductal carcinomas in situ, and palpable tumors. In DMBA+hCG-treated rats, the incidence of microscopic and palpable tumors was markedly reduced. In these animals, alpha- and beta-inhibin immunoreactivity was elevated in the non-tumoral mammary glands in association with lobule formation and in the tumors. Inhibin A and B mRNAs were also elevated in the mammary tissue, and c-myc and c-jun were induced by the hormonal treatment. DMBA alone did not modify the expression of these genes. Our findings indicate that inhibin production and gene activation are associated with both mammary gland differentiation and tumor regression. PMID: 10487517 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 283: Endocr J. 1999 Apr;46(2):253-61. Regulation of c-fos gene induction and mitogenic effect of transforming growth factor-beta1 in rat articular chondrocyte. Osaki M, Tsukazaki T, Yonekura A, Miyazaki Y, Iwasaki K, Shindo H, Yamashita S. Department of Orthopaedic Surgery, Nagasaki University School of Medicine, Japan. We have previously reported that type I transforming growth factor beta (TGF-beta1) is a potent stimulator of cell growth in articular chondrocytes. In this study, we examined the mechanism of TGF-beta1 induced cellular proliferation by using cultured rat articular chondrocytes (CRAC). A time-course study of [3H]thymidine incorporation upon TGF-beta1 (1 ng/mL) or 10% fetal bovine serum stimulation revealed that TGF-beta1 directly stimulates DNA synthesis in CRAC. Pretreatment with H7, an inhibitor for protein kinase C (PKC), completely blocks TGF-beta1-induced proliferation. Since TGF-beta1 has been shown to transduce signals through MAP kinase cascades, we investigated the induction of several protooncogenes by Northern blotting. TGF-beta1 addition causes an immediate and transient induction of c-fos but not myc or jun mRNA. Furthermore, this c-fos expression is not inhibited by cycloheximide, but is completely abolished by pretreatment with TPA, so that the c-fos gene is a direct target of TGF-beta1 signalling and PKC is involved in this c-fos induction. To refine our understanding of TGF-beta1 regulation of the c-fos promoter region, we performed chloramphenicol acetyltransferase (CAT) assays. A serial deletion analysis of the c-fos promoter region reveals a TGF-beta1 responsive element in a region between -403 and -329 bp upstream of the transcription initiation site. We attempted gel shift assays on this response element with CRAC nuclear extracts. Although this region contains a sis-inducible binding element, we fail to detect specific DNA-protein complexes. Our results, however, suggest that TGF-beta1 acts as a primary mitogen in CRAC and this mitogenic activity requires PKC activation. Finally, the subsequent induction of c-fos expression occurs through an as yet unidentified transactivation mechanism. PMID: 10460009 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 284: Toxicol Sci. 1999 Jul;50(1):98-105. Enhanced transcription factor DNA binding and gene expression induced by arsenite or arsenate in renal slices. Parrish AR, Zheng XH, Turney KD, Younis HS, Gandolfi AJ. Department of Anesthesiology, College of Medicine, University of Arizona, Tucson 85721, USA. Although the kidney represents a target for the accumulation and toxicity of arsenic, little is known about the molecular targets of arsenic in this organ. Therefore, these studies were designed to examine the molecular impact of arsenite [As(III)] and arsenate [As(V)] at low (nanomolar) concentrations. Precision-cut rabbit renal cortical slices were challenged with As(III) or As(V) for up to 8 h. Neither form of the metal induced overt cytotoxicity as assessed by intracellular K+ levels over this time period at concentrations from 0.01-10 microM. In addition, no alterations in the expression of Hsp 60, 70, or 90 were observed. However, induction of heme oxygenase-1 (Hsp 32) was seen following a 4-h challenge with As(III), but not with As(V). As(III) and As(V) induced DNA binding of AP-1 at 2- and 4-h exposure; following a 6-h exposure there was no difference. Although no alteration in the DNA binding activity of ATF-2 was induced by As(III) or As(V), both forms enhanced the DNA binding activity of Elk-1. Enhanced DNA binding activity of AP-1 and Elk-1 correlated with increased gene expression of c-fos, but not c-jun, at 2 h. c-myc gene expression was also induced by As(III) and As(V), albeit at a later time point (6 h). These results suggest that acute arsenic challenge, by either As(III) or As(V), is associated with discrete alterations in the activity of signaling pathways and gene expression in renal tissue. PMID: 10445758 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 285: Calcif Tissue Int. 1999 Aug;65(2):133-8. Molecular characterization of gene expression changes in ROS 17/2.8 cells cultured in diffusion chambers in vivo. Onyia JE, Hale LV, Miles RR, Cain RL, Tu Y, Hulman JF, Hock JM, Santerre RF. Skeletal Disease Research Group, 0403, Endocrine Division, Lilly Research Labs, Indianapolis, Indiana 46285 USA. Transplantation of diffusion chambers (DC) containing osteoblast-like cells to extraskeletal sites has been highly studied and proven to be a useful technique to investigate the process of osteoblast differentiation and bone formation. To investigate the molecular basis of osteogenesis in DC, we examined the temporal pattern of gene expression of the proliferation marker histone H4, immediate early response genes (IEGs), c-fos, c-jun, c-myc, osteoblast phenotype-associated genes, osteocalcin (OC), osteopontin (OP), type I collagen (COL1A1), alkaline phosphatase (ALP), parathyroid hormone receptor (PTHR) and matrix modifying enzyme, matrix metalloproteinase-9 (MMP-9). DC containing ROS 17/2.8 were implanted intraperitoneally into rat hosts and cultured in vivo for various times up to 56 days. Histological analysis of von Kossa stained sections of the DC contents showed a well-organized connective tissue and the production of mineralized matrices/nodules. In contrast, histological examination of DC containing Rat-2 fibroblast cells revealed the lack of an organized mineralized matrix. Molecular analysis of DC containing ROS 17/2.8 cells at 0, 3, 10, 28, and 56 days demonstrated a time-dependent decrease in DNA content associated with cell death. In the surviving cells, an increase in histone H4 mRNA (consistent with an increase in cell proliferation) was evident by 3-10 days and thereafter expression returned to control levels. In vitro, ROS 17/2.8 cells expressed detectable levels of c-fos, c-jun, c-myc, OC, OP, ALP, COL1A1, and PTHR but not MMP-9. In vivo, the expression of c-fos increased 2-fold in 3-28 days and by 56 days was 4-5 fold above control levels. In 3-10 days, c-jun expression increased 1.6-1.8-fold above control levels. In contrast, by day 28, c-jun expression decreased to control levels, but increased to 2.1-fold above control by 56 days. c-myc mRNA expression increased 3-fold within 3 days and then dropped to below control values by 10-56 days. After transplantation in vivo, the expression of OC and PTHR decreased to undetectable levels. Similarly, ALP mRNA decreased to or = 4 mm in diameter), invasiveness (smooth vs. invading margins), and other properties (piling vs. spread) induced by 3-methylcholanthrene in Balb/c-3T3 cells. The transformed focal cells were used in in vitro studies including anchorage-independent analysis, focal reconstruction, gene transfection using NIH-3T3 host cells, and Southern blotting to assess amplification of five proto-oncogenes (K-ras, H-ras, c-fos, c-jun, c-myc) and a tumor suppressor (p53) gene. Results showed that 1) there was a significant increase in anchorage-independent growth of all five types of foci ranging from 7-12%; 2) all five morphological types of transformed foci showed 8-15% focal reconstruction; 3) DNA from all five types of transformed foci induced transformation in NIH-3T3 cells at a level significantly above the control DNA; 4) gene amplification studies indicated amplification in both K-ras and H-ras proto-oncogenes; however, c-fos, c-jun, and c-myc did not show DNA amplification. The tumor suppressor gene (p53) was activated and the increase was up to 3-fold over the normal Balb/c-3T3 DNA. These findings are consistent with our hypothesis that all five morphologically different foci have preneoplastic potential and that any foci of size > or = 2 mm regardless of invasiveness and piling should be scored as positive during the transformation assay. PMID: 9882012 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 319: Oncogene. 1998 Dec 3;17(22):2901-13. Structure and transcriptional regulation of BKJ, a novel AP-1 target gene activated during jun- or fos-induced fibroblast transformation. Hartl M, Bister K. Institute of Biochemistry, University of Innsbruck, Austria. The BKJ gene was originally identified based on its specific transcriptional activation in jun-transformed avian fibroblasts. We now show that BKJ is a direct transcriptional target of the AP-1 transcription factor components Jun and Fos. The complete structural organization of the quail BKJ gene was determined by nucleotide sequence analysis and transcriptional mapping. The gene contains three exons with the coding region confined to the third exon. A major mRNA species of 0.8 kb and a minor one of 1.3 kb are produced by variable usage of two transcriptional initiation sites. The BKJ promoter region contains two authentic AP-1 binding sites. By transactivation of reporter gene constructs and direct binding of Jun recombinant protein, the proximal AP-1 element was shown to be essential for BKJ promoter activation. Using polyclonal antiserum directed against recombinant BKJ protein, the activation of BKJ in jun-transformed avian fibroblasts was also demonstrated at the protein level. BKJ is a novel gene related to the avian beta-keratin gene family whose members display highly specific expression patterns during embryogenesis and epidermal development. Activation of BKJ in fibroblasts by retroviral or deregulated cellular jun or fos alleles may contribute to cell transformation. PMID: 9879996 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 320: Biochim Biophys Acta. 1998 Dec 22;1443(3):334-42. Absence of 60-Hz, 0.1-mT magnetic field-induced changes in oncogene transcription rates or levels in CEM-CM3 cells. Jahreis GP, Johnson PG, Zhao YL, Hui SW. Membrane Biophysics Laboratory, Department of Molecular and Cellular Biophysics, Roswell Park Cancer Institute, Buffalo, NY 14263, USA. Our objective was to assess the reproducibility of the 60-Hz magnetic field-induced, time-dependent transcription changes of c-fos, c-jun and c-myc oncogenes in CEM-CM3 cells reported by Phillips et al. (Biochim. Biophys. Acta, 1132 (1992) 140-144). Cells were exposed to a 60-Hz magnetic field (MF) at 0.1 mT (rms), generated by a pair of Helmholtz coils energized in a reinforcing (MF) mode, or to a null magnetic field when the coils were energized in a bucking (sham) mode. After MF or sham exposure for 15, 30, 60 or 120 min, nuclei and cytoplasmic RNA were extracted. Transcription rates were measured by a nuclear run-on assay, and values were normalized against either their zero-time exposure values, or against those of the c-G3PDH (housekeeping) gene at the same time points. There was no significant difference, at P=0.05, detected between MF and either sham-exposed or control cells at any time point. Transcript levels of the oncogenes were measured by Northern analysis and normalized as above. No significant difference (P=0.05) in transcript levels between MF and either sham-exposed or control cells was detected. PMID: 9878814 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 321: Cell Growth Differ. 1998 Dec;9(12):983-8. Inhibition of G1 cyclin-dependent kinase activity in cell density-dependent growth arrest in human fibroblasts. Afrakhte M, Heldin NE, Westermark B. Department of Genetics and Pathology, University Hospital, Uppsala, Sweden. The growth of normal fibroblasts in culture ceases as the cells reach saturation density. Although cells in dense cultures express functionally active growth factor receptors, they are essentially refractory to the mitogenic activity of growth factors. Northern blot analysis revealed that immediate early genes, c-myc, c-fos and c-jun are induced by mitogen in dense cultures. However, these cells fail to express the late G1 genes as E2F-1, cdc25A, and cyclin A in response to mitogen stimulation. Furthermore, because pRb-phosphorylation is a key event in G1 progression, here we show that in dense cultures, pRb remains in its active (hypophosphorylated) form after stimulation by mitogens. We also show that the kinase activity of cyclin-dependent kinases that are indispensable for the phosphorylation of pRb in late G1 phase was decreased on increasing cell density. The reduced kinase activity may be caused by the observed increase in cyclin-dependent kinase inhibitors and the reduction of cdc25A expression in dense cells. PMID: 9869298 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 322: Blood. 1999 Jan 1;93(1):260-7. Constitutive and interleukin-7/interleukin-15 stimulated DNA binding of Myc, Jun, and novel Myc-like proteins in cutaneous T-cell lymphoma cells. Qin JZ, Dummer R, Burg G, Dobbeling U. Department of Dermatology, University Hospital Zurich, Zurich, Switzerland. Members of the Myc and Jun/Fos gene families have been found to be expressed in late stages of cutaneous T-cell lymphoma (CTCL) and may be responsible for the transition from low-grade to high-grade tumors. The composition of these complexes is an important parameter, as the different homo- and heterodimeric jun and myc complexes can have gene transcription activating or suppressing activities. We determined the composition of the jun and myc DNA-binding complexes in three CTCL cell lines and malignant cells of seven Sezary patients by electrophoretic mobility shift assays (EMSAs) and "supershift" assays in which specific antibodies against the different members of the tested gene families were included in the binding reactions. Complexes containing JunD were found in three cell lines and two patients. The three cell lines and one patient contained also c-Myc/Max heterodimers. Because c-Myc/Max heterodimers are strong gene transcription activators and are necessary for cell-cycle progression, they may play a role in the progression of CTCL. JunD may also promote cell-cycle progression and influence the expression of cell death survival genes. Interleukin-7 (IL-7) and IL-15, which have been identified as growth factors for CTCL cells, stimulated the DNA binding of JunD and two novel c-Myc recognition site (E-box) binding proteins, but not the DNA binding of c-Myc/Max heterodimers. PMID: 9864169 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 323: Mol Cell Biol. 1999 Jan;19(1):889-98. Cyclosporin A promotes translational silencing of autocrine interleukin-3 via ribosome-associated deadenylation. Nair AP, Hirsch HH, Colombi M, Moroni C. Institute for Medical Microbiology, University of Basel, Basel, Switzerland. Translation is regulated predominantly by an interplay between cis elements at the 3' and 5' ends of mRNAs and trans-acting proteins. Cyclosporin A (CsA), a calcineurin antagonist and blocker of interleukin-2 (IL-2) transcription in T cells, was found to inhibit translation of IL-3 mRNA in autocrine mast cell tumor lines. The mechanism involved ribosome-associated poly(A) shortening and required an intact AU-rich element in the 3' untranslated region. FK506, another calcineurin inhibitor, shared the effect. The translational inhibition by CsA was specific to oncogenically induced lymphokines IL-3 and IL-4 but not to IL-6, c-jun, and c-myc, which are expressed in the nonmalignant precursor cells. Furthermore, no translational down-regulation of the mRNA was observed in IL-3-transfected precursor cells. These data suggest that translational silencing is associated with the tumor phenotype. PMID: 9858612 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 324: Hematol Cell Ther. 1998 Oct;40(5):217-21. Molecular pathophysiology of chronic myelogenous leukemia. Turhan AG, Solary E, Vainchenker W, Dusanter-Fourt I. Translational Research-Cell Therapy Laboratory-INSERM U362, Institut Gustave Roussy, Villejuif, France. It is currently well established that chronic myelogeneous leukemia (CML) results from the activation of multiple signalling pathways by the Philadelphia chromosome (Ph1) and its molecular counterpart, the BCR-ABL oncogene. Deletion and site-directed mutagenesis experiments have determined the critical regions of the oncogene for its interaction with major signalling pathways but the roles of the latter in the resulting leukemic phenotypes are not well understood. Several major signalling pathways shown to be activated by BCR-ABL, including RAS, MYC, JUN, STAT, PI-3K and NF-KB are briefly discussed in this paper. Other signalling molecules are also clearly involved, including p62-DOK, p95-VAV, CRK-L, p12O-CBL and focal adhesion proteins. Recent experimental evidence also indicates that negative regulatory proteins could be activated in cells expressing BCR-ABL and their inhibition during the course of the disease could play a role in the progression towards the acute phase. We finally discuss the evidence indicating that at least in experimental systems BCR-ABL has a clear anti-apoptotic activity and that BCR-ABL achieves this effect by acting upstream of the procaspase-3. Publication Types: Review Review, Tutorial PMID: 9844814 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 325: DNA Cell Biol. 1998 Nov;17(11):951-6. Insulin activates the hepatitis B virus X gene through the activating protein-1 binding site in HepG2 cells. Choi BH, Park CJ, Rho HM. Department of Molecular Biology and Research Center for Cell Differentiation, Seoul National University, Korea. Insulin stimulates cellular oncogenic activators such as c-jun, c-fos, and c-myc; and hepatitis B virus (HBV) X, a viral transactivator, is known to induce liver cancer in transgenic mice. In this respect, the effect of insulin on the expression of HBx protein was investigated in HepG2 cells. Insulin-stimulated transcription from the HBV X promoter in a dose-dependent manner was assessed by chloramphenicol acetyltransferase (CAT) assay. A mutation preventing AP-1 binding to the E element abolished the activation of the HBV X promoter by insulin. In addition, insulin stimulated the minimal thymidine kinase (tk) gene promoter activity through both the HBV E element and the consensus AP-1 binding site in HepG2 cells. An electrophoretic mobility shift assay (EMSA) using insulin-treated HepG2 nuclear extracts showed that insulin actually enhanced the binding of nuclear proteins to the HBV E element as well as to the consensus AP-1 binding site. Both HBV E and AP-1 oligonucleotides were effective competitors for this binding. These results showed that insulin elevated the expression of HBx protein through the AP-1 binding site of HBV EnI. We suggest that insulin can augment the role of HBx in the development of hepatocellular carcinoma (HCC) in HBV-infected liver, probably through interaction with other cellular oncogenes. PMID: 9839804 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 326: Scand J Immunol. 1998 Nov;48(5):551-6. Differential oncogene and TNF-alpha mRNA expression in bone marrow cells from systemic lupus erythematosus patients. Alvarado-de la Barrera C, Alcocer-Varela J, Richaud-Patin Y, Alarcon-Segovia D, Llorente L. Department of Immunology and Rheumatology, Instituto Nacional de la Nutricion Salvador Zubiran, Tlalpan, Mexico. The aim of the study was to investigate the bone marrow expression of genes involved in cell growth and apoptosis in patients with systemic lupus erythematosus (SLE). Spontaneous expression of bcl-2, bax, c-myc. c-fos, c-jun, p53, Fas and tumour necrosis factor (TNF)-alpha by bone marrow cells was measured using either semiquantitative or quantitative reverse transcription polymerase chain reaction in SLE patients (n = 8) and in eight normal control subjects. The expression of bcl-2 was found to be higher in SLE patients than in controls. Bone marrow cells from SLE patients showed significant down-regulation of bax, c-myc, c-fos and p53 (P < or = 0.05), as compared to normal controls. In both SLE patients and controls the expression of c-jun and Fas was very low or undetectable. Finally, TNF-alpha gene expression was higher in bone marrow samples from SLE patients than in those of controls (P= 0.01). The abnormal expression of genes regulating cell growth and apoptosis in bone marrow cells from SLE patients may help explain the presence of autoreactive cells in secondary lymphoid organs and peripheral blood of SLE patients. PMID: 9822266 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 327: Clin Cancer Res. 1996 May;2(5):847-54. Paclitaxel induces programmed cell death in MDA-MB-468 human breast cancer cells. McCloskey DE, Kaufmann SH, Prestigiacomo LJ, Davidson NE. The Johns Hopkins Oncology Center, Johns Hopkins University School of Medicine, Baltimore, Maryland 21231, USA. The ability of paclitaxel, one of the most active chemotherapeutic agents against breast cancer, to induce programmed cell death in hormone-independent MDA-MB-468 human breast cancer cells was assessed. Treatment of MDA-MB-468 cells led to growth inhibition, high-molecular-weight and oligonucleosomal DNA fragmentation, and apoptosis-associated morphological changes after either 3- or 24-h exposure to paclitaxel concentrations >/=10 nM. Additionally, cleavage products of poly(ADP-ribose) polymerase and lamin B1, two proteins that are cleaved early in the execution phase of programmed cell death, were detected. Quantitative studies indicated that exposure to paclitaxel for 24 h resulted in more DNA fragmentation than did 3-h exposure. Rapid induction of the early-response gene c-jun but not c-myc was associated with paclitaxel treatment. The ability of paclitaxel to induce high-molecular-weight DNA fragmentation and apoptosis-associated morphological changes in three other breast cancer cell lines was also established. These data suggest that paclitaxel, an agent known to stabilize microtubules and prevent cell division but not to act directly on DNA, induces programmed cell death in breast cancer cells. PMID: 9816240 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 328: Clin Cancer Res. 1995 Feb;1(2):207-14. PAX5 expression correlates with increasing malignancy in human astrocytomas. Stuart ET, Kioussi C, Aguzzi A, Gruss P. Department of Molecular Cell Biology, Max-Planck Institute for Biophysical Chemistry, Am Fassberg, 37077 Gottingen, Germany. Rearrangements concerning chromosome 9p are a late event in the progression of human astrocytic tumors to their most malignant form. The expression of PAX5, which maps to chromosome 9p13, was studied in primary human brain tumors of astrocytic origin. Whereas murine Pax5 is not expressed in the forebrain at any stage, PAX5 expression was increased in a range of astrocytomas (WHO grades II-IV) which originated in the forebrain. Expression of PAX5 was limited to those cells which also expressed the oncogenes myc, fos, or jun singularly or in combination. The epidermal growth factor receptor was highly expressed in glioblastoma multiform tumors in areas which were also highly PAX5 positive. We conclude that the missappropriate expression of PAX5 may aid in promoting the progression of astrocytomal malignancy. PMID: 9815975 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 329: Int Immunol. 1998 Oct;10(10):1421-33. IL-4 and IL-13 specifically increase adhesion molecule and inflammatory cytokine expression in human lung fibroblasts. Doucet C, Brouty-Boye D, Pottin-Clemenceau C, Jasmin C, Canonica GW, Azzarone B. U268 INSERM Hopital Paul Brousse, Villejuif, France. Subepithelial fibrosis in the bronchi of asthmatics is the result of an irreversible lung fibroblast activation, triggered by cytokines secreted by IL-4- and IL-5-activated inflammatory cells. Here, we provide evidence that human lung fibroblasts (ICIG7 cells) express a single class of high-affinity IL-4 receptor (IL-4R). This receptor is functional and composed of at least the IL-4Ralpha and IL-13Ralpha1 chains in the absence of the IL-2Rgamma chain. The IL-4Ralpha is efficiently internalized at 37 degrees C within 15 min in the presence of IL-4, whereas this process is slower with IL-13. In ICIG7 cells, IL-4 triggers the tyrosine phosphorylation of at least two proteins (110 and 180 kDa), and up-regulates the transcription of c-fos, c-jun and c-myc proto-oncogenes. In addition, the secretion of several cytokines [IL-6, granulocyte colony stimulating factor and granulocyte macrophage colony stimulating factor (GM-CSF)] as well as the expression of beta1 integrin and VCAM-1 adhesion molecules are augmented by IL-4. IL-13 displays similar biological activities, but less effectively than IL-4. On the other hand, ICIG7 cells could constitute a lung fibroblast population defined by the spontaneous release of several pro-inflammatory cytokines (IL-6, IL-11 and GM-CSF) and cell surface phenotype (CD4 and Thy-1). Through this peculiar cytokine pattern and the IL-4/IL-13-dependent activities, these cells could act as effector cells in the pathogenesis of asthma, triggering and maintaining the recruitment, homing and activation of bone marrow-derived inflammatory cells, and playing a role in the remodeling process of the airways. PMID: 9796908 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 330: Mol Cell Biochem. 1998 Oct;187(1-2):211-20. Biological effects of a relatively low concentration of 1-beta-D-arabinofuranosylcytosine in K562 cells: alterations of the cell cycle, erythroid-differentiation, and apoptosis. Yamada H, Horiguchi-Yamada J, Nagai M, Takahara S, Sekikawa T, Kawano T, Itoh K, Fukumi S, Iwase S. Department of Internal Medicine (IV), Aoto Hospital, Institute of DNA Medicine, The Jikei University School of Medicine, Tokyo, Japan. Therapeutic strategies for leukemia are directed to induction of differentiation and apoptosis as well as growth inhibition. One of the key antileukemic agents, 1-beta-D-arabinofuranosylcytosine (ara C), is clinically applied according to these therapeutic aims. However, the molecular effects of 0.1 microg/ml of ara C, a concentration that corresponds to the serum level in leukemic patients on a conventional dose of ara C, have not been well disclosed. Here, we addressed these issues using K562 cells which derived from a blastic crisis of chronic myeloid leukemia. DNA synthesis of treated cells was suppressed from 1-6 h. But, it recovered at 12 h and no further inhibition was observed. The number of cells was not decreased but DNA fragmentation was observed at 72 h. The number of erythroid-differentiated cells also increased to 30% at 72 h. Along with treatment, no marked alteration of mRNAs for cell cycle-regulating genes was found and the retinoblastoma gene product remained hyperphosphorylated throughout treatment. The expression of mRNAs for apoptosis-regulating genes also remained unchanged, except for slight down-regulation of Bax. c-myc protein was not found later than 48 h, and Max mRNA was downregulated. c-jun was immediately induced, followed by the fluctuated expression level along with treatment. These findings suggest that the 0.1 microg/ml ara C changed the proliferation, differentiation and death of K562 cells in a biphasic manner. In the early phase, DNA synthesis was inhibited without altering the expression of cell cycle regulating-genes. In the latter phase, cell death and erythroid- differentiation occurred in accordance with the down-regulation of c-myc. PMID: 9788759 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 331: Oncogene. 1998 Sep 10;17(10):1189-94. Normal development, oncogenesis and programmed cell death. Liebermann DA. Fels Institute for Cancer Research and Molecular Biology, Philadelphia, Pennsylvania 19140, USA. Meeting's Report -- June 2, 1998, Sugarload Estate Conference Center, Philadelphia, Pennsylvania, USA. A symposium on Normal Development, Oncogenesis and Programmed Cell Death, was held at the Sugarload Estate Conference Center, Philadelphia, Pennsylvania, USA sponsored by the Fels Cancer Institute, Temple University School of Medicine, with the support of the Alliance Pharmaceutical Corporation. The symposium was organized by Drs Dan A Liebermann and Barbara Hoffman at the Fels. Invited speakers included: Dr Andrei V Gudkov (University of Illinois) who started the symposium talking about 'New cellular factors modulating the tumor suppressor function of p53'; Dr Yuri Lazebnik (Cold Spring Harbor Laboratories) spoke about 'Caspases considered as enemies within'; Dr E Premkumar Reddy (Fels Institute, Temple University) talked about recent exciting findings in his laboratory regarding 'JAK-STATs dedicated signaling pathways'; Dr Michael Greenberg (Harvard University) spoke about 'Signal transduction pathways that regulate differentiation and survival in the developing nervous system'; Dr Richard Kolesnick's (Memorial Sloan-Kettering Cancer Center) talk has been focused at 'Stress signals for apoptosis, including Ceramide and c-Jun Kinase/Stress-activated Protein Kinase'; Dr Barbara Hoffman (Fels Institute, Temple University) described research, conducted in collaboration with Dr Dan A Liebermann, aimed at deciphering the roles of 'myc, myb, and E2F as negative regulators of terminal differentiation', using hematopoietic cells as model system. Dr Daniel G Tenen (Harvard Medical School), described studies aimed at understanding the 'Regulation of hematopoietic cell development by lineage specific transcription regulators'. Dr George C Prendergast (The Wistar Institute) talked about the 'Myc-Bin1 signaling pathway in cell death and differentiation. Dr Ruth J Muschel (University of Pennsylvania) spoke about work, conducted in collaboration with Dr WG McKenna, aimed at gaining a better understanding of 'Radioresistance and the cell cycle'. Finally Dr Donald Kufe concluded the symposium (Dana Farber Cancer Institute, Harvard Medical School) describing studies that were performed in his laboratory addressing the 'Role for the c-Abl tyrosine kinase in genetic recombination'. Publication Types: Congresses PMID: 9771961 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 332: J Toxicol Environ Health A. 1998 Sep 25;55(2):121-31. Modulation of proto-oncogene expression by polychlorinated biphenyls in 3T3-L1 cell line. Gribaldo L, Sacco MG, Casati S, Zucchi I, Dosanjh MK, Catalani P, Marafante E. European Centre for the Validation of Alternative Methods-J.R.C., Ispra, Italy. The effects of two substituted polychlorinated biphenyls, the 3,4,5,3',4,5' (PCB-169) and the 2,3,4,2',4',5' (PCB-138) forms, were examined on the expression of c-myc, c-jun, c-ras, and jun-b in 3T3-L1 cells. Northern blot analysis demonstrated that the two PCBs, which exhibit a coplanar and di-ortho-substituted configuration, activated these oncogenes differently. PCB-138 markedly induced overexpression of ras, jun, and myc, whereas PCB-169 led to the overexpression of jun-b. High-performance liquid chromatography analysis of the cell samples treated in medium without serum revealed a higher intracellular concentration of the 2,3,4,2',4',5'-hexachlorobiphenyl (hexaCB), whereas the 3,4,5,3',4'5'-hexaCB reached the same concentration in the sonicated samples of cells with or without serum. These results indicated that there was a relationship between PCB structure, bioavailability, and the capacity to stimulate oncogene expression. PMID: 9761132 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 333: Hepatology. 1998 Oct;28(4):959-70. Comment on: Hepatology. 1998 Oct;28(4):906-13. Analysis of liver regeneration in mice lacking type 1 or type 2 tumor necrosis factor receptor: requirement for type 1 but not type 2 receptor. Yamada Y, Webber EM, Kirillova I, Peschon JJ, Fausto N. Department of Pathology, University of Washington School of Medicine, Seattle, WA 98195-7705, USA. We used KO mice lacking either TNF receptor 1 (TNFR-1) or receptor 2 (TNFR-2) to determine whether signaling at the start of liver regeneration after partial hepatectomy (PH) involves only one or both TNF receptors and to analyze in more detail the abnormalities caused by lack of TNFR-1 receptor, which is required for the initiation of liver regeneration. Lack of TNFR-2 had little effect on NF-kappaB and STAT3 binding, and no effect in interleukin-6 production after PH, but caused a delay in AP-1 and C/EBP binding and in the expression of c-jun and c-myc messenger RNA (mRNA). In contrast to mice lacking TNFR-1, which had deficient hepatocyte DNA synthesis and massive lipid accumulation in hepatocytes, TNFR-2 KO mice had normal liver structure and similar levels of hepatocyte DNA replication as those of wild type mice. We conclude that TNFR-1, but not TNFR-2, is necessary for liver regeneration, and that NF-kappaB and STAT3 binding are activated by signals transduced by TNFR-1. Inhibition of AP-1 and C/EBP binding and in the expression of c-jun and c-myc mRNA in the first 4 hours after PH, as well as the apparent lack of Fos in AP-1 complexes, had no effect on the timing or extent of DNA replication. Publication Types: Comment PMID: 9755232 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 334: Blood. 1998 Sep 15;92(6):1957-66. The zinc finger transcription factor Egr-1 activates macrophage differentiation in M1 myeloblastic leukemia cells. Krishnaraju K, Hoffman B, Liebermann DA. Fels Institute for Cancer Research and Molecular Biology, and Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, USA. We previously have shown that the zinc finger transcription factor Egr-1 blocked granulocytic differentiation of HL-60 cells, restricting differentiation along the monocytic lineage. Egr-1 also was observed to block granulocyte colony-stimulating factor (G-CSF)-induced differentiation of interleukin-3 (IL-3)-dependent 32Dcl3 hematopoietic precursor cells, endowing the cells with the ability to be induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) for terminal differentiation along the macrophage lineage. To better understand the function of Egr-1 as a positive modulator of monocytic differentiation, in this work we have studied the effect of ectopic expression of Egr-1 on the murine myeloblastic leukemic cell line M1, which is induced for differentiation by the physiological inducer IL-6. It is shown that, unlike in HL-60 and 32Dcl3 cells, ectopic expression of Egr-1 in M1 cells resulted in activation of the macrophage differentiation program in the absence of differentiation inducer. This included the appearance of morphologically differentiated cells, decreased growth rate in mass culture, and cloning efficiency in soft agar, and expression of endogenous c-myb and c-myc mRNAs was markedly downregulated. Untreated M1Egr-1 cells also exhibited cell adherence, expression of Fc and C3 receptors, and upregulation of the myeloid differentiation primary response genes c-Jun, junD, and junB and the late genetic markers ferritin light-chain and lysozyme. Ectopic expression of Egr-1 in M1 cells also dramatically increased the sensitivity of the cells for IL-6-induced differentiation, allowed a higher proportion of M1 cells to become terminally differentiated under conditions of optimal stimulation for differentiation, and decreased M1 leukemogenicity in vivo. These findings demonstrate that the functions of Egr-1 as a positive modulator of macrophage differentiation vary, depending on the state of lineage commitment for differentiation of the hematopoietic cell type. Copyright 1998 by The American Society of Hematology. PMID: 9731053 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 335: Br J Urol. 1998 Aug;82(2):261-6. Tissue damage and regeneration of ureteric smooth muscle in rats with obstructive uropathy. Chuang YH, Chuang WL, Liu KM, Chen SS, Huang CH. Department of Neurology, School of Medicine, Kaohsiung Medical College, Taiwan, ROC. OBJECTIVE: To investigate regeneration in obstructed ureters and to elucidate the role of hyperplasia in the thickening of the smooth muscle layer in the late stages of complete ureteric obstruction. MATERIALS AND METHODS: The expression of Ki-67 antigen, c-Fos, c-Jun and c-Myc in the smooth muscle layer of obstructed ureters was determined using immunohistochemistry in 40 Sprague-Dawley rats. After unilateral ligation of the ureter, five rats each were killed for examination at 1, 3, 7, 10, 14, 21, 28 and 42 days after ligation: dive rats that underwent a sham operation were also examined as controls. RESULTS: The severity of hydroureter and thickening of the smooth muscle layer progressed consistently in the ligated ureters, but no mitosis was detected in myocytes within 14 days of ligation. Fibrosis in the smooth muscle layer appeared 21 days after ligation and progressed. There was no expression of Ki-67 antigen and oncoproteins until 14 days after ligation. The expression of Ki-67 and c-Myc increased gradually to a peak after 28 days, then declined. However, the expression of c-Fos and c-Jun was low and transient. CONCLUSION: Cell regeneration is impaired in the damaged muscle layer of obstructed ureters. Only hypertrophy and not hyperplasia of the smooth muscle layer developed during the course of complete ureteric obstruction in this rat model of obstructive uropathy. PMID: 9722764 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 336: J Biol Chem. 1998 Sep 4;273(36):23046-54. Oxidative stress activates the human histidine decarboxylase promoter in AGS gastric cancer cells. Hocker M, Rosenberg I, Xavier R, Henihan RJ, Wiedenmann B, Rosewicz S, Podolsky DK, Wang TC. Medizinische Klinik mit Schwerpunkt Hepatologie und Gastroenterologie, Universitatsklinikum Charite, Campus Virchow Klinikum, Humboldt Universitat, Berlin, Germany. Oxidant stress is thought to play a role in the pathogenesis of many gastric disorders. We have recently reported that histidine decarboxylase (HDC) promoter activity is stimulated by gastrin through a protein kinase C- and extracellular signal-regulating kinase (ERK)-dependent pathway in gastric cancer (AGS-B) cells, and this transcriptional response is mediated by a downstream cis-acting element, the gastrin response element (GAS-RE). To study the mechanism through which oxidant stress affects gastric cells, we examined the effects of hydrogen peroxide (H2O2) on HDC promoter activity and intracellular signaling in AGS-B cells. H2O2 (10 mM) specifically activated the HDC promoter 10-12-fold, and this activation was blocked by both mannitol and N-acetylcysteine. Hydrogen peroxide treatment of AGS-B cells increased the phosphorylation and kinase activity of ERK-1 and ERK-2, but did not affect Jun kinase tyrosine phosphorylation or kinase activity. In addition, treatment of AGS-B cells with H2O2 resulted in increased c-fos/c-jun mRNA expression and AP-1 activity, and also led to increased phosphorylation of epidermal growth factor receptor (EGFR) and Shc. H2O2-dependent stimulation of HDC promoter activity was completely inhibited by kinase-deficient ERKs, dominant-negative (N17 and N15) Ras, and dominant-negative Raf, and partially blocked by a dominant-negative EGFR mutant. In contrast, protein kinase C blockade did not inhibit H2O2-dependent induction of the HDC promoter. Finally, deletion analysis demonstrated that the H2O2 response element could be mapped to the GAS-RE (nucleotides 2 to 24) of the basal HDC promoter. Overall, these studies suggest that oxidant stress activates the HDC promoter through the GAS-RE, and through an Ras-, Raf-, and ERK-dependent pathway at least partially involving the EGFR. PMID: 9722530 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 337: Br J Pharmacol. 1998 Jul;124(6):1227-37. Epigallocatechin suppression of proliferation of vascular smooth muscle cells: correlation with c-jun and JNK. Lu LH, Lee SS, Huang HC. Department of Pharmacology, College of Medicine, National Taiwan University, Taipei. 1. The mechanisms of the antiproliferative effect of epigallocatechin, one of the catechin derivatives found in green tea, in vascular smooth muscle cells were studied. The proliferative response was determined from the uptake of tritiated thymidine. 2. In the concentration range of 10(-6) to 10(-4) M, catechin, epicatechin, epigallocatechin, epicatechin gallate and epigallocatechin, epigallocatechin gallate, concentration-dependently inhibited the proliferative response stimulated by serum in rabbit cultured vascular smooth muscle cells. Catechin and epicatechin were less effective in inhibiting the serum-stimulated smooth muscle cell proliferation, indicating that the galloyl group may be important for full inhibitory activity. 3. Epigallocatechin (EGC) inhibited the proliferative responses in different cells including rat aortic smooth muscle cells (A7r5 cells), rabbit cultured aortic smooth muscle cells, human coronary artery smooth muscle cells, and human CEM lymphocytes in a concentration-dependent manner. The possible mechanisms of the antiproliferative effect of EGC were further studied in A7r5 cells. 4. The membranous protein tyrosine kinase activity stimulated by serum in A7r5 cells was significantly reduced by 10(-5) M EGC. In contrast, the cytosolic protein kinase C activity stimulated by phorbol ester was unaffected by directly incubating with EGC (10(-6)-10(-4) M). 5. We also performed Western blot analysis using the anti-phosphotyrosine monoclonal antibody PY20. EGC (10(-5) M) reduced the levels of tyrosine phosphorylated proteins with different molecular weights, indicating that EGC may inhibit the protein tyrosine kinase activity or stimulate the protein phosphatase activity. 6. Reverse transcription-polymerase chain reaction analysis of c-fos, c-jun and c-myc mRNA levels demonstrated that c-jun mRNA level after serum-stimulation was significantly reduced by 10(-5) M EGC. However, the reduction of c-fos and c-myc mRNA levels by 10(-5) M EGC did not achieve significance. 7. Western blot analysis using the antibody against JNK (c-jun N-terminal kinase) and ERK (extracellular signal-regulated kinase) demonstrated that the level of phosphorylated JNK1, but not phosphorylated ERK1 and ERK2, was reduced by 10(-5) M EGC. Direct measurement of kinase activity by immune complex kinase assay confirmed that JNK1 activity was inhibited by EGC treatment. These results demonstrate that EGC preferentially reduced the activation of JNK/SAPK (stress-activated protein kinase) signal transduction pathway. 8. It is suggested that the antiproliferative effect of epigallocatechin on vascular smooth muscle cells may partly be mediated through inhibition of protein tyrosine kinase activity, reducing c-jun mRNA expression and inhibiting JNK1 activation. Tea catechins may be useful as a template for the development of drugs to prevent the pathological changes of atherosclerosis and post-angioplasty restenosis. PMID: 9720795 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 338: Biochem Pharmacol. 1998 Jun 15;55(12):1963-71. Doxorubicin-induced alterations of c-myc and c-jun gene expression in rat glioblastoma cells: role of c-jun in drug resistance and cell death. Pourquier P, Montaudon D, Huet S, Larrue A, Clary A, Robert J. Universite Victor Segalen Bordeaux 2 and Institut Bergonie, France. We studied the effect of doxorubicin on the expression of c-myc and c-jun in the rat glioblastoma cell line C6 and its doxorubicin-resistant variant C6 0.5, at equitoxic exposures. For quantitation, the mRNA levels of these oncogenes were related to those of two domestic genes, beta-actin and glyceraldehyde phosphate dehydrogenase. After a transient overexpression of the genes during the first hour of incubation, there was a selective, dose-dependent down-regulation of both genes by doxorubicin in the sensitive cells. In the resistant cell line, c-myc expression was also decreased in response to doxorubicin incubation, but the expression of c-jun remained unchanged over the whole range of concentrations. In contrast, vincristine had no effect on the amounts of c-myc and c-jun mRNAs in either line. The effect of doxorubicin on the mRNA levels of c-jun was also observed on the JUN proteins by immunoblotting, but the MYC protein levels remained unchanged upon doxorubicin treatment. There was a significant correlation between the levels of c-myc and c-jun gene expression and the degree of growth inhibition induced by doxorubicin. In addition, doxorubicin induced a fragmentation of DNA in sensitive cells, but not in resistant cells, thus revealing a resistance to apoptosis in this line. Doxorubicin-induced cell death did not appear to be mediated by p53 in either cell line. PMID: 9714316 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 339: J Cell Sci. 1998 Sep;111 ( Pt 17):2519-27. Evidence for MAP kinase activation during mitotic division. Chiri S, De Nadai C, Ciapa B. Groupe de Recherche Sur l'Interaction Gametique (GRIG), CJF 9504 INSERM, Faculte de Medecine, Avenue de Valombrose, Cedex 02 France. MAP kinases have been implicated in the control of a broad spectrum of cellular events in many types of cells. In somatic cells, MAP kinase activation seems to be triggered after exit from a quiescent state (in G0 or G2) only and then inactivated by entry into a proliferative state. In oocytes of various species, a one-time activation of MAP kinase that is apparently not repeated during the succeeding mitotic cycles occurs after meiotic activation. However, several reports suggest that a myelin basic protein (MBP) kinase activity, unrelated to that of maturation promoting factor, can sometimes be detected during mitotic divisions in various types of cells and oocytes. We have reinvestigated this problem in order to determine the origin and the role of MBP kinase that is stimulated at time of mitosis in the fertilized eggs of the sea urchin Paracentrotus lividus. We used anti-ERK1 antibodies or substrates specific for different MAP kinases, and performed in-gel phosphorylation experiments. Our results suggest that an ERK1-like protein was responsible for part of the MBP kinase activity that is stimulated during the first mitotic divisions. Furthermore, we observed that wortmannin, an inhibitor of PI 3-kinase that arrests the fertilized sea urchin eggs at the prometaphase stage, inhibited the inactivation of MAP kinase normally observed when the eggs divide, suggesting a role for PI 3-kinase in the deactivation process of MAP kinase. We also discuss how the activities of MPF and MAP kinase may be interconnected to regulate the first mitotic divisions of the early sea urchin embryo. PMID: 9701551 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 340: Biochem Pharmacol. 1998 Jul 15;56(2):163-71. Tissue remodelling in the adrenal gland. Wolkersdorfer GW, Bornstein SR. Department of Internal Medicine III, University of Leipzig, Germany. wolkg@server3.medizin.uni-leipzig.de Adaptation of the adrenal gland to the demands of the organism is regulated functionally and structurally. Three common hypotheses on zonation in the adrenal gland, the migrational, zonal, and transformation field theories, try independently to reconcile the findings on structure, proliferation, and cell death. The classical theories on zonation are revisited in the light of recent data on cell death and renewal. In accordance with data on cell death as immunoreactivity against FAS(CD 95), an apoptosis-inducing receptor, in situ end labelling of fragmented DNA, and ultrastructural analyses, programmed cell death (PCD) occurs throughout the whole organ. The angiotensin II receptor subtypes described in the adrenal allow an additional regulation of tissue homeostasis by proliferative and even by the antiproliferative effects of the angiotensin II type 2 receptor. Proto-oncogenes are involved in the regulation of cell cycle and PCD, and adrenocorticotropin asserts its tissue integrating and differentiating effects by regulating proto-oncogenes such as c-jun, c-fos, jun-B and c-myc. Polypeptides involved in proliferation and DNA repair, such as proliferating cell nuclear antigen and Ki-67, have been found within zones of expected cell senescence. The expression of the class II major histocompatibility complex on normal adrenocortical cells allows cell-to-cell communication with the immune system and may trigger the Fas/Fas-ligand system to permit tissue regression and decreasing activity in both systems. In summary, new data allow us to reappraise and to reconcile the classical theories. Apoptosis is a physiological process in the adrenal gland. There is a differential regulation of apoptosis in the different zones. An investigation of this process may elucidate the basic mechanisms of adrenal zonation. Publication Types: Review Review, Tutorial PMID: 9698069 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 341: J Biol Chem. 1998 Aug 14;273(33):21423-9. Myb-dependent regulation of thrombospondin 2 expression. Role of mRNA stability. Bein K, Ware JA, Simons M. Angiogenesis Research Center, Cardiovascular Division, Department of Medicine Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts 02215, USA. The nuclear transcription factor c-Myb, which is highly expressed in hematopoietic cells, has been shown to be functional in NIH 3T3 cells: cells that do not possess detectable levels of c-Myb. To identify endogenous target genes of c-Myb in fibroblasts, RNA isolated from NIH 3T3 cells stably transfected with a full-length or a dominant negative c-myb construct (GREMyb and GREMEn, respectively) was subjected to differential display analysis. 5'-Rapid amplification of cDNA ends of a selected band, sequencing, and a nucleotide homology search led to the identification of thrombospondin 2 (TSP 2) as the gene product repressed in GREMyb and induced in GREMEn cells. The pattern of TSP 2 expression during the cell cycle was consistent with c-myb-dependent regulation. The possibility that the identified transcript was TSP 1, a homologous product known to be repressed by v-Src, c-Jun, and v-Myc, was ruled out by using a TSP 2-specific DNA probe and by showing a distinct pattern of regulation of TSP 1 and TSP 2 expression. Nuclear run-on and TSP 2 promoter-reporter (chloramphenicol acetyltransferase) assays showed similar transcriptional levels in GREMyb and NIH 3T3 cells. However, mRNA stability studies showed a much shorter TSP 2 mRNA half-life in GREMyb compared with wild type NIH 3T3 cells, suggesting that c-myb affects TSP 2 expression via a post-transcriptional mechanism. The implications of a protooncogene-mediated suppression of TSP expression are discussed. PMID: 9694906 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 342: J Cancer Res Clin Oncol. 1998;124(6):307-14. Molecular pathology of hemangiopericytomas accompanied by severe hypoglycemia: oncogenes, tumor-suppressor genes and the insulin-like growth factor family. Pavelic K, Cabrijan T, Hrascan R, Vrkljan M, Lipovac M, Kapitanovic S, Gall-Troselj K, Bosnar MH, Tomac A, Grskovic B, Karapandza N, Pavelic LJ, Kurslin B, Spaventi S, Pavelic J. Division of Molecular Medicine Ruder Boskovic Institute, Zagreb, Croatia. pavelic@rudjer.irb.hr Relatively little is known about molecular genetic events that participate in the genesis and progression of hemangiopericytoma. In this study, we describe two cases of hemangiopericytoma accompanied by severe hypoglycemia. Tumor cells from patient 1 exhibited insulin-growth factor I (IGF I) and insulin-like growth factor I receptor (IGF IR) mRNA transcripts. Tumor cells from patient 2 exhibited IGF II, IGF IR and IGF binding proteins 1-3 mRNA. Serum from patient 2 contained IGF II, mostly in a large molecular form ("big" IGF II); the IGF II level did not change after the tumor removal. The presence of IGF IR in tumor cells was confirmed by immunoprecipitation with antibodies that recognize human IGF IR subunit (visualized as a 460-kDa band). The hemangiopericytoma cells derived from patient 1 expressed 210000 IGF I receptors/cell. Specific binding of IGF I to the tumor cell membrane fraction was higher in tissue from patient 1, while the tissue of patient 2 showed relatively low IGF I binding. In contrast, IGF II binding was much higher in tissue from patient 2. Both tumor tissues showed positive immunostaining for c-Jun; one tumor showed strong immunostaining for c-Myc, H-Ras and p53, while the other exhibited strong reaction with H-Ras antibodies only. No loss of the heterozygosity at the genes APC, NFI and nm23-H1 loci in tumor tissue obtained from patient 1 was found. In effect, our results suggest multiple molecular genetic changes in hemangiopericytoma -- activation of some oncogenes and the IGF growth factor family. IGF ligands together with IGF IR could be responsible for hypoglycemia and perhaps the transformed phenotype. Publication Types: Case Reports PMID: 9692837 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 343: Dig Dis Sci. 1998 Jul;43(7):1526-36. Systemic short-chain fatty acids rapidly alter gastrointestinal structure, function, and expression of early response genes. Tappenden KA, McBurney MI. Department of Food Science and Human Nutrition, University of Illinois, Urbana 21801, USA. Luminal and systemic short chain fatty acids (SCFA) stimulate mucosal proliferation but the mechanism(s) is unclear. This study examined acute effects of systemic SCFAs on gastrointestinal structure and function and signals potentially mediating SCFA-induced mucosal proliferation. Male Sprague-Dawley rats (246+/-2 g) received nutrients as either standard total parenteral nutrition (TPN) or an isoenergetic, isonitrogenous formulation containing SCFAs (TPN + SCFA). Animals were randomized to one of five treatments: standard TPN for 72 hr, TPN + SCFA for 72 hr, or standard TPN followed by TPN + SCFA for the final 6, 12, and 24 hr. SCFAs reduced (P < 0.003) ileal protein within 6 hr. Jejunal GLUT2 expression was increased (P=0.0001) in all SCFA groups and ileal GLUT2 protein in the 6-, 12-, and 24-hr SCFA groups (P < 0.05). SCFAs increased (P < 0.003) ileal proglucagon abundance following 6, 12, and 24 hr, and plasma GLP-2 concentration following 12 hr (P < 0.03). Jejunal c-myc expression was increased (P < 0.001) following 6, 12, and 24 hr of SCFAs. SCFAs increased ileal c-myc, c-jun, and c-fos expression following 24 hr (P < 0.02), 12 hr (P < 0.05) and 6, 12, and 24 hr (P=0.0001), respectively. In conclusion, systemic SCFAs increase plasma GLP-2 and ileal proglucagon mRNA, GLUT2 expression and protein, and c-myc, c-jun, and c-fos expression. PMID: 9690391 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 344: Am J Physiol. 1998 Jul;275(1 Pt 1):L21-9. Increased lung inflation induces gene expression after pneumonectomy. Gilbert KA, Rannels DE. Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA. Rapid hyperplastic growth of the remaining lung is initiated by partial pneumonectomy in many mammalian species. The response restores normal tissue structure and function. Although physiological control of compensatory lung growth is documented, little is known about the molecular mechanisms that underlie the process. The aim of this study was to investigate the role of mechanical signals in the induction of immediate-early gene (IEG) expression after pneumonectomy. Expression of c-fos and junB increased nine- and fourfold, respectively, in the right lung within 30 min after left pneumonectomy in rats. In contrast, changes in expression of c-jun and c-myc were not observed. When isolated lungs were subjected to elevated airway pressures in vitro, expression of c-fos and junB was induced in a time- and dose-dependent manner similar to that observed in vivo. Similarly, in vitro lung perfusion induced c-fos and junB expression in the absence of increasing lung inflation. These results support the premise that rapid changes in IEG expression after pneumonectomy are initiated by mechanical signaling in the remaining lung. Elevated IEG expression may contribute to initiation of compensatory lung growth. PMID: 9688931 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 345: Antimicrob Agents Chemother. 1998 Aug;42(8):1923-30. Ciprofloxacin induces an immunomodulatory stress response in human T lymphocytes. Riesbeck K, Forsgren A, Henriksson A, Bredberg A. Department of Medical Microbiology, Lund University, Malmo University Hospital, S-205 02 Malmo, Sweden. riesbeck@mikrobiol.mas.lu.se Exposure of cells to adverse environmental conditions invokes a genetically programmed series of events resulting in the induction of specific genes. The fluoroquinolone antibiotic ciprofloxacin has recently been reported to upregulate interleukin-2 (IL-2) gene induction. In the present investigation, the effect of ciprofloxacin at supratherapeutic concentrations on immediate-early (<2 h) gene expression in primary human peripheral blood lymphocytes was studied with Northern blots. In addition, transcriptional activity of IL-2 and metallothionein enhancer and promoter regions and transcription factors AP-1, NF-kappaB, and NF-AT were analyzed by chloramphenicol acetyltransferase (CAT) and electrophoretic mobility shift assays, respectively. The concentration of c-fos, c-jun, c-myc, junB, and fra-1 mRNAs was increased in activated peripheral blood lymphocytes incubated with ciprofloxacin compared to that in untreated controls. Ciprofloxacin increased CAT activity in stimulated lymphocytes transfected with plasmids containing either the IL-2 or metallothionein enhancer. Furthermore, among the transcription factors tested, AP-1 activity was increased in stimulated purified T helper lymphocytes incubated with ciprofloxacin compared to drug-free controls. Taken together, ciprofloxacin increased the levels of immediate-early transcripts, enhanced IL-2 and metallothionein promoter induction, and upregulated AP-1 concentrations in primary lymphocytes, reflecting a program commonly observed in mammalian stress responses. PMID: 9687385 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 346: Exp Cell Res. 1998 Aug 1;242(2):401-9. Hepatocyte growth factor-induced expression of ornithine decarboxylase, c-met, and c-myc is differently affected by protein kinase inhibitors in human hepatoma cells HepG2. Desiderio MA, Pogliaghi G, Dansi P. Institute of General Pathology and CNR Center for Research on Cell Pathology, University of Milano, via Mangiagalli, Milan, 31-20133, Italy. desi@imiucca.csi.unimi.it Binding of hepatocyte growth factor (HGF) to its receptor Met induces autophosphorylation and activation of the tyrosine kinase activity. In HGF-treated HepG2 cells, we studied: (i) the expression patterns of early (c-myc, c-jun, and c-fos) and delayed-early (ornithine decarboxylase and c-met) response genes and (ii) the possible involvement of protein kinase transducers in the control of the expression of c-met and of other genes eventually induced downstream. c-met and c-myc mRNAs peaked 1-2 h after HGF, while c-jun and c-fos mRNAs slightly increased at 1 h. Ornithine decarboxylase activity was induced earlier (4 h) than the mRNA (8-10 h). The transducers involved in HGF-triggered gene inductions were investigated using different protein kinase inhibitors: genistein for the receptor tyrosine kinase, herbimycin A for the nonreceptor tyrosine kinase (pp60(c-src)), wortmannin for phosphatidylinositol 3-kinase (PI3K) and H7 for protein kinase C (PKC). The similarity of responses to PKC inhibition led to suppose that c-myc and ornithine decarboxylase mRNAs were induced sequentially along the same transduction pathway triggered by HGF. Ornithine decarboxylase activity seemed to be largely regulated by phosphorylation(s). The mRNA expression of c-jun was likely to undergo a negative regulation through a mechanism involving PI3K, while that of c-met seemed to be almost independent from various protein kinases (PI3K, pp60(c-src), and PKC). Copyright 1998 Academic Press. PMID: 9683527 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 347: Oncogene. 1998 Jun 11;16(23):3057-68. Transition from SCLC to NSCLC phenotype is accompanied by an increased TRE-binding activity and recruitment of specific AP-1 proteins. Risse-Hackl G, Adamkiewicz J, Wimmel A, Schuermann M. Zentrum fur Innere Medizin, Abteilung Hamatologie/Onkologie, Philipps-Universitat Marburg, Germany. Transitions from small cell (SCLC) to non-small cell lung cancer (NSCLC) cells have been documented both in vitro and in vivo and are thought to be an important step during tumor progression of human small cell lung cancer towards a treatment-resistant tumor state. We have screened NSCLC and SCLC cell lines for differences in the composition of nuclear transcription factors using consensus oligonucleotide sequences (SRE, Ets, TRE, CRE, B-motif, GAS, E-box). We found NSCLC cells to exhibit significantly higher AP-1 binding activity than SCLC cells consistent with the increased expression of CD44, an AP-1 target gene. To gain more insight into the molecular mechanisms underlying these differences, we analysed SCLC cell lines (NCI-N592 and NCI-H69) which were phenotypically transformed into NSCLC-type cells by transfection with activated H-ras and c-myc oncogenes. In these cells, ras-induced transition is accompanied by a strong induction of AP-1-binding activity along with increased expression of CD44 mRNA and protein. When analysing the composition of the AP-1 complex in more detail and comparing ras-induced versus phorbol ester-induced changes, we found Fra-1 to be the major component induced in ras-transfected but not in phorbol-ester treated or non-treated parental SCLC cells. This finding is paralleled by the observation that among the various members of the Fos and Jun family analysed (c-Fos, FosB, Fra-1, Fra-2, c-Jun, JunD, JunB) fra-1 is the only gene to be exclusively expressed in NSCLC cells but not in cells of SCLC origin. Our data, thus, point to a histiotype-related mechanism of recruitment among AP-1 proteins which may have bearings on the fate of lung cancer development. PMID: 9662339 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 348: Prostaglandins Leukot Essent Fatty Acids. 1998 Apr;58(4):271-81. Inhibition of EGF-dependent mitogenesis by prostaglandin E2 in Syrian hamster embryo fibroblasts. Hsi LC, Eling TE. National Institute of Environmental Health Sciences, Laboratory of Molecular Carcinogenesis, Eicosanoid Biochemistry Section, Research Triangle Park, NC 27709, USA. Lipid metabolism can play an important role in the development and progression of human cancers. We have used Syrian hamster embryo (SHE) fibroblasts as a model system to study how lipid metabolites can alter cell proliferation and apoptosis. For example, the linoleic acid metabolite 13(S)-HpODE enhances EGF-dependent growth by inhibiting de-phosphorylation of the EGFR which leads to activation of the MAP kinase pathway. In contrast, the arachidonic acid metabolite, PGE2, inhibits EGF-dependent mitogenesis and the expression of the proto-oncogenes c-myc, c-jun, and jun-B. In this study, we have investigated the mechanism by which PGE2 attenuates these responses by studying the EGF signaling cascade in SHE cells. PGE2 pretreatment caused a concentration-dependent decrease in EGF-dependent phosphorylation of MAP kinase and a corresponding inhibition of EGF-stimulated MAP kinase activity. Pretreatment of the SHE cells with PGE2 had little effect on the magnitude of EGF-dependent receptor auto-phosphorylation and the phosphorylation of GAP suggesting a down-stream target. Treatment of cells with forskolin and EGF causes similar inhibition of MAP kinase phosphorylation as observed with PGE2 and EGF. Since PGE2 elevates cAMP in these cells, it may act by altering cAMP accumulation. Raf-1 activity can be inhibited by a cAMP-dependent process. Raf-1 activity, measured by phosphorylation of Mek-1, was attenuated by the addition of PGE2. To determine if inhibition of Raf-1 activity causes inhibition of the MAP kinase pathway, cells were concomitantly incubated with PGE2 and EGF. Inhibition of MAP kinase phosphorylation was observed. From these data, we propose that in SHE cells PGE2 increases cAMP levels, which in turn causes inhibition of Raf-1 activity. The MAP kinase pathway is thus downregulated which decreases mitogenesis and proto-oncogene expression. This study demonstrates that an arachidonic acid metabolite can modulate phosphorylation and activity of key signal transduction proteins in a growth factor mitogenic pathway. PMID: 9654400 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 349: Biochem Biophys Res Commun. 1998 Jun 29;247(3):773-9. Different modes of cell death induced by 5-fluoro-2'-deoxyuridine in two clones of the mouse mammary tumor FM3A cell line. Kakutani T, Ebara Y, Kanja K, Hidaka M, Matsumoto Y, Nagano A, Wataya Y. Faculty of Pharmaceutical Sciences, Okayama University, Japan. The mode of cell death induced by 1 microM 5-fluoro-2'-deoxyuridine (FdUrd) changed in a wild-type F28-7 clone of mouse mammary tumor FM3A cells after a six-month culture. In the original stocked F28-7 clone, FdUrd-induced cell death was accompanied by necrosis-like cell swelling and DNA fragmentation to 100-200 kbp. In subclone F28-7-A isolated from F28-7 cells, which had been cultured for six months, apoptotic bodies and nucleosomal DNA-ladder fragments were observed with the treatment. Furthermore, we investigated the differences in FdUrd-induced intracellular signals between these clones. In F28-7 cells, FdUrd induced increases in caspase-3-like activity, and the mRNA levels of the c-jun, c-fos and c-myc genes, which were greater and earlier than those in F28-7-A cells. Moreover, intracellular acidification occurred in F28-7-A cells treated with FdUrd, though it was not observed in F28-7 cells. These findings suggest that FdUrd-induced cell death occurred through the death program to cell lysis (necrosis) without apoptosis when the induction of these intracellular signals was very high and when intracellular acidification was deficient. Investigation of the differences in the mode of FdUrd-induced cell death between these clones would be important for elucidating the molecular mechanism of pivotal events guiding cells toward either apoptosis or necrosis. PMID: 9647769 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 350: J Immunol. 1998 Jun 15;160(12):5898-906. Erratum in: J Immunol 1999 Jul 15;163(2):1093. CpG oligodeoxyribonucleotides rescue mature spleen B cells from spontaneous apoptosis and promote cell cycle entry. Yi AK, Chang M, Peckham DW, Krieg AM, Ashman RF. Medical Services, Department of Veterans Affairs, Iowa City, IA 52246, USA. Isolated murine splenic B cells undergo spontaneous apoptosis. Motifs containing unmethylated CpG dinucleotides in bacterial DNA or in synthetic oligodeoxynucleotides (ODN) are known to activate murine B cells. Now we show that ODN that induce spleen B cell cycle entry also inhibit spontaneous apoptosis in a sequence-specific fashion. Reversal of the CG to GC abolished activity. Methylation of the central cytosine decreased activity. When CpG is preceded by a cytosine or followed by a guanine, activity was abolished. Other substitutions at the same positions had no effect. Dose-response curves for apoptosis protection and G1 entry suggested that a uniform population of ODN recognition sites controlled downstream ODN effects. A CpG ODN with a nuclease-resistant phosphorothioate backbone (S-ODN) was also active, and increased the levels of c-myc, egr-1, c-jun, bclXL, and bax mRNA and c-Myc, c-Jun, Bax, and BclXL protein in spleen B cells. Levels of c-myb, myn, c-Ki-ras, and bcl2 mRNA remained unchanged. When protein synthesis was inhibited, at 16 h ODN-induced cell cycle entry was abolished and apoptosis protection was partially preserved. Under these conditions, c-Myc was still present, but c-Jun and BclXL were not detected. Our results suggest that CpG containing ODN motifs provide signals for both survival and cell cycle entry. Single base changes determine whether this signal proceeds through a rate-limiting step governing at least two steps in apoptosis (plasma membrane transition, DNA cleavage) and two phases of the cell cycle (G1 and S phase entry). This biologic action is associated with increased c-Myc, c-Jun, and BclXL expression. PMID: 9637502 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 351: Crit Rev Immunol. 1998;18(3):185-220. IL-2-induced cellular events. Gomez J, Gonzalez A, Martinez-A C, Rebollo A. Department of Immunology and Oncology, Centro Nacional de Biotecnologia, Universidad Autonoma, Campus de Cantoblanco, Madrid, Spain. In this review we discuss several molecules that are attractive candidates as transducing molecules involved in signaling processes. IL-2 receptor signaling is a complex process involving a large number of molecules: Ras, Rho, PI3 kinase, PKC, Akt, transcription factors NF-AT, and NF-kappaB and some target genes such as bcl-2, c-myc, c-jun and c-fos. Ras and Rho have been defined as dual molecules because Ras- and Rho-initiated signals can either promote or inhibit apoptosis. Several studies have contributed to the delineation of a signaling pathway structured in three independent channels designated channels 1, 2, and 3. These three channels serve as major landmarks: Lck-c-fos/c-jun (channel 1), Syk-myc (channel 2), and a pathway leading to actin organization/bcl-2 expression (channel 3). The detailed hierarchical organization of these three channels is presented throughout the review and the model is depicted in the figure. Publication Types: Review PMID: 9637410 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 352: Mol Cell Biol. 1998 Jun;18(6):3620-32. Human T-cell leukemia virus type 1 Tax and cell cycle progression: role of cyclin D-cdk and p110Rb. Neuveut C, Low KG, Maldarelli F, Schmitt I, Majone F, Grassmann R, Jeang KT. Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892-0460, USA. Human T-cell leukemia virus type 1 is etiologically linked to the development of adult T-cell leukemia and various human neuropathies. The Tax protein of human T-cell leukemia virus type I has been implicated in cellular transformation. Like other oncoproteins, such as Myc, Jun, and Fos, Tax is a transcriptional activator. How it mechanistically dysregulates the cell cycle is unclear. Previously, it was suggested that Tax affects cell-phase transition by forming a direct protein-protein complex with p16(INK4a), thereby inactivating an inhibitor of G1-to-S-phase progression. Here we show that, in T cells deleted for p16(INK4a), Tax can compel an egress of cells from G0/G1 into S despite the absence of serum. We also show that in undifferentiated myocytes, expression of Tax represses cellular differentiation. In both settings, Tax expression was found to increase cyclin D-cdk activity and to enhance pRb phosphorylation. In T cells, a Tax-associated increase in steady-state E2F2 protein was also documented. In searching for a molecular explanation for these observations, we found that Tax forms a protein-protein complex with cyclin D3, whereas a point-mutated and transcriptionally inert Tax mutant failed to form such a complex. Interestingly, expression of wild-type Tax protein in cells was also correlated with the induction of a novel hyperphosphorylated cyclin D3 protein. Taken together, these findings suggest that Tax might directly influence cyclin D-cdk activity and function, perhaps by a route independent of cdk inhibitors such as p16(INK4a). PMID: 9584203 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 353: Ann N Y Acad Sci. 1998 May 15;839:561-3. Detection of c-Myc, c-Fos, and c-Jun-like products in the lizard (Podarcis s. sicula) testis. Chieffi P, Angelini F, Pierantoni R. Dipartimento di Fisiologia Umana e Funzioni Biologiche Integrate F. Bottazzi, II Universita di Napoli, Naples, Italy. PMID: 9629217 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 354: Cytokine. 1998 May;10(5):331-6. Function of the human interleukin 4 receptor (IL-4R)-derived acidic motif revealed by cytoplasmic domain chimeras of the IL-4R alpha chain and the IL-2R beta chain. Deutsch HH, Chung J, Kalthoff FS. Novartis Research Institute, Department of Immunology, Vienna, Austria. Interleukin 2 (IL-2)- and IL-4-mediated stimulation of survival and growth, reflected by the induction of bcl2 and c-myc, respectively, depends on the integrity of the membrane-proximal region (S-region) in the IL-2 receptor beta-chain (IL-2R beta) and the haematopoietin homology box1-containing region of the IL-4 receptor alpha-chain (IL-4R alpha). In contrast to IL-4, IL-2 induces the expression of c-fos and c-jun family genes, mediated by the acidic region (A-region) within the cytoplasmic domain of IL-2R beta. A highly acidic motif is also present in IL-4R alpha, and evidence in favour and against its importance has been published. The authors have constructed chimeric receptors between IL-2R beta and IL-4R alpha by substitution of either the S-region or the A-region of IL-2R beta with sequences derived from IL-4R alpha. These chimeras were stably transfected into BA/F3 cells and assayed for the capacity to restore functions of IL-2 beta, such as growth mediation by IL-2 and the induction of proto-oncogenes (c-myc, c-junB and c-fos). Replacement of both the S- and A-region of IL-2R beta with IL-4R alpha derived regions of similar size and cytoplasmic location supported growth-stimulation by IL-2 as well as proto-oncogene induction. In contrast, all IL-2R functions were lost by exchange of the S-region with the corresponding part of IL-4R alpha. Induction of c-junB and c-fos RNA as an indicator of A-region function, however, was maintained in an IL-2R beta chimera containing the acidic box-bearing region of IL-4R alpha. These data indicate a functional role of the acidic region in the IL-4R alpha-chain. PMID: 9619370 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 355: Biochemistry (Mosc). 1998 Feb;63(2):149-54. Changes in structural organization of the nucleosome fiber during protooncogene activation. Terent'ev AA, Kostyuk GV, Boikov PY. Institute of Chemical Physics, Russian Academy of Sciences, Chernogolovka, Moscow Region, 142432 Russia. boikov@icp.ac.ru Dynamics of structural changes in the chromatin of rat liver cells were studied during activation of the c-myc, c-fos, and c-jun protooncogenes. Nuclei were isolated at various stages of gene activation using cycloheximide and chromatin was subjected to partial nucleolysis by endogenous Ca2+/Mg2+-DNAse and extracted with low magnesium buffer; the nucleosome fragmentation was analyzed. DNA was isolated from the soluble chromatin and low-molecular-weight fragments were separated by electrophoresis through a composite gel system (3 and 1% agarose). This modification of DNA electrophoresis protocol improves the efficiency of separation of the low-molecular-weight DNA fragments. Increased protooncogene activity is associated with decreased content of DNAse-sensitive soluble chromatin. The fractional content of the c-fos gene is increased in the soluble chromatin. In control nuclei, endogenous Ca2+/Mg2+-DNAse cleaves mononucleosomes containing DNA fragments with variable length. Upon protooncogene activation, mono- and oligonucleosomes with elongated DNA fragments are predominantly formed. The changes involve about 40% of the nuclear chromatin. The data suggest that protooncogene activation is associated with reorganization at the nucleosome level of the structural organization of the chromatin. PMID: 9526106 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 356: Zhonghua Yi Xue Za Zhi. 1997 Mar;77(3):194-6. [Growth regulatory mechanisms of recombinant human interleukin-6 on human lung carcinoma cell lines] [Article in Chinese] Fu J, Wu B, Zheng J. Department of Pathology, Beijing Medical University. OBJECTIVES: To detect the activity of interleukin-6 (rhIL-6) on human lung cancer and the influences of recombinant human IL-6 (rhIL-6) on the in vitro growth of high-metastatic human lung giant cell carcinoma cell line PG and low-metastatic human lung adenocarcinoma cell line PAa. METHODS: The effects of rhIL-6 on the expressions of IL-6, IL-6R, c-myc, c-fos and c-jun mRNAs were analyzed by Northern blotting hybridization. RESULTS: rhIL-6 could stimulate the in vitro growth of both PG and PAa cells, in a dose-and-time-dependent manner. Both PG and PAa cells were shown to express IL-6 and IL-6R identified by reverse transcription PCR analysis. Northern blotting hybridization revealed that rhIL-6 up-regulated mRNA levels of IL-6, IL-6R and c-myc rather than those of c-fos and c-jun. CONCLUSION: These data suggest that rhIL-6 may promote the proliferation of human lung cancer cell lines PG and PAa, which might, at least partly, be related to the up-regulation of expressions of IL-6, IL-6R, c-myc gene transcript. PMID: 9596958 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 357: J Cardiovasc Pharmacol. 1998 May;31(5):786-93. Amlodipine inhibition of serum-, thrombin-, or fibroblast growth factor-induced vascular smooth-muscle cell proliferation. Stepien O, Gogusev J, Zhu DL, Iouzalen L, Herembert T, Drueke TB, Marche P. Universite Rene Descartes & Department of Pharmacology, CNRS URA 1482, CHU Necker, Paris, France. Atherosclerosis, like several other vascular diseases, exhibits structural and functional abnormalities resulting partially from an exaggerated proliferation of vascular smooth-muscle cells (VSMCs). Ca2+ channel blockers, such as amlodipine, have been suggested to retard or even prevent the progression of atherosclerosis. To determine the mechanisms involved in these effects, we investigated the influence of amlodipine on VSMC proliferation by using rat aortic VSMCs in culture. Amlodipine (0.1-10 microM) inhibited serum-, basic fibroblast growth factor (bFGF)-, and thrombin-induced VSMC proliferation and DNA synthesis in a concentration-dependent manner, as demonstrated by cell count and bromodeoxyuridine (BrdU)-incorporation measurements, respectively. Delayed addition of amlodipine after VSMC stimulation showed that the drug exerted its effect early in G1 phase of the cell cycle. This observation was confirmed by the finding that amlodipine did not influence DNA synthesis in VSMCs arrested to the G1/S boundary by hydroxyurea treatment. Consistent with its effects on VSMC growth/proliferation, amlodipine also decreased c-myc, c-fos, and c-jun protooncogene expression induced by serum, thrombin, or bFGF within 1 h after cell activation, as assessed by semiquantitative reverse transcriptase (RT)-polymerase chain reaction (PCR) analysis. The calcium channel agonist Bay K 8644, which counteracted the inhibition by nifedipine of bFGF-, thrombin- or serum-induced DNA synthesis, was ineffective to antagonize the inhibitory effect of amlodipine. The aforementioned effects of amlodipine were of similar amplitude, irrespective of the growth-enhancing agent used. This strongly indicates that amlodipine acts downstream of receptor activation to exert its antiproliferative action, probably early in the G1 phase of the cell cycle. Moreover, the lack of antagonistic effect between amlodipine and Bay K 8644 suggests that, in addition to its L-type Ca2+ channel inhibitory effect, amlodipine inhibits other intracellular signaling pathways. Such an interference of amlodipine with mitogenic signaling pathways might contribute to confer a blood vessel-protecting potential on amlodipine. PMID: 9593080 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 358: Melanoma Res. 1997 Aug;7 Suppl 2:S7-17. Role of interleukin (IL)-2 and IL-15 in the tumour progression of a melanoma cell line MELP, derived from an IL-2 progressor patient. Doucet C, Meazza R, Pottin-Clemenceau C, Scudeletti M, Brouty-Boye D, Ferrini S, Alileche A, Taoufik Y, Jasmin C, Azzarone B, Indiveri F. Unite 268 INSERM, Hopital Paul Brousse, Villejuif, France. MELP is an interleukin (IL)-2 receptor (IL-2R; alpha+ beta+ gamma-) melanoma cell line that was derived, before the beginning of the immunotherapy, from a patient whose metastasis increased in size during treatment with IL-2/interferon-alpha. In these cells, continuous culture in the presence IL-2 (1000 UI/ml) causes the selection of a cell sub-line (termed MILG) expressing the gamma-chain which is tumorigenic in nude mice. Here, we further analysed the characteristics of MELP and MILG cells as well as clones selected at limiting dilution in the presence of high concentrations of IL-2 or IL-15, or those selected after transfection for the expression of a human IL-2 transgene (MELP-CL1). MELP cells, but not six other melanomas cell lines, shed two soluble immunosuppressive molecules, CD25 and intercellular adhesion molecule-1, whose levels also strongly increase in vivo during immunotherapy. In vitro MELP cells express transcripts for IL-6, transforming growth factor, basic fibroblast growth factor and vascular-endothelial growth factor. Cloning at limiting dilution was obtained in culture fed with IL-2 or IL-15. All these clones, as MILG cells, express the transcript for the IL-2R gamma chain. This could favour improved interactions with cytokines using this chain. By contrast, MELP-CL1 cells, which secrete low amounts of biologically active IL-2 (200 UI/10(6) cells) exhibit a phenotype and growth characteristics similar to those of the parental MELP cells. Indeed, a crosslinking experiment with 125I-IL-2, has showed that MELP and MELP-CL1 cells display a scant IL-2 binding ability that is strongly increased in MELP cells fed for 1 week with 1000 UI/ml IL-2. These cells, as well as MILG cells express a betagamma-complex which can also bind IL-15. IL-2 induces a rapid tyrosine phoshorylation in MILG cells, which is followed by a prolonged induction of c-fos and c-jun genes. By contrast, in MELP cells IL-2 only causes a delayed induction of c-myc gene. All MELP derivatives, but not MILG cells, express the transcripts for IL-15, which is not secreted but is present as an intracellular protein. All MELP cells express the transcript for the IL-15R alpha chain. MELP-CL1 cells are not tumorigenic in nude mice, whereas MILG cells form rapidly growing tumours in 75% of the mice. Coinjection at the same site of MILG and MELP-CL1 cells causes the rapid regression of MILG tumours in 80% of the mice, whereas their bilateral injection causes the rapid development of MILG tumours in 100% of the nude mice. Finally, treatment in nude mice of MILG cells with low amounts of IL-2 (1000 UI per mouse) and IL-15 (50 ng per mouse) induces the development of much more aggressive tumours.The expression of functional IL-2Rs in a subset of human melanomas could be responsible for tumour progression. Publication Types: Case Reports PMID: 9578412 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 359: Gynecol Oncol. 1998 Mar;68(3):280-7. Value of glutathione S-transferase pi and the oncogene products c-Jun, c-Fos, c-H-Ras, and c-Myc as a prognostic indicator in endometrial carcinomas. Yokoyama Y, Sagara M, Sato S, Saito Y. Department of Obstetrics and Gynecology, Hirosaki University School of Medicine, Japan. OBJECTIVE: To examine the relationship between the expressions of glutathione S-transferase pi (GST-pi) and four oncogene products, c-Jun, c-Fos, c-H-Ras, and c-Myc, and clinicopathological prognostic factors and patients' prognosis in endometrial carcinomas, and to assess their prognostic value in endometrial carcinomas. METHODS: Specimens of endometrial carcinoma obtained from 63 patients were investigated immunohistochemically using respective specific antibodies. RESULTS: The overall positive rates in 63 carcinoma specimens were 34.9% for GST-pi, 44.4% for c-Jun, 34.9% for c-Fos, 47.6% for c-H-Ras, and 54.0% for c-Myc. Multivariate analysis revealed that GST-pi expression correlated independently with paraaortic lymph node (PAN) metastasis, and c-Jun expression was independently related to pelvic lymph node (PLN) and PAN metastasis. The prognosis of patients with a GST-pi-positive tumor was significantly poorer than that of those with a GST-pi-negative tumor (P < 0.05). The patients with c-Jun-positive tumor also had a significantly worse prognosis than those with c-Jun-negative tumor (P < 0.05). No significant relationship between the expressions of the remaining three oncogene products, c-Fos, c-H-Ras, and c-Myc, and the examined prognostic factors and clinical outcome was apparent. CONCLUSION: These results suggest that the expressions of GST-pi and c-Jun may reflect the metastatic potential of endometrial carcinomas and that their expressions of endometrial carcinoma may be useful as a prognostic indicator for predictive testing. PMID: 9570981 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 360: Oncogene. 1998 Mar 26;16(12):1525-31. The OMgp gene, a second growth suppressor within the NF1 gene. Habib AA, Gulcher JR, Hognason T, Zheng L, Stefansson K. Department of Neurology, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA. The Oligodendrocyte-Myelin glycoprotein gene (OMgp) is placed within an intron of the NF1 gene. Neurofibromin, the product of NF1, acts as a RasGAP and suppresses growth; inactivating mutations in NF1 lead to neurofibromatosis type 1. We report that OMgp also has growth suppressive effects and downregulates mitogenic signaling pathways closely related to those influenced by neurofibromin. Overexpression of OMgp alters mitogenic signaling in NIH3T3 fibroblasts. Cells overexpressing OMgp grow more slowly in serum compared to controls and show a partial G1 block upon cell cycle analysis. PDGF is the primary mitogen for fibroblasts in serum. Overexpression of OMgp alters PDGF signaling in fibroblasts which results in a block of mitogenic signaling. PDGF induced activation of c-Src is blocked, as is the induction of c-Myc and c-Fos, while tyrosine phosphorylation of the PDGFbeta receptor, PLCgamma1 and induction of c-Jun are intact. Although a number of genes embedded within other genes have been described, the biological significance of this arrangement remains unknown. We demonstrate here that structurally unrelated products of two such genes may exercise closely related functions. Our data also raise the possibility of a role for OMgp in disorders of cell proliferation such as NF1. PMID: 9569019 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 361: Mol Cell Biol. 1998 May;18(5):2659-67. Increased c-fos mRNA expression by human fibroblasts contracting stressed collagen matrices. Rosenfeldt H, Lee DJ, Grinnell F. Department of Cell Biology and Neuroscience, University of Texas Southwestern Medical School, Dallas 75235-9039, USA. We studied early changes in gene expression during fibroblast contraction of stressed collagen matrices. The level of c-fos mRNA increased dramatically and peaked 50 to 60 min after matrix contraction was initiated. This response did not require serum and could not be accounted for simply by disruption of the actin cytoskeleton. Increased c-fos mRNA levels required Ca2+ influx but not the cyclic AMP or extracellular signal-regulated kinase (ERK 1/2) signaling pathways, both of which are activated when fibroblasts contract stressed collagen matrices. The levels of two other immediate-early genes, fosb and c-jun, also increased transiently after fibroblast contraction, whereas the levels of fra-1, fra-2, c-myc, and the transcription factor NF-kappaB remained the same, indicating that fibroblast contraction caused changes in a selective group of genes. The increase in c-fos mRNA during contraction of stressed collagen matrices may reflect a unique role for c-fos in mechanoregulated events at the end of wound repair. PMID: 9566885 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 362: Cancer Lett. 1998 Mar 13;125(1-2):141-8. Epithelial cell proliferation in the digestive tract induced by space restriction and water-immersion stress. Hori T, Wanibuchi H, Yano Y, Otani S, Nishikawa A, Osugi H, Kinoshita H, Fukushima S. First Department of Pathology, Osaka City University Medical School, Osaka, Japan. The effects of space restriction and water-immersion stress on epithelial cell proliferation in the digestive tract, with special attention to the esophagus, stomach and duodenum, in 8-week-old SD male rats were examined. Histological assessment revealed spotted hemorrhagic lesions in the fundus of the glandular stomach, accompanied by statistically increased 5-bromo-2'-deoxyuridine (BrdU) labeling index in the fundic and pyloric regions. Furthermore, biochemical analysis demonstrated an increased activity of ornithine decarboxylase (ODC) and spermidine/spermine N1-acetyltransferase (SAT), known as key late-limiting enzymes of the polyamine pathway, in the gastric fundus. The stress may induce a remarkable increase in expression of c-fos, c-jun and c-myc mRNAs in both fundic and pyloric regions of the glandular stomach. There were no remarkable changes in the esophagus. These results indicate that space restriction and water-immersion stress induced cell proliferation in the glandular stomach through overexpression of proto-oncogenes and increased ODC and SAT activities that might be related to the promotion of gastric carcinogenesis. PMID: 9566708 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 363: J Cell Biochem. 1998 May 1;69(2):189-200. Inhibition of PPAR alpha/RXR alpha-mediated direct hyperplasia pathways during griseofulvin-induced hepatocarcinogenesis. Nagao Y, French BA, Cai Y, French SW, Wan YJ. Department of Pathology, Harbor-UCLA Medical Center, Torrance, California 90509, USA. Chronic griseofulvin (GF) feeding induces preneoplastic foci followed by hepatocellular carcinoma in the mouse liver. Our previous study suggested that GF-induced hepatocellular proliferation had a different mechanism from that of peroxisome proliferator (PP)-induced direct hyperplasia. The GF-induced hepatocellular proliferation was mediated through activation of immediate early genes such as Fos, Jun, Myc, and NFKB. In contrast, PP-induced direct hyperplasia does not involve activation of any of these immediate early genes. It has been shown that nuclear hormone receptors including peroxisome proliferator activated receptors (PPARs) and retinoid x receptors (RXRs) play important roles in mediating the pleiotropic effects of PPs. To examine the possible roles of PPARs and RXRs during non-PP-induced hepatocellular proliferation and the interaction between PP and non-PP-induced proliferation, we have studied the expression of the PPAR and RXR genes in the GF model using northern blot hybridizations and gel retardation assays. The data showed that the expression of PPARalpha and RXRalpha genes was down-regulated in the livers containing preneoplastic nodules and in the liver tumors induced by GF. The mRNA down-regulation was accompanied by a decrease in the amount of nuclear protein-bound to peroxisome proliferator and retinoic acid responsive elements. Down-regulation was also associated with the suppressed expression of the PPARalpha/RXRalpha target genes (i.e., acyl-Co oxidase and cytochrome P450 4A1) and the catalase gene. The RXR-gamma gene was also down-regulated, but the RARalpha, beta, and gamma and PPARbeta and gamma genes were up-regulated. These results indicated that the hepatocarcinogenesis induced by GF is accompanied by suppression of the PPARalpha/RXRalpha-mediated direct hyperplasia pathway. The differential expression of these nuclear hormone receptors reveals a new aspect for understanding the individual roles and intercommunication of PPAR, RXR, and RAR isoforms in the liver. PMID: 9548566 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 364: J Biol Chem. 1998 Mar 13;273(11):6013-8. The polycystic kidney disease 1 gene product mediates protein kinase C alpha-dependent and c-Jun N-terminal kinase-dependent activation of the transcription factor AP-1. Arnould T, Kim E, Tsiokas L, Jochimsen F, Gruning W, Chang JD, Walz G. Renal Division, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA. Autosomal dominant polycystic kidney disease (ADPKD) is a common hereditary disorder that accounts for 8-10% of end stage renal disease. PKD1, one of two recently isolated ADPKD gene products, has been implicated in cell-cell and cell-matrix interactions. However, the signaling pathway of PKD1 remains undefined. We found that the C-terminal 226 amino acids of PKD1 transactivate an AP-1 promoter construct in human embryonic kidney cells (293T). PKD1-induced transcription is specific for AP-1; promoter constructs containing cAMP response element-binding protein, c-Fos, c-Myc, or NFkappaB-binding sites are unaffected by PKD1. In vitro kinase assays revealed that PKD1 triggers the activation of c-Jun N-terminal kinase (JNK), but not of mitogen-activated protein kinases p38 or p44. Dominant-negative Rac-1 and Cdc42 mutations abrogated PKD1-mediated JNK and AP-1 activation, suggesting a critical role for small GTP-binding proteins in PKD1-mediated signaling. Several protein kinase C (PKC) inhibitors decreased PKD1-mediated AP-1 activation. Conversely, expression of the C-terminal domain of PKD1 increased PKC activity in 293T cells. A dominant-negative PKC alpha, but not a dominant-negative PKC beta or delta, abrogated PKD1-mediated AP-1 activation. These findings indicate that small GTP-binding proteins and PKC alpha mediate PKD1-induced JNK/AP-1 activation, together comprising a signaling cascade that may regulate renal tubulogenesis. PMID: 9497315 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 365: Biochem Mol Biol Int. 1998 Feb;44(2):217-24. Inductions of immediate early genes (IEGS) and ref-1 by human chorionic gonadotropin in murine Leydig cell line (MA-10). Suzuki S, Nagaya T, Suganuma N, Tomoda Y, Seo H. Department of Endocrinology and Metabolism, Research Institute of Environmental Medicine, Nagoya University, Japan. The effect of human chorionic gonadotropin (hCG) on the expression of immediate early genes (IEGs) including all members of fos and jun family, and c-myc was studied using mouse Leydig cell line (MA-10 cells) by Northern blot analyses. In addition, the induction of ref-1 which enhances DNA binding of fos/jun proteins was also analyzed. HCG induced a rapid and transient expression of c-fos, fosB, c-jun, junB, junD and c-myc with a peak at 30 min to 1 h. In contrast, induction of fra-1 mRNA was delayed with a peak at 3 hr. However, fra-2 mRNA was immediately increased by hCG with a peak at 1 h. The ref-1 mRNA was expressed before the stimulation and its level was not altered by hCG at least for 8 hr. The differential induction of IEGs and continuous expression of ref-1 mRNA suggest an important role of their gene products on the regulation of Leydig cell function by hCG. PMID: 9530505 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 366: Carcinogenesis. 1998 Mar;19(3):471-7. Prevention of estrogen carcinogenesis in the hamster kidney by ethinylestradiol: some unique properties of a synthetic estrogen. Li JJ, Hou X, Bentel J, Yazlovitskaya EM, Li SA. Kansas Cancer Institute, Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City 66160-7312, USA. jli1@kumc.edu Ethinylestradiol (EE) has evident paradoxical effects on cancer risk for human breast and hepatic cancer which parallel in some respects its effects on estrogen-induced neoplasms in the hamster kidney and liver. EE has been shown to be only weakly carcinogenic in the hamster kidney, but the most potent carcinogenic estrogen in the hamster liver following prolonged treatment. Unexpectedly, when EE and potent carcinogenic estrogens, such as diethylstilbestrol (DES), 17beta-estradiol (E2) and Moxestrol (MOX), are administered concomitantly, estrogen-induced carcinogenesis in the kidney is completely prevented. In studying this novel finding, we found that, compared with E2 exposure alone, EE at 0.05 and 1.0 nM significantly (P < 0.001) inhibited the rise in proliferation of cultured primary hamster proximal renal tubular (PRT) cells in the presence of E2 (1.0 nM). Consistent with these findings, combined EE + DES treatment for 5.0 months reduced hamster kidney c-myc, c-fos and c-jun RNA expression to 43, 37 and 52%, respectively, compared with levels observed after DES treatment alone. Interestingly, TAM + DES treatment for the same period also resulted in the same low level of RNA expression of these proto-oncogenes. c-MYC, c-FOS and c-JUN protein products were comparably reduced after either EE + DES or TAM + DES treatment. It appears that c-fos expression and c-FOS protein levels in the hamster kidney were more responsive to TAM inhibition. These data demonstrate that EE possesses unique anti-tumorigenic properties in vivo in the hamster kidney. Additionally, the observed anti-estrogen-like effect of EE on cell proliferation of cultured PRT cells suggests that EE may interfere critically with estrogen receptor (ER)-mediated mitogenic pathway(s) affected by potent carcinogenic estrogens, thus preventing subsequent gene dysregulation and, hence, tumor development. Based on competition studies, the differential binding of EE to hamster kidney ER relative to that of the other estrogens (E2, DES, MOX) appears not to contribute to the prevention of estrogen carcinogenesis at this organ site by EE. PMID: 9525282 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 367: J Neurochem. 1998 Apr;70(4):1474-83. Extracellular calcium deprivation in astrocytes: regulation of mRNA expression and apoptosis. Chiesa R, Angeretti N, Del Bo R, Lucca E, Munna E, Forloni G. Biology of Neurodegenerative Disorders Laboratory, Istituto di Ricerche Farmacologiche Mario Negri, Milano, Italy. Cell viability and gene expression were studied in primary astroglial cells cultured in a nominally calcium-free medium. Ca2+ deprivation reduced progressively the astrocytes' viability, starting from 12 h; the restoration of a normal Ca2+ concentration (1.8 mM) in the medium after 12-h deprivation reversed the degenerative effect within 24 h. Biochemical and morphological examinations indicated that cell death induced by Ca2+ deprivation was mediated by apoptosis. This was associated with the expression of c-fos, c-jun, and c-myc, which, with different time courses, were induced in astrocytes after Ca2+ deprivation. Furthermore, shifting to a Ca2+-free medium modified the expression of Ich-1S transcript and rapidly increased intracellular cyclic AMP, which has been implicated in the transcriptional activation of immediate-early genes. The absence of Ca2+ in the medium reduced the expression of constitutive proteins such as alpha-actin, clusterin, glial fibrillary acidic protein, amyloid precursor protein, and glucose-6-phosphate dehydrogenase. The expression of these mRNAs was reduced >50% after 8 h of Ca2+ deprivation, when the effect on cell viability was negligible. When Ca2+ deprivation was prolonged for 24 h the expression of mRNA dropped completely, and restoration of the Ca2+ ions in the medium for 48 h did not reverse this effect. In contrast with general assumption, the apoptotic machinery in astrocytes is activated similarly not only by increased Ca2+ influx but also with the extracellular Ca2+ deprivation. PMID: 9523564 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 368: Biochem Biophys Res Commun. 1998 Mar 6;244(1):167-71. The protein kinase C activator, phorbol ester, elicits disparate functional responses in androgen-sensitive and androgen-independent human prostatic cancer cells. Henttu P, Vihko P. Biocenter Oulu, University of Oulu, Finland. The protein kinase C (PKC) activator 12-O-tetradecanoyl-phorbol-13-acetate (TPA) activated cell death in androgen-sensitive LNCaP cells but not in androgen-independent DU-145 or PC-3 cells, whose growth was significantly decreased by PKC inhibitors staurosporine and H7. All cell lines had similar levels of total PKC activities which, however, differed on their dependency on Ca2+ ions and lipid and were regulated differently by TPA. Furthermore, expression of the immediate early genes c-fos and c-jun was up-regulated by TPA only in LNCaP and DU-145 cells, whereas PC-3 cells failed to express c-fos mRNA. The regulation of the c-myc mRNA by TPA correlated inversely with activation of cell death being down-regulated in LNCaP cells, and slightly increased in the androgen-independent cell lines. These results suggest that the PKC signal transduction pathway functions differently in androgen-sensitive and insensitive prostatic cells. PMID: 9514905 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 369: Gynecol Oncol. 1998 Feb;68(2):218. Papers to Appear in Forthcoming Issues [No authors listed] Concomitant Endometrial Hyperplasia in Patients with Endometrial Carcinoma Fatih Gucer, Olaf Reich, Karl Tamussino, Armin A. Bader, Doris Pieber, Wolfgang Scholl, Josef Haas, and Edgar Petru Decline in Incidence of Endometrial Cancer Following Increase in Prescriptions for Opposed Conjugated Estrogens in a Prepaid Health Plan Harry K. Ziel, William D. Finkle, and Sander Greenland Treatment for Fertility and Risk of Ovarian Tumors of Borderline Malignancy Fabio Parazzini, Eva Negri, Carlo La Vecchia, Simona Moroni, Anna Polatti, Francesca Chiaffarino, and Elena Ricci Immunohistochemical Expression of Thymidine Phosphorylase in Human Endometrial Cancer Ritsuto Fujiwaki, Kohkichi Hata, Kohji Iida, Mikio Koike, and Kohji Miyazaki Palliative Benefit of External-Beam Radiation in the Management of Platinum Refractory Epithelial Ovarian Carcinoma Daphna Gelblum, Borys Mychalczak, Lois Almadrones, David Spriggs, and Richard Barakat Virilizing Ovarian Tumor of Low Malignant Potential Associated with Antecedent Tamoxifen Use for Breast Cancer Daniel V. Surbek, Irene Hoesli, Joachim Torhorst, Alfonso C. Almendral, Athanassios Dellas, and Wolfgang Holzgreve Value of Glutathione S-Transferase pi and the Oncogene Products c-Jun, c-Fos, c-H-Ras, and c-Myc as a Prognostic Indicator in Endometrial Carcinomas Yoshihito Yokoyama, Morio Sagara, Shigemi Sato, and Yoshiharu Saito Prevalence, Risk Factors, and Accuracy of Cytologic Screening for Cervical Intraepithelial Neoplasia in Women with the Human Immunodeficiency Virus Mitchell Maiman, Rachel G. Fruchter, Alexander Sedlis, Joseph Feldman, Patrick Chen, Robert D. Burk, and Howard Minkoff The Value of Squamous Cell Carcinoma Antigen as a Predictor of Nodal Metastasis in Cervical Cancer Nobuhiro Takeshima, Yasuo Hirai, Katsuyoshi Katase, Kenji Yano, Kazuhiro Yamauchi, and Katsuhiko Hasumi Five-Year Survival after Second-Line Cisplatin-Based Intraperitoneal Chemotherapy for Advanced Ovarian Cancer Fernando O. Recio, M. Steven Piver, Ronald E. Hempling, and Deborah L. Driscoll Evidence for a Common Etiology for Endometrial Carcinomas and Malignant Mixed Mullerian Tumors Alice Zelmanowicz, Allan Hildesheim, Mark E. Sherman, Susan R. Sturgeon, Robert J. Kurman, Rolland J. Barrett, Michael L. Berman, Rodrigue Mortel, Leo B. Twiggs, George D. Wilbanks, and Louis A. Brinton Estimation of Probability of Malignancy Using a Logistic Model Combining Physical Examination, Ultrasound, Serum CA 125, and Serum CA 72-4 in Postmenopausal Women with a Pelvic Mass: An International Multicenter Study E. M. J. Schutter, C. Sohn, P. Kristen, V. Mobus, G. Crombach, M. Kaufmann, H. Caffier, R. Kreienberg, A. A. Verstraeten, and P. Kenemans Influence of Quantity of Lymph-Vascular Space Invasion on the Risk of Nodal Metastases in Women with Early-Stage Squamous Cancer of the Cervix Lynda D. Roman, Juan C. Felix, Laila I. Muderspach, Taz Varkey, Alexander F. Burnett, Dajun Qian, and C. Paul Morrow Unsuspected Primary Tubal Carcinoma during Operative Laparoscopy Rene Wenzl, Rainer Lehner, Michael Drager, Stefan Jirecek, Christian Gamper, and Paul Sevelda An HIV-Infected Woman with Choriocarcinoma Presenting with a Nasal Mass Somsak Tangtrakul, Vasant Linasmita, Sarikapan Wilailak, Somkeart Srisupandit, Sunchai Bullangpoti, and Nathapong Israngura Na Ayudhya Incidence of Ovarian Cancer in Women with Prior Hysterectomy in Japan Nobuo Yaegashi, Shinji Sato, and Akira Yajima Copyright 1998 Academic Press. PMID: 9514817 [PubMed - as supplied by publisher] --------------------------------------------------------------- 370: Biol Reprod. 1998 Mar;58(3):778-85. Phorbol ester inhibition of estrogen-induced uterine deoxyribonucleic acid synthesis. Kirkland JL, Murthy L, Thomazy V, Hyder SM, Stancel GM. Department of Pediatrics, Baylor College of Medicine, Houston, Texas 77030, USA. kirkland@bcm.tmc.edu Protein kinase C (PKC) is a key regulatory enzyme in the control of growth and differentiated function in many cell types. Recently it has become clear that cross talk occurs between PKC and steroid hormone-activated signaling pathways. In this work we have thus used the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) to investigate the relationship between PKC activation and estrogen-induced proliferation in an in vivo model of hormone action, the immature rat uterus. A single injection of estradiol (E2) to immature female animals increases DNA synthesis in all major uterine cell types. Administration of TPA alone, simultaneous administration of TPA and E2, or administration of TPA 12 h after E2 did not alter uterine DNA synthesis. However, administration of TPA 6 h after E2 markedly decreased [3H]thymidine incorporation and the labeling indices in uterine epithelial, stromal, and myometrial cells. This inhibition represents a decrease in DNA synthesis per se rather than a change in the time course of the tissue response to the hormone. The inhibitory effect of TPA was reversible within 72 h and did not appear to be due to a decrease in the level or degree of occupancy of uterine estrogen receptors. These results suggest that a discrete regulatory event(s) in the pathway of estradiol-induced proliferation is inhibited by PKC activation approximately 6 h after hormonal stimulation. PMID: 9510966 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 371: Genomics. 1998 Feb 15;48(1):87-92. Genomic structure and chromosomal localization of the mouse LIM domain-binding protein 1 gene, Ldb1. Yamashita T, Agulnick AD, Copeland NG, Gilbert DJ, Jenkins NA, Westphal H. Laboratory of Mammalian Genes and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892-2790, USA. The LIM domain is a structural motif that is well conserved throughout evolution in a variety of factors known to play important roles in development and cell regulation. Ldb genes encode LIM domain-binding (Ldb) factors. Here we report on the structural organization and chromosomal localization of the mouse Ldb1 gene. It contains at least 10 exons and spans approximately 4 kb of genomic DNA. The transcription initiation site is located 462 bp upstream of the translation initiation codon ATG as determined by 5'-RACE. Sequencing analysis of the 5'-flanking region shows TATA and CCAAT motifs as well as potential binding sites for GATA, CF-1, PEA3, HRE, APRRE, RARE, Myc, and c-Jun. Ldb1 maps to the distal region of mouse chromosome 19 that is syntenic with human chromosome 10q. PMID: 9503020 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 372: Cancer Res. 1998 Mar 1;58(5):1027-33. Ets-2 transdominant mutant abolishes anchorage-independent growth and macrophage colony-stimulating factor-stimulated invasion by BT20 breast carcinoma cells. Sapi E, Flick MB, Rodov S, Kacinski BM. Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, Connecticut 06520-8040, USA. eva.sapi@yale.edu Activation of the macrophage colony-stimulating factor receptor (CSF-1R) by its cognate ligand CSF-1 dramatically increases the tumorigenicity and invasive potential of both normal and neoplastic mammary epithelial cells. Recent studies have suggested that the Ets2 transcription factor plays a central role in mediating CSF-1R-induced mitogenesis in fibroblasts. To determine whether the Ets2 transcription factor can also mediate CSF-1- and CSF-1R-stimulated signaling pathways in mammary epithelial cells, we expressed a dominant negative mutant, Ets2, in the CSF-1R- and Ets2-positive BT20 breast carcinoma cell line and examined its effects on CSF-1-induced cellular invasion and on colony formation in soft agar. Our data show that stable expression of the mutant Ets2 in BT20 cells completely inhibits the formation of soft agar colonies and abolishes the CSF-1-stimulated invasion of these cells through a barrier of reconstituted basement membrane (Matrigel). We have also demonstrated that the expression of this Ets2 mutant is capable of interrupting the CSF-1R-regulated intracellular signaling pathways by inhibiting CSF-1-induced c-myc, c-fos, and c-jun expression in BT20 cells. Our results are the first demonstration of an important role for the Ets2 transcription factor in the regulation of the anchorage-independent growth and cellular invasiveness of neoplastic mammary epithelial cells. PMID: 9500466 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 373: Cancer Res. 1998 Mar 1;58(5):867-70. Suppression of cellular transformation by glucocorticoid receptor mutants. Kameda T, Iba H. Department of Gene Regulation, Institute of Medical Science, University of Tokyo, Japan. Glucocorticoid receptor (GR) has been shown to suppress activator protein 1 (AP-1)-mediated transcription by several molecular mechanisms. We previously showed that activation of endogenous AP-1 is essential for cellular transformation induced by oncogenes, such as v-src and c-Ha-ras. In the present study, we have analyzed whether high levels of GR expression suppress cellular transformation caused by these oncogenes. To eliminate the ligand effects that induce the transcriptional stimulation via glucocorticoid response elements, we constructed two GR mutants: CD-GR-1, lacking the COOH-terminal portion, including both the ligand and Hsp90-binding domains, and tau1TDCD-GR-1, a derivative of CD-GR-1 lacking the tau1 transactivation domain. When these GR mutants were expressed in chicken embryonic fibroblasts by retroviral vectors, they translocated into the nucleus without addition of glucocorticoid to the culture medium, and they suppressed cellular transformation caused by v-src and c-Ha-ras, as well as by c-fos and c-jun. Cellular transformation by v-myc was not suppressed by these mutants. Such suppressive effects of these GR mutants have a very similar oncogene dependency to that of dominant-negative mutants of AP-1. This suggests that GR can be altered to suppress cellular transformation by inhibiting endogenous AP-1 activity without activating glucocorticoid response elements. PMID: 9500440 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 374: Arterioscler Thromb Vasc Biol. 1998 Feb;18(2):227-34. Shear stress activates p60src-Ras-MAPK signaling pathways in vascular endothelial cells. Jalali S, Li YS, Sotoudeh M, Yuan S, Li S, Chien S, Shyy JY. Department of Bioengineering and Institute for Biomedical Engineering, University of California, San Diego, La Jolla 92093-0412, USA. The aim of this study was to elucidate the upstream signaling mechanism that mediates the fluid shear stress activation of mitogen-activated protein kinases (MAPKs), including c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinases (ERKs), in vascular endothelial cells (ECs). Our results indicate that p60src is rapidly activated by fluid shear stress in bovine aortic endothelial cells (BAECs). Shear stress induction of the hemagglutinin (HA) epitope-tagged HA-JNK1 and the Myc epitope-tagged Myc-ERK2 was significantly attenuated by v-src(K295R) and c-src(K295R), the kinase-defective mutants ofv-src and c-src, respectively. HA-JNK1 and Myc-ERK2 were activated by c-src(F527), a constitutively activated form of p60src, and the activation was abolished by RasN17, a dominant-negative mutant of p2lras. In contrast, although HA-JNK1 and Myc-ERK2 were also activated by RasL61, an activated form of p21ras, the activation was not affected by v-src(K295R). These results indicate that p60src is upstream to the Ras-JNK and Ras-ERK pathways in response to shear stress. The shear stress inductions of the promoters of monocyte chemotactic protein-1 (MCP-1) and c-fos, driven by TPA-responsive element (TRE) and serum-responsive element (SRE), respectively, were attenuated by v-src(K295R). This attenuation is associated with decreased transcriptional activities of c-Jun and Elk-1, the transcription factors targeting TRE and SRE, respectively. Thus, p60src plays a critical role in the shear stress activation of MAPK pathways and induction of Activating Protein-1 (AP- 1)/TRE and Elk-1/SRE-mediated transcription in ECs. PMID: 9484987 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 375: Br J Cancer. 1998 Feb;77(4):663-9. Erratum in: Br J Cancer 1998 Apr;77(7):1198. Prognostic value of ERBB-1, VEGF, cyclin A, FOS, JUN and MYC in patients with squamous cell lung carcinomas. Volm M, Rittgen W, Drings P. Department of Oncological Diagnostics and Therapy, German Cancer Research Center, Heidelberg. Patients with previously untreated squamous cell lung carcinomas were evaluated to see if combining the expression of molecular and cellular factors with the most important clinical prognostic factors could improve the diagnostic ability to predict prognosis. For this reason, immunohistochemistry was used to examine the squamous cell lung carcinomas from 121 patients for their expression of ERBB-1, vascular endothelial growth factor (VEGF), cyclin A, FOS, JUN and MYC. Median survival was shorter for patients with ERBB-1-, VEGF-, cyclin A-, FOS-, or JUN-positive tumours. For those patients with positive lymph node involvement, the survival times were also shorter in the VEGF-positive, cyclin A-positive and FOS-positive groups. Multivariate analysis independently demonstrated a significant prognostic value for lymph node involvement, VEGF and FOS. PMID: 9484827 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 376: Oncogene. 1998 Feb 5;16(5):655-60. Apolipoprotein A-1 is a negative target of v-Jun overexpression. Hadman M, Lin W, Bush L, Bos TJ. Department of Microbiology and Immunology, Eastern Virginia Medical School, Norfolk 23501, USA. The product of the Jun oncogene influences a variety of processes including cell proliferation and differentiation. Jun exerts its influence by binding to the promoter and enhancer regions of a number of different target genes resulting in their activation or repression. We describe here the isolation and characterization of a gene differentially downregulated upon overexpression of v-Jun but not c-Jun. DNA and amino acid homology search analysis revealed this gene to be identical to chicken apolipoprotein A-1, the major component of high density lipoprotein (HDL). The half life of apolipoprotein A-1 RNA remains constant in the presence or absence of v-Jun overexpression suggesting downregulation by v-Jun is at the level of promoter activity. Consistent with this hypothesis, apolipoprotein A-1 upstream promoter fragments active in normal and c-Jun expressing CEF are inactive in v-Jun transformed CEF. Analysis of expression of apolipoprotein A-1 in CEF overexpressing other oncogenes revealed a similar downregulation by Myc and v-Src but not c-Fos, v-Ha-Ras, c-Src or c-Ski. Our findings point to a potential regulatory affect on cholesterol metabolism by v-Jun, as a result of altered levels of apolipoprotein A-1 message expression. PMID: 9482111 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 377: Chem Biol Interact. 1997 Dec 12;108(1-2):1-18. Erratum in: Chem Biol Interact 1998 Jul 3;114(1-2):143. Potential effect of sodium nitrite on the expression of nuclear proto-oncogenes during 2-acetyl aminofluorene-induced hepatocarcinogenesis in rats. Hsu JD, Hsu CL, Chou FP, Wen PH, Wang CJ. Department of Pathology, Chung Shan Medical and Dental College, Hospital, Tauchung, Taiwan, People's Republic of China. 2-acetyl aminofluorene (AAF) reacts in acidic conditions with nitrous fume yielding N-nitroso-AAF (N-NO-AAF), as previously described, that exerts more toxic and mutagenic effects than its parental compound. In this study, the effect of sodium nitrite (NaNO2) on the tumorigenicity of AAF in rats fed with AAF and NaNO2 was observed. Wistar rats were divided into five groups: group I served as control; group II were treated with NaNO2 (0.3%); group III was given 0.02% AAF alone; groups IV and V received both AAF and NaNO2 (0.2 and 0.3% respectively) in their diet for 12 weeks. At the end of the experiment, all rats in groups III, IV and V developed early stage phenomena of hepatocellular carcinoma, including hepatomegaly with variable-sized foci and neoplastic nodules. Severe damage was observed in the rats treated with AAF and NaNO2. Feeding of AAF (0.02%) for 3 months elevated the levels of c-Fos, c-Jun and c-Myc proteins in the rat livers. The AAF-induced c-Jun, c-Fos and c-Myc expressions were significantly magnified (P < 0.001) by NaNO2. These data confirmed that the strengthening of AAF-induced hepatocarcinogenesis by NaNO2 should be associated with its enhancing effect on the AAF-induced increases in the expressions of c-Jun, c-Fos and c-Myc. PMID: 9463517 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 378: Kidney Int. 1998 Feb;53(2):350-7. Morphine modulates proliferation of kidney fibroblasts. Singhal PC, Sharma P, Sanwal V, Prasad A, Kapasi A, Ranjan R, Franki N, Reddy K, Gibbons N. Department of Medicine, Long Island Jewish Medical Center, New Hyde Park, New York 11040, USA. Renal interstitial scarring is an important component of heroin-associated nephropathy. Kidney fibroblasts have been demonstrated to play a role in the development of renal scarring in a variety of renal diseases. We studied the effect of morphine, an active metabolite of heroin, on the proliferation of kidney fibroblasts. Morphine at a concentration of 10(-12) M enhanced (P < 0.001) the proliferation of kidney fibroblasts (control, 67.5 +/- 2.0 vs. morphine, 112.2 +/- 10.1 x 10(4) cells/well). [3H]thymidine incorporation studies further confirmed these results. Morphine at concentrations of 10(-12) M to 10(-10) M also modulated mRNA expression of early growth related genes (c-fos, c-jun and c-myc). Morphine at concentrations of 10(-8) to 10(-4) M promoted apoptosis of kidney fibroblasts and also enhanced the synthesis of p53 by kidney fibroblasts. We speculate that morphine-induced kidney fibroblast proliferation may be mediated through the activation of early growth related genes, whereas morphine induced kidney fibroblast apoptosis may be mediated through the generation of p53. The present in vitro study provides a hypothetical basis for the role of morphine in the development of renal interstitial scarring in patients with heroin-associated nephropathy. PMID: 9461094 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 379: Cancer Lett. 1997 Dec 9;120(2):235-41. Capsaicin can alter the expression of tumor forming-related genes which might be followed by induction of apoptosis of a Korean stomach cancer cell line, SNU-1. Kim JD, Kim JM, Pyo JO, Kim SY, Kim BS, Yu R, Han IS. Department of Biology, University of Ulsan, South Korea. Capsaicin (CAP) has been known to inhibit some tumor development in vivo (J.J. Jang, S.H. Kim, T.K. Yun, Inhibitory effect of capsaicin on mouse lung tumor development, in vivo, J. Korean Med. Sci. 3 (1989) 49-53; J.J. Jang, K.J. Cho, Y.S. Lee, J.H. Bae, Different modifying responses of capsaicin in a wide-spectrum initiation model of F344 rat, J. Korean Med. 6 (1991) 31-36) [1,2] even though its mechanism of action is not well understood. The objectives of this study were to examine the effect of CAP on expression of tumor forming-related genes in a Korean stomach tumor cell, SNU-1. We used slot blot hybridization to investigate its effect on a wide spectrum of proto-oncogenes. It was found that CAP enhanced the transcripts of two proto-oncogenes (c-myc and c-Ha-ras) and tumor suppressor gene p53. While a low concentration of CAP (0.01 microM) did not significantly increase the level of p53 transcript in SNU-1, it did increase it by a factor of 3.5 at a 10 microM dose of CAP. Consequently, SNU-1 cells are sensitive to CAP in the overexpression of tumor suppressor gene, p53 and proto-oncogenes, c-myc and c-Ha-ras, but not those of c-erbB-2, c-jun and bcl-2 genes. Both cell death and DNA fragmentation were shown in SNU-1 cells with treatment of CAP. Our results suggest that CAP induces apoptotic cell death in human gastric cancer cells (SNU-1) in vitro which may be possibly mediated by the overexpression of p53 and/or c-myc genes. Because cell suicide is arguably the most potent natural defense against cancer, the correlation between the induction of apoptosis and the change of tumor forming-related gene expression after CAP treatment should be further studied in detail. PMID: 9461043 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 380: Biochemistry (Mosc). 1997 Sep;62(9):1021-5. Expression of protooncogenes during lymphocyte activation by growth factors. Bulanova EG, Budagyan VM, Yarilin AA, Mazurenko NN. Institute of Immunology, Russian Ministry of Health Care, Moscow, Russia. Effects of growth factors of non-immune origin including somatotropin (ST) and platelet-derived growth factor (PDGF) on the expression of the proteins encoded by c-fos, c-myc, c-fun, and c-ets family protooncogenes were studied for the first time. The dynamics of the oncoprotein expression in activated CD(3+)-lymphocytes was investigated by immunoblotting. The accumulation of the Fos and Myc proteins was enhanced in T-lymphocytes treated with ST, PDGF, or phytohemagglutinin; the accumulation was maximum at 30-60 min and decreased in 2 h; the data indicate that the oncoproteins participate in the early lymphocyte activation by various growth factors. The Jun protein appears only in 3 h after the onset of lymphocyte activation; this suggests independent participation of Fos in the early stages of lymphocyte activation prior to the appearance of Jun, preceding the joint action of Fos and Jun within the AP-1 transcription complex. The products of the c-ets family are differentially activated by the studied growth factors. Resting lymphocytes actively accumulate the Ets-1 protein; ST and PDGF activation decreases Ets-1 expression in 2 h. The Ets-2 protein is not detected in resting cells and PDGF-activated lymphocytes, whereas lymphocyte activation by ST is associated with accumulation of Ets-2. The data suggest that the product of the c-ets-1 gene is more important in the regulation of resting cells and the product of the c-ets-2 gene is important during activation of lymphocytes by ST. The results indicate that activation of lymphocytes with growth factors of non-immune origin is mediated by several signal transduction pathways. PMID: 9457764 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 381: J Biochem Mol Toxicol. 1998;12(2):79-82. Immediate-early gene expression during regenerative and mitogen-induced liver growth in the rat. Holden PR, Odum J, Soames AR, Foster JR, Elcombe CR, Tugwood JD. Zeneca Central Toxicology Laboratory, Alderley Park, Macclesfield, England, UK. The nongenotoxic carcinogens phenobarbitone (PB) and methyl clofenapate (MCP) and the hepatomitogen pregnenolone 16 alpha carbonitrile (PCN) are direct inducers of hepatic S-phase in rats, whereas the S-phase seen after partial hepatectomy is regenerative. We have investigated S-phase and immediate-early gene expression (c-myc and c-jun) in rat liver following these treatments to study the differences in gene expression associated with direct vs. regenerative responses. Both partial hepatectomy (one- and two-thirds) and mitogen treatment caused an increase in hepatic S-phase that peaked around 36 hours. Two-thirds partial hepatectomy caused the greatest increase in S-phase followed by one-third partial hepatectomy, then the mitogens PCN, MCP, and PB in that order. This order of response was also seen with c-jun and to a lesser degree with c-myc expression, suggesting that immediate-early gene expression might be linked not only to regenerative S-phase but also to direct mitogen-induced responses. PMID: 9443064 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 382: Jpn J Cancer Res. 1997 Nov;88(11):1052-62. Inhibition of development of N,N'-dimethylhydrazine-induced rat colonic aberrant crypt foci by pre, post and simultaneous treatments with 24R,25-dihydroxyvitamin D3. Salim EI, Wanibuchi H, Taniyama T, Yano Y, Morimura K, Yamamoto S, Otani S, Nishizawa Y, Morii H, Fukushima S. First Department of Pathology, Osaka City University Medical School. It has recently been reported that new vitamin D3 derivatives can exert inhibitory effects on colon carcinogenesis in rats. In the present study the chemopreventive potential of 24R,25-dihydroxyvitamin D3 (24R,25(OH)2vitamin D3) was assessed in a murine model of colon carcinogenesis. In experiment 1, male 6-week-old F344 rats were administered N,N'-dimethylhydrazine (DMH) 20 mg/kg s.c. once a week 4 times. The rats were fed 24R,25(OH)2vitamin D3 at 10 ppm in the diet prior to (pre), together with (simultaneous) or after (post) DMH treatment. Modifying effects were assessed using aberrant crypt foci (ACF), putative preneoplastic lesions, as the end point markers in this model of colon carcinogenesis. After 8 weeks, pre and more markedly simultaneous administration of 24R,25(OH)2vitamin D3 was found to have reduced the total numbers of ACF and significantly inhibited the development of foci. After 16 weeks, numbers of foci with > or = 4 crypts, which are more likely to progress to tumors, were significantly reduced. The most pronounced inhibition of ACF development was noted in rats fed the 24R,25(OH)2vitamin D3 after DMH administration. The reduction was particularly marked in the proximal colon. Blood levels of calcium were not significantly increased over the control levels in groups administered DMH and the vitamin. Immunohistochemical staining showed numbers of proliferating cell nuclear antigen-positive cells to be lower in the colonic epithelia of rats fed the vitamin D3 metabolite than in the controls. In experiment 2, the effect of 24R,25(OH)2vitamin D3 on the alterations in c-fos, c-myc and c-jun oncogene expression in response to DMH administration was examined by northern blot analysis. The early increase in expression of ornithine decarboxylase (ODC) activity was not altered by 24R,25(OH)2vitamin D3. The results suggest that 24R,25(OH)2vitamin D3 is a cancer chemopreventive agent which may suppresses DMH induction of lesions and their subsequent development via an antiproliferative action. PMID: 9439680 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 383: Blood. 1998 Jan 15;91(2):406-18. The SH3 domain contributes to BCR/ABL-dependent leukemogenesis in vivo: role in adhesion, invasion, and homing. Skorski T, Nieborowska-Skorska M, Wlodarski P, Wasik M, Trotta R, Kanakaraj P, Salomoni P, Antonyak M, Martinez R, Majewski M, Wong A, Perussia B, Calabretta B. Kimmel Cancer Institute, Jefferson Medical College, Philadelphia, PA 19107, USA. To determine the possible role of the BCR/ABL oncoprotein SH3 domain in BCR/ABL-dependent leukemogenesis, we studied the biologic properties of a BCR/ABL SH3 deletion mutant (delta SH3 BCR/ABL) constitutively expressed in murine hematopoietic cells. delta SH3 BCR/ABL was able to activate known BCR/ABL-dependent downstream effector molecules such as RAS, PI-3kinase, MAPK, JNK, MYC, JUN, STATs, and BCL-2. Moreover, expression of delta SH3 BCR/ABL protected 32Dcl3 murine myeloid precursor cells from apoptosis, induced their growth factor-independent proliferation, and resulted in transformation of primary bone marrow cells in vitro. Unexpectedly, leukemic growth from cells expressing delta SH3 BCR/ABL was significantly retarded in SCID mice compared with that of cells expressing the wild-type protein. In vitro and in vivo studies to determine the adhesive and invasive properties of delta SH3 BCR/ABL-expressing cells showed their decreased interaction to collagen IV- and laminin-coated plates and their reduced capacity to invade the stroma and to seed the bone marrow and spleen. The decreased interaction with collagen type IV and laminin was consistent with a reduced expression of alpha 2 integrin by delta SH3 BCR/ABL-transfected 32Dcl3 cells. Moreover, as compared with wild-type BCR/ABL, which localizes primarily in the cytoskeletal/membrane fraction, delta SH3 BCR/ABL was more evenly distributed between the cytoskeleton/membrane and the cytosol compartments. Together, the data indicate that the SH3 domain of BCR/ABL is dispensable for in vitro transformation of hematopoietic cells but is essential for full leukemogenic potential in vivo. PMID: 9427693 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 384: J Exp Ther Oncol. 1996 Nov;1(6):335-41. The association of the H-ras oncogene and steroid hormone receptors in gynecological cancer. Zachos G, Varras M, Koffa M, Ergazaki M, Spandidos DA. Institute of Biological Research and Biotechnology, National Hellenic Research Foundation, Athens, Greece. Expression of the ras family of cellular oncogenes is associated with tumorigenicity, invasiveness and metastatic potential in a variety of human carcinomas. Additionally, H-ras cooperates with glucocorticoids and with ovarian hormones in cell transformation and in the development of mammary carcinomas. Steroids are considered to be tumor promoters and their levels influence the cure rates and survival of the patients with gynecological lesions. It is proposed that they exert tumor promoting activity by transcriptional regulation of nuclear proto-oncogenes, such as c-fos, c-jun, and c-myc. The human H-ras gene contains within its first and fourth introns, sequences that are specifically recognized by glucocorticoid and estrogen receptors, respectively. Using gel retardation assays, the level of steroid receptor binding in H-ras elements has been compared, employing nuclear extracts from human endometrial and ovarian lesions and from the adjacent normal tissue. Elevated binding of the glucocorticoid and estrogen receptors in the corresponding H-ras elements in almost all tissue pairs tested has been found. It is suggested that the H-ras proto-oncogene is hormonally regulated and directly implicated in human gynecological cancer through elevated, steroid-induced gene expression. Publication Types: Review Review, Tutorial PMID: 9414422 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 385: Science. 1997 Dec 5;278(5344):1812-5. Requirement of NF-kappaB activation to suppress p53-independent apoptosis induced by oncogenic Ras. Mayo MW, Wang CY, Cogswell PC, Rogers-Graham KS, Lowe SW, Der CJ, Baldwin AS Jr. Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA. The ras proto-oncogene is frequently mutated in human tumors and functions to chronically stimulate signal transduction cascades resulting in the synthesis or activation of specific transcription factors, including Ets, c-Myc, c-Jun, and nuclear factor kappa B (NF-kappaB). These Ras-responsive transcription factors are required for transformation, but the mechanisms by which these proteins facilitate oncogenesis have not been fully established. Oncogenic Ras was shown to initiate a p53-independent apoptotic response that was suppressed through the activation of NF-kappaB. These results provide an explanation for the requirement of NF-kappaB for Ras-mediated oncogenesis and provide evidence that Ras-transformed cells are susceptible to apoptosis even if they do not express the p53 tumor-suppressor gene product. PMID: 9388187 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 386: Nord Med. 1997 Oct;112(8):271-5. [Apoptosis: molecular aspects] [Article in Danish] Lovschall H, Kassem M, Mosekilde L. Afd for Tandsygdomslaere, Odontologisk Institut, Aarhus Universitet. Many signals and external stimuli regulate the apoptosis activity by interaction with the genome. These stimuli include morphogenetic signals, physiological factors, and environmental influence. The signals mediate their effect on cells with suitable receptors, relevant signalling pathways, and competence to execute the apoptosis cascade. Apoptosis is triggered indirectly by deprivation of survival factors, or directly by intercellular cell death signalling factors, and also by unbalanced intracellular messenger molecules, which are, more or less, involved in regulation of both programmed cell death and survival. Several genes are involved in regulation of cell survival and apoptosis: bcl-2/bax, p53, c-myc and transcription factors such as cdk, c-myc, c-fos and c-jun. Apparently, apoptosis could be triggered by increased or inhibited gene expression as well as biochemical reactions without changed gene expression. The morphological changes during apoptosis reflect a cascade of genetic and biochemical reactions in the cell. In the signal transduction pathway both secondary messenger Ca2+, different kinases, and polyamines are involved. Cysteine proteases cleave cytoskeletal proteins, endonucleases divide DNA into fragments, and transglutaminases cross-link macromolecules. Degradative enzymes such as proteases, endonucleases and transglutaminases are activated during apoptosis, leading to cellular collapse and formation of vesicular apoptotic bodies. Both increased and inhibited apoptosis activity may have pathological consequences. New therapeutic strategies aim to counteract dysregulation of apoptosis in specific tissues by pharmacological intervention. Thus there is a need for identification of molecules and gene products involved in regulation of apoptosis activity and clarification of the conditions where this knowledge may be used. PMID: 9411394 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 387: J Biol Chem. 1997 Nov 28;272(48):30455-62. Fluid shear stress activation of focal adhesion kinase. Linking to mitogen-activated protein kinases. Li S, Kim M, Hu YL, Jalali S, Schlaepfer DD, Hunter T, Chien S, Shyy JY. Department of Bioengineering and Institute for Biomedical Engineering, University of California, San Diego, La Jolla, California 92093, USA. shyy@bioeng.ucsd.edu Shear stress, the tangential component of hemodynamic forces, activates the extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) signal transduction pathways in cultured vascular endothelial cells to induce the transcriptional activation of many immediate early genes. It appears that integrins, protein-tyrosine kinases, and the structural integrity of actin are important factors involved in these shear stress-induced responses. The underlying molecular events were investigated by the application of a shear stress of 12 dyn/cm2 on bovine aortic endothelial cells (BAEC). We found that such a shear stress increased the tyrosine phosphorylation and the kinase activity of focal adhesion kinase (FAK) and its association with growth factor receptor binding protein 2 (Grb2) in a rapid and transient manner, suggesting that FAK may be linked to these mitogen-activated protein kinase signaling pathways through a Grb2.Son of sevenless (Sos) complex. FAK(F397Y), which encodes a dominant negative mutant of FAK, attenuated the shear stress-induced kinase activity of Myc epitope-tagged ERK2 and hemagglutinin epitope-tagged JNK1. DeltamSos1, encoding a dominant negative mutant of Sos in which the guanine nucleotide exchange domain has been deleted, also attenuated shear stress activation of Myc-ERK2 and hemagglutinin-JNK1. Pretreating the confluent BAEC monolayers with a blocking type anti-vitronectin receptor monoclonal antibody had similar inhibitory effects in these shear stress-activated ERKs and JNKs. Confocal microscopic observation further demonstrated that FAK tended to cluster with vitronectin receptor near the abluminal side of the sheared BAEC. These results demonstrate that FAK signaling is critical in the shear stress-induced dual activation of ERK and JNK. PMID: 9374537 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 388: Fundam Appl Toxicol. 1997 Nov;40(1):129-37. Expression of the immediate-early genes, c-fos, c-jun, and c-myc: a comparison in rats of nongenotoxic hepatocarcinogens with noncarcinogenic liver mitogens. Hasmall SC, Pyrah IT, Soames AR, Roberts RA. Zeneca Central Toxicology Laboratory, Macclesfield, Cheshire, United Kingdom. The involvement of the immediate-early (IE) genes c-fos, c-jun, and c-myc in regenerative liver hyperplasia is accepted, but their involvement in direct hyperplasia is uncertain. We have examined the hypothesis that the ability to induce IE genes may reflect the hepatocarcinogenic potential of a chemical. The ability of 1,4-dichlorobenzene (DCB) (300 mg/kg) (a noncarcinogenic rat liver mitogen), diethylhexyl phthalate (DEHP) (950 mg/kg), and chlorendic acid (120 mg/kg) (both nongenotoxic hepatocarcinogens) to induce c-fos, c-jun, and c-myc expression in rat liver was determined by Northern blot analysis and by in situ hybridization. Results were correlated to hepatic labeling index (LI) as determined by incorporation of BrdU in each of three lobes for each of three male F344 rats per group. Carbon tetrachloride (CCl4) (2 ml/kg) was used as a positive control. Increased LI was preceded by elevated expression of all three IE genes after CCl4, but also after DCB and DEHP, although induction by these was less marked. In all cases, there was considerable interanimal variation within groups, but little interlobe variation. Interestingly, there was a good correlation (r2 > or = 0.85) between c-myc expression and LI, but not between LI and c-fos or c-jun. Despite the disparate carcinogenic potential of DEHP and DCB, both chemicals induced similar patterns of IE gene expression, suggesting that this cannot distinguish hepatocarcinogenic liver mitogens from noncarcinogenic liver mitogens. These data assist in the evaluation of IE gene expression both as a marker of direct versus regenerative hyperplasia and as an indicator of the hepatocarcinogenic potential of liver mitogens. PMID: 9398495 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 389: Hua Xi Yi Ke Da Xue Xue Bao. 1996 Jun;27(2):117-21. [Transcriptional expression of oncogenes and Rb antioncogene in experimental atherosclerotic lesions] [Article in Chinese] Wang H, Liu B, Fu M, Zeng C. Apolipoprotein Research Unit, Institute of Biochemistry and Molecular Biology, Chengdu. Atherosclerosis (AS) is characterized by the proliferation of the smooth muscle cells (SMC) in the arterial wall. Its pathogenesis might be associated with overexpression of oncogenes in SMC. Gorden and Barrett et al found that sis mRNA level elevated in human atherosclerotic plaques 5-12 fold above level present in normal artery. But the transcriptional expression of c-fos, c-myc, c-jun, H-ras, v-erb-B oncogenes and Rb antioncogene in atherosclerotic lesion has not yet been reported. A study on these oncogenes and Rb gene expression in artherosclerotic lesions in rabbits fed on high cholesterol diet were assayed by the dot blot hybridization using alpha-32P-labelled oncogenes and Rb gene fragments as the probes. After fed with the high cholesterol diet for six months, the plasma cholesterol levels in AS rabbits were significantly increased (1300 +/- 240 mg/dl vs 67.1 +/- 11.5 mg/dl). The atherosclerotic plaques covered 91% +/- 11% of the intimal aortic surface of aorta thoracalis. The results showed that the atherosclerotic plaques contained 3-4 fold more v-sis, c-fos and c-myc mRNA (P < 0.01), 2 fold more c-jun and H-ras mRNA (P < 0.01), and less Rb mRNA (P < 0.05) than those in the normal aortic arteries. But the expression of v-erb-B gene in atherosclerotic plaques remained unchanged. These results indicate that the abnormal expression of v-sis, c-myc, c-fos, c-jun, and H-ras oncogenes and Rb antioncogene may play an important role in arterial SMC proliferation and pathogenesis of atherosclerosis. PMID: 9389022 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 390: FEBS Lett. 1997 Oct 13;416(1):113-6. Mechanisms of cycloheximide-induced apoptosis in liver cells. Alessenko AV, Boikov PYa, Filippova GN, Khrenov AV, Loginov AS, Makarieva ED. Institute of Biochemical Physics RAS, Moscow, Russia. aless@center.chph.ras.ru Cycloheximide in sublethal doses caused apoptosis in liver cells in vivo, inducing c-myc, c-fos, c-jun and p53 genes and accumulation of sphingosine, a toxic product of the sphingomyelin cycle. These data support the hypothesis that continuous synthesis of labile protective proteins is required to restrain apoptosis in liver; sphingosine might be important in mediating cycloheximide-induced apoptosis as an endogenous modulator of protein kinase C activity. PMID: 9369245 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 391: Cancer. 1997 Oct 15;80(8 Suppl):1581-7. Mechanisms of the development of osteoblastic metastases. Goltzman D. Department of Medicine, Royal Victoria Hospital and McGill University, Montreal, Quebec, Canada. Although several neoplasms may produce osteoblastic metastases, carcinoma of the prostate is by far the most common. Biochemical and histologic studies indicate that osteolysis also is a manifestation of prostate carcinoma. Furthermore, factors such as parathyroid hormone-related peptide, which mediate osteolysis in other cancers, also appear to be operative in the bone breakdown induced by prostate carcinoma. However, the most unique skeletal effect of this tumor is its consistent capacity to stimulate osteoblasts to deposit new bone. Several bone growth factors have been detected in prostatic tissue and may contribute to this process. These include transforming growth factor-beta, fibroblast growth factor, and bone morphogenetic proteins. The author isolated an amino-terminal fragment (ATF) of the protease urokinase (uPA) from the conditioned medium of the prostate carcinoma cell line PC-3 and demonstrated that this fragment has mitogenic activity for osteoblastic cells. The activity appears to reside in an epidermal growth factor-like growth factor domain (GFD) within the ATF. Subsequently, the author cloned the rat uPA receptor (uPAR). uPAR is known to bind the ATF and can permit the uPA molecule to exhibit focal proteolysis. It was shown that the ATF also can induce c-myc, c-jun, and c-fos in osteoblastic cells. This effect of ATF can be mimicked by the GFD and suggests that this signalling pathway in osteoblasts is via the uPAR. Consequently, the uPA molecule may contribute to growth factor effects in osteoblasts via the NH2-terminal fragment and to tumor invasiveness via its COOH-terminal proteolytic domain. This scenario is supported by results from studies with uPA-overexpressing prostate carcinoma cells in rats. Additional studies will be required to further define the mechanisms of interaction of prostate carcinoma and other cancers with bone but each site of molecular interaction may provide a therapeutic window for curtailing the effects of these tumors on the skeleton. Publication Types: Review Review, Tutorial PMID: 9362425 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 392: Gen Comp Endocrinol. 1997 Nov;108(2):173-81. Proto-oncogene activity in the testis of the lizard, Podarcis s. sicula, during the annual reproductive cycle. Chieffi P, Angelini F, Pierantoni R. Dipartimento di Fisiologia Umana e Funzioni Biologiche Integrate "Filippo Bottazzi," II, Universita di Napoli, Naples, 80138, Italy. Since proto-oncogenes play a central role in the regulation of cellular growth and differentiation, localization of MYC, FOS, and JUN proteins has been studied in the testis of the lizard, Podarcis s. sicula, during the annual reproductive cycle by immunocytochemistry using antisera against c-myc, c-fos, and c-jun products. MYC was localized in the nuclei of spermatogonia (SPG), I and II spermatocytes (SPC), and spermatids (SPT). Strong immunoreactivity was detected in Sertoli cells just prior to the onset of the early spring spermatogenic wave coinciding with the androgen peak. FOS protein was present in the nuclei of SPG and SPC. In SPG an exclusive nuclear localization was seen during the active spermatogenic period (February-March and September). A perinuclear localization was observed during other months. Immunoreactivity in Sertoli cells was also observed during the periods of active spermatogenesis. JUN protein was localized in the cytoplasm of SPG as well as in I and II SPC and was detected in the nuclei of I and II SPC during April and October when spermatogenic waves occur. These data suggest that proto-oncogene activities have regulatory roles in the spermatogenesis of the lizard. Copyright 1997 Academic Press. Copyright 1997 Academic Press PMID: 9356213 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 393: Br J Urol. 1997 Oct;80(4):548-53. Early genetic and cellular responses in the smooth muscle layer of obstructed ureters in a rat model of obstructive uropathy. Chuang YH, Chuang WL, Liu KM, Chen SS, Huang CH. Department of Neurology, School of Medicine, Kaohsiung Medical College, Taiwan, ROC. OBJECTIVE: To investigate the early genetic and cellular responses in the smooth muscle layer of completely obstructed ureters, and to determine whether myocytes proliferate (hyperplasia) in the ureters during the early stage of obstructive uropathy. MATERIALS AND METHODS: The study comprised 35 female Sprague-Dawley rats which had undergone unilateral ligation of their ureters. After ureteric ligation, five rats each were killed and examined at 0.5, 1, 2, 4, 8, 16 and 24 h after ligation. The proximal portion of the ureters was prepared for light and electron microscopy. The expression of c-Fos, c-Jun, c-Myc and Ki-67 antigen was assessed immunohistochemically. RESULTS: c-Fos and c-Jun were detected 2 h after ligation and the expression of these two proteins reached a maximum after 4 h, becoming undetectable 16 h after ligation. The expressions of c-Fos and c-Jun were strongly correlated (r = 0.9854, P < 0.001). The expressions were of c-Myc and Ki-67 antigen was not detected within 24 h after ureteric ligation. The amount of rough endoplasmic reticulum (rER) in the ligated ureters increased soon after complete ligation and the increase continued throughout the period of ureteric obstruction (r = 0.9699, P < 0.001). The change in rER was also significantly correlated with the expression of c-Fos and c-Jun within 8 h after ligation. CONCLUSION: The expression of c-Fos and c-Jun, but not c-Myc, might contribute to the hypertrophy of the ureteric smooth muscle layer during the course of complete ureteric obstruction. There is no hyperplasia in ureteric smooth muscle in the early stages of obstructive uropathy. PMID: 9352690 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 394: Toxicol Pathol. 1997 Sep-Oct;25(5):433-40. Dissimilar characteristics of N-methyl-N-nitrosourea-initiated foci and tumors promoted by dichloroacetic acid or trichloroacetic acid in the liver of female B6C3F1 mice. Latendresse JR, Pereira MA. ManTech Environmental Technology, Toxicology Division, Armstrong Laboratory, Wright-Patterson Air Force Base, Ohio 45433, USA. Dichloroacetic acid (DCA) and trichloroacetic acid (TCA) are metabolites of the industrial solvent and environmental contaminant trichloroethylene (TCE), as well as contaminants of chlorinated drinking water. Human exposure to these chemicals is of concern as all three have been shown to increase liver tumor incidence in mice. Differences in dose-response curves, progression to cancer, and postexposure regression of lesions suggest that TCA and DCA work through different mechanisms. The purpose of this study was to further characterize the proliferative hepatocellular lesions promoted by TCA and DCA using biomarkers of cell growth, differentiation, and metabolism in liver sections to better delineate the distinctions in the mechanism of the two chloroacetates. Fifteen-day-old female mice were initiated with 25 mg/kg N-methyl-N-nitrosourea. The initiated mice were administered DCA or TCA (20.0 mmol/L) in drinking water from age 49 days until euthanasia at age 413 days. The pathologic assessment showed that the foci of altered hepatocytes and tumors occurring in the animals promoted with DCA were eosinophilic and positive immunohistochemically for TGF-alpha, c-jun, c-myc, CYP 2E1, CYP 4A1, and glutathione S-transferase-pi (GST-pi). The DCA lesions also were essentially negative for c-fos and TGF-beta, but nontumor hepatocytes were consistently TGF-beta-positive. In contrast, tumors promoted by TCA were predominantly basophilic, lacked GST-pi, and stained variably; usually, more than 50% of the tumor hepatocytes were essentially negative for the other biomarkers. This study demonstrates some striking differences in certain molecular biomarkers of cell growth, differentiation, and metabolism between DCA and TCA. The results also suggest some potential growth signal transduction pathways that may contribute to the DCA promotion of tumors, further support the premise that these two chloroacetates promote hepatocarcinogenesis in different ways, and provide a rational basis for a similar comparison with TCE. Such a comparison should give some insight as to whether DCA, TCA, or both are playing a significant role in the murine liver carcinogenesis of the parent compound, TCE. PMID: 9323830 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 395: Nephron. 1997;77(2):225-34. Effect of morphine on renomedullary interstitial cell proliferation and matrix accumulation. Singhal PC, Sharma P, Gibbons N, Franki N, Kapasi A, Wagner JD. Department of Medicine, Long Island Jewish Medical Center, New Hyde Park, N.Y. 11040, USA. Renal interstitial scarring is an important feature of heroin-associated nephropathy. We studied the effect of morphine, an active metabolite of heroin, on cultured rat renal medullary interstitial cell (RMIC) proliferation and matrix accumulation. Morphine (10(-12) M) enhanced (p < 0.001) the proliferation of RMIC (control, 15.0+/-0.5 vs. morphine, 20.4+/-1.1 x 10(4) cells/ml). This effect of morphine was dose and time dependent. [3H]thymidine and bromodeoxyuridine incorporation studies confirmed the mitogenic effect of morphine on RMIC. Morphine also enhanced mRNA expression for c-jun and c-myc on RMIC. However, nalbuphine, a non-addicting alkaloid did not modulate the proliferation of RMIC. Morphine enhanced the accumulation of collagen type I in a dose-dependent manner and also increased (p < 0.001) the accumulation of collagen type III at a high concentration (control, 1,291+/-55.8 vs. morphine, 10(-4) M, 2,697.6+/-257.8 ng/microg protein). Morphine did not modulate the accumulation of laminin or fibronectin. Neutralizing antibody to IL-6 inhibited the effect of morphine on RMIC. H7, a protein kinase C inhibitor, also attenuated the morphine-induced RMIC proliferation. The present study provides a basis for a hypothesis that morphine may be playing a role in the development of renal interstitial pathology in patients with heroin addiction. PMID: 9346391 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 396: J Biol Chem. 1997 Oct 24;272(43):27015-24. Gastrin and phorbol 12-myristate 13-acetate regulate the human histidine decarboxylase promoter through Raf-dependent activation of extracellular signal-regulated kinase-related signaling pathways in gastric cancer cells. Hocker M, Henihan RJ, Rosewicz S, Riecken EO, Zhang Z, Koh TJ, Wang TC. Gastrointestinal Unit and Department of Medicine, Massachusetts General Hospital, Boston, Massachusetts 02114, USA. Gastrin stimulates transcription of the human histidine decarboxylase (HDC) gene through binding to the G-protein-coupled cholecystokinin-B/gastrin receptor. We have explored the possibility that mitogen-activated protein kinase cascades play a role in mediating the effects of gastrin on transcription in a gastric cancer (AGS-B) cell line. Gastrin and phorbol 12-myristate 13-acetate (PMA) treatment of AGS-B cells was found to increase the phosphorylation of tyrosine residues of extracellular signal-regulated kinases (ERKs) 1 and 2 and increase ERK activity as determined by the in vitro phosphorylation of myelin basic protein. Reporter gene assays also demonstrated that gastrin and PMA stimulated Elk-1- and c-Myc-dependent transactivation, consistent with gastrin- and PMA-induced activation of ERKs. Overexpression of wild type ERK-1 and ERK-2 or activation of endogenous ERKs using activated MEK-1 (mitogen-activated protein kinase kinase or ERK kinase) overexpression stimulated HDC promoter activity in a dose-dependent fashion. Interruption of the ERK-related pathway using expression vectors for kinase-deficient ERKs or an ERK-specific phosphatase (PAC-1) blocked gastrin- and PMA-stimulated HDC promoter activity. In contrast, inhibition of the Jun kinase pathway using an interfering dominant negative SEK-1 (stress-activated protein kinase/ERK-1) mutant did not inhibit HDC promoter activity. Furthermore, whereas gastrin stimulated phosphorylation of Shc proteins and association with Grb2, activation of the HDC promoter was not influenced by expression of dominant negative Ras (N15 or N17) proteins. However, gastrin stimulated Raf-1 kinase activity, and activation of the HDC promoter was blocked by coexpression of a dominant negative Raf-1 construct. Overall, these data demonstrate that gastrin regulates HDC transcription in a Rafdependent, Ras-independent fashion predominantly through activation of the ERK-related pathway. PMID: 9341140 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 397: Mol Cells. 1997 Aug 31;7(4):537-43. Characterization of the murine cyclin D2 gene: exon/intron organization and promoter activity. Jun DY, Kim MK, Kim IG, Kim YH. Department of Microbiology, College of Natural Sciences, Kyungpook National University, Taegu, Korea. Cyclin D2 is normally expressed in G1 and promotes progression through G1 of the cell cycle. From a murine genomic library constructed with spleen DNA, two overlapping genomic clones of cyclin D2 were isolated. These clones contain most of the exon of cyclin D2 except exon 5. Characterization of these clones revealed that murine cyclin D2 mRNA spans over 18 kb and 5 exons ranging from 149 to approximately 462 bp in length, and suggested that exon 5 may be at least >5 kb downstream from exon 4. Primer extension analysis of cyclin D2 mRNA isolated from murine activated T cells detected 5 putative sites of transcription initiation. These are located at - 499, - 417, - 391, - 373, and - 349 relative to the translation start site, which is given as + 1. No consensus sequence for TATA box existed at an appropriate position within the promotor region. Instead, several putative transcriptional factor binding sites for C/EBP, PEA3, AP2, NF-Y, Sp1, c-Myc, GATA-1, AP1, v-Myb, and CREB were detected. The 5'-flanking region of the cyclin D2 gene up to nucleotide - 945 shared about 61% sequence homology between mouse and human. Functional analysis of promoter activity of the 5'-flanking region of cyclin D2 suggested that the region - 1,100 to - 805 including C/EBP, PEA3, AP2, NF-Y, c-Myc, and Sp1 may have a major positive regulatory activity for expression of cyclin D2. PMID: 9339900 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 398: Cell Biochem Funct. 1997 Sep;15(3):153-62. Characterization and partial purification of a keratinocyte-derived growth factor with wound-healing properties. Kratz G, Jonzon B, Hultgard-Nilsson A, Haegerstrand A. Department of Plastic and Reconstructive Surgery, Karolinska Hospital, Stockholm, Sweden. We have previously described the mitogenic and wound-healing properties of keratinocyte-conditioned medium (KCM). In this study we investigated the effect of KCM on the activation of second messenger systems and the expression of proto-oncogene in cultured human skin fibroblasts. We also present a partial purification of the factor responsible for the mitogenic and wound-healing effects of KCM. KCM was shown to increase the expression of the proto-oncogenes c-fos, c-myc and c-jun. The effect of KCM on three second messenger systems was investigated. The extracellular release of choline metabolites was increased by 40 per cent when cells were stimulated with KCM whereas the formation of cAMP and hydrolysis of phosphatidyl inositol (PI) was unaffected. KCM was purified by ion exchange chromatography and filtration. The biologically active fraction was eluted from an SP column and retained its activity after filtration through a 3-kDa filter. The fraction was inactivated by heat and acid, indicative of a peptide origin. Furthermore, the active fraction was shown to increase the extracellular release of choline metabolites and to stimulate re-epithelialization in wounds in human skin in vitro comparable to KCM. The study indicates that human keratinocytes produce a < 3 kDa peptide which may be partly responsible for the growth stimulatory and wound-healing properties of KCM. PMID: 9377793 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 399: FASEB J. 1997 Oct;11(12):965-72. The activated anaplastic lymphoma kinase increases cellular proliferation and oncogene up-regulation in rat 1a fibroblasts. Wellmann A, Doseeva V, Butscher W, Raffeld M, Fukushima P, Stetler-Stevenson M, Gardner K. Laboratory of Pathology, National Institutes of Health, Bethesda, Maryland 20892-1500, USA. More than 60% of anaplastic large-cell lymphomas (Ki-1 lymphoma) are associated with a t(2;5)(p23;q35) translocation that produces an 80 kDa hyperphosphorylated chimeric protein (p80) derived from the fusion of the anaplastic lymphoma kinase (ALK) with nucleophosmin (NPM). The NPM-ALK chimeric gene is an activated tyrosine kinase that has been shown to be a potent oncogene. We have developed a cellular model for the study of p80 action in rat 1a fibroblasts. Expression of cDNA's encoding NPM-ALK (p80) in rat 1a fibroblasts induces anchorage-independent growth in soft agar and promotes foci formation in culture. Cells expressing exogenous p80 showed significantly increased proliferation characterized by accelerated cell cycle entry into S-phase. Consistent with increased G0/G1 to S-phase transition, there is also marked up-regulation of cyclin A and cyclin D1 expression. In addition, p80 transformed cells showed elevated expression of several immediate early genes involved in cellular proliferation, including fos, jun, and c-myc. DNA binding analysis of nuclear extracts prepared from p80 transformed cells reveal marked up-regulation of AP-1 DNA binding activity. Functional AP-1-specific transfection assays also show up-regulation of AP-1-dependent transcriptional activation. These finding demonstrate that p80 transformed rat 1a fibroblast can be a highly useful model system for the molecular and biochemical characterization of the mechanisms of action of this interesting new oncogene. PMID: 9337149 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 400: Am J Physiol. 1997 Sep;273(3 Pt 1):C1020-9. Polyamines modulate transcription but not posttranscription of c-myc and c-jun in IEC-6 cells. Patel AR, Wang JY. Department of Surgery, University of Maryland Medical School, Baltimore, USA. The goal of the current study was to examine whether polyamines are involved in the regulation of transcription and posttranscription of the protooncogenes c-myc and c-jun in intestinal epithelial cells. Studies were conducted in the IEC-6 cell line derived from rat small intestinal crypt cells. Administration of alpha-difluoromethylornithine (DFMO), a specific inhibitor of polyamine synthesis, for 4 or 6 days not only almost completely depleted total (whole) cellular and nuclear polyamines but also significantly decreased expression of the protooncogenes c-myc and c-jun in IEC-6 cells. Using nuclear run-on transcription assay, we demonstrated that the basal rate of transcription of c-myc was decreased by 55% at 4 days and by 60% at 6 days in the DFMO-treated cells. The c-jun transcription in DFMO-treated cells was decreased by 75% at 4 days and 85% at 6 days. The transcription rates of c-myc and c-jun were dramatically stimulated by 5% dialyzed fetal bovine serum (dFBS) in normal quiescent cells. However, polyamine depletion significantly prevented the increased transcription of these two genes in the DFMO-treated cells exposed to 5% dFBS. Furthermore, direct administration of spermidine to isolated nuclei from polyamine-deficient (caused by DFMO) cells resulted in a 2- to 2.5-fold increase in c-myc and c-jun transcription. There were no significant changes in the half-lives of c-myc and c-jun mRNAs between the controls and the DFMO-treated cells. These results indicate that 1) polyamines are required for the transcription of the protooncogenes c-myc and c-jun in IEC-6 cells and 2) depletion of intracellular polyamines has no effect on posttranscriptional regulation of c-myc and c-jun mRNAs. These findings suggest that polyamines play an important role in the regulation of the transcription of protooncogenes, and this may be one mechanism by which polyamines modulate mucosal cell division. PMID: 9316423 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 401: Cancer Lett. 1997 Sep 16;118(1):101-7. Alteration of proto-oncogenes during apoptosis in the oral squamous cell carcinoma cell line, SAS, induced by staurosporine. Abiko Y, Arai J, Mitamura J, Kaku T. Department of Oral Pathology, School of Dentistry, Health Sciences University of Hokkaido, Japan. Staurosporine (ST) has been reported to induce apoptosis in many kinds of cultured cells. The pathway of the apoptosis induced by ST is still not clear. Certain proto-oncogene expressions have been shown to be involved in the apoptotic pathway. The present study characterized apoptosis induced by ST in the oral squamous cell carcinoma cell line, SAS, focusing on the alteration of proto-oncogene expression. SAS showed typical apoptotic features upon exposure to ST. We compared the level of gene expression in apoptosis induced by ST with that by withdrawal of serum, which is a common system to induce apoptosis. By RT-PCR analysis, ST-induced apoptosis showed c-fos and c-jun up-regulation, whereas serum withdrawal-induced apoptosis showed c-jun up-regulation and the same levels of p21/waf-1 and c-myc. These results indicate that ST can rapidly induce apoptosis in SAS, possibly via a c-fos and c-jun pathway. PMID: 9310266 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 402: Leuk Res. 1997 Jul;21(7):589-94. Differential effects of phorbol ester on signaling and gene expression in human leukemia cells. Hass R, Prudovsky I, Kruhoffer M. Institute of Anatomy, University Clinic Charite, Berlin, Germany. Human U937 myeloid leukemia cells were treated with different concentrations of 12-O-tetradecanoylphorbol-13-acetate (TPA) to determine signals that contribute to growth arrest and differentiation. While 0.5 nM TPA had little if any effect, exposure of U937 cells to higher TPA concentrations (5-500 nM) revealed a complete growth arrest after 48 h. Cytosolic PKC activity decreased by 50% after exposure to 0.5 nM TPA and by 80 and 95% after stimulation with 5 nM and 50 nM TPA, respectively. Simultaneously, the PKC activity in the particulate fraction of U937 cells increased accordingly. These events were associated with induction of a differentiated monocytic phenotype. Expression of the c-myc gene was down-regulated and c-jun and c-fms transcripts increased following exposure to 5-500 nM TPA. In contrast, exposure to 0.5 nM TPA decreased c-myc expression and increased c-jun transcripts only transiently between 4 and 8 h while little if any effect was detectable on c-fms mRNA expression and subsequent differentiation. Taken together, these data suggest that a certain initial threshold of PKC activation is required for induction of a differentiated monocytic phenotype while beyond this threshold, a growth-arrested and differentiated state in these human leukemic cells can be maintained regardless of TPA concentrations. PMID: 9301678 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 403: Cell Growth Differ. 1997 Sep;8(9):951-61. p53-independent induction of p21WAF1/CIP1 expression in pericentral hepatocytes following carbon tetrachloride intoxication. Serfas MS, Goufman E, Feuerman MH, Gartel AL, Tyner AL. Department of Genetics, University of Illinois at Chicago 60607, USA. The cyclin-dependent kinase, proliferating cell nuclear antigen, and stress-activated protein kinase/c-jun NH2 terminal kinase inhibitor p21WAF1/CIP1 can induce G1 arrest, and its expression coincides with the cessation of replication in many systems. We examined expression of p21 during the early stages of carbon tetrachloride intoxication in the mouse liver and observed a dramatic increase in p21 RNA levels between 4 and 8 h after administration. p21 expression, visualized by in situ hybridization, is induced in pericentral hepatocytes before carbon tetrachloride-induced necrosis. Examination of c-fos and c-myc expression patterns confirm that these immediate-early genes are induced in similar regions of the mouse liver. p21 induction is not dependent on p53; we observed similar levels and localization of p21 in wild-type and p53 null animals. Immunohistochemical localization of p21 and CCAAT/enhancer-binding protein expression shows that p21 protein accumulation is limited to a subset of CCAAT/enhancer-binding protein-positive hepatocytes. A second peak of periportal and intermediate zone-specific p21 gene expression, appearing 1-2 days after injection, is also p53 independent and may represent cell cycle checkpoints or postmitotic growth arrest. Sporadic p21 expression was also detected in pairs of hepatocytes distributed throughout the liver acini in healthy animals. Together, these data suggest several roles for p21 in the liver in response to toxicity, regeneration, and growth inhibition. PMID: 9300178 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 404: Exp Hematol. 1997 Sep;25(10):1042-50. Differential regulation by hematopoietic growth factors of apoptosis and mitosis in acute myeloblastic leukemia cells. Murohashi I, Yoshida K, Handa A, Murayoshi M, Yoshida S, Jinnai I, Bessho M, Hirashima K. First Department of Internal Medicine, Saitama Medical School, Japan. We evaluated the effects of various hematopoietic growth factors (HGFs) on the prevention of apoptosis in blasts from 19 patients with acute myeloblastic leukemia (AML) by assessing DNA ladder formation. After incubation without HGF, apoptosis was noted in all but two patients. HGFs prevented, did not affect, or enhanced apoptosis in 39 (60%), 14 (22%), or 12 (18%) of 65 suspension cultures, respectively. HGFs that prevented apoptosis also stimulated and/or synergized blast colony formation in 35 of 39 corresponding methylcellulose cultures. HGFs that alone stimulated colony formation also prevented apoptosis in all but two of 28 corresponding suspension cultures. In contrast, HGFs that did not prevent apoptosis also failed to stimulate growth in 17 of 26 corresponding methylcellulose cultures. HGFs that enhanced apoptosis alone never stimulated colony formation. After incubation, we noted enhanced c-fos and cjun genes as well as induction of p21 protein. An appropriate dose of HGF elevated c-fos, reduced c-jun and p21, induced G1/S transition, and inhibited apoptosis. In two patients, apoptosis was not induced after incubation. Cells not treated with HGF expressed no c-fos, c-jun, or c-myc, and remained in G0/G1. Taken together, our results support the conclusion that not only c-fos, cjun, and c-myc, but also p53 and p21 are required for blast apoptosis. HGF differentially prevents apoptosis and induces mitosis, and both events seem to be integral to the self-renewal of AML clonogenic cells. PMID: 9293901 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 405: Placenta. 1997 Sep;18(7):577-85. In vitro cultured human term cytotrophoblast: a model for normal primary epithelial cells demonstrating a spontaneous differentiation programme that requires EGF for extensive development of syncytium. Morrish DW, Dakour J, Li H, Xiao J, Miller R, Sherburne R, Berdan RC, Guilbert LJ. Signal Transduction Laboratories, University of Alberta, Edmonton, Canada. Normal human term cytotrophoblast cells prepared by trypsin-DNAse I digestion with and without secondary immunological purification with CD9 antibodies were investigated for the expression of morphological and genetic markers of proliferation and differentiation. After 24 h of culture, the cell preparations demonstrated spontaneous formation of microvilli and formation of small syncytial units as assessed by desmoplakin staining and FITC-dextran microinjection. EGF was required for mature syncytial formation. Compared to log-phase proliferating HeLa cells, uptake of [3H]thymidine incorporation was low and quickly decreased to negligible levels. Expression of the proto-oncogenes c-myc, c-fos, and c-jun and histone 2A decreased rapidly in the first 24 h of culture in both cell preparations, followed by an increase in expression of c-fos and junB over the next 3 days of culture. Proto-oncogene changes were similar in attached and suspension cells. Spontaneous increases in alpha hCG, pregnancy-specific beta(1)-glycoprotein and 3 beta-hydroxysteroid dehydrogenase (3 beta OHSD) occurred within 1 day in both cell preparations. EGF receptor blocking antibodies did not inhibit minor degrees of spontaneous syncytial formation nor inhibit spontaneous expression of alpha hCG or 3 beta OHSD mRNA, but did prevent extensive synctialization induced by EGF. The results demonstrate that term cytotrophoblast cells even in serum-free conditions or suspension culture rapidly commit to a non-proliferative differentiation program in culture which includes limited syncytialization and marked hormone mRNA expression. However, EGF is required for extensive syncytial development. PMID: 9290154 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 406: J Clin Endocrinol Metab. 1997 Sep;82(9):2962-5. Metastatic prolactinoma: effect of octreotide, cabergoline, carboplatin and etoposide; immunocytochemical analysis of proto-oncogene expression. Hurel SJ, Harris PE, McNicol AM, Foster S, Kelly WF, Baylis PH. Department of Medicine, Royal Victoria Infirmary, Newcastle upon Tyne, United Kingdom. A 49-yr-old woman presented with an extensive prolactinoma (serum PRL > 10,000 mU/L, normal range < 450 mU/L). Over a 5-yr period following transsphenoidal surgery and pituitary irradiation, she became increasingly resistant to high doses of bromocriptine and underwent transfrontal surgery followed by stereotactic radiotherapy. In spite of these treatments, serum prolactin estimations rose progressively to > 100,000 mU/L. Magnetic resonance imaging scanning demonstrated a massive cystic tumor invading the temporal lobes, extending into the cervical and thoracic spine, with metastases to cervical lymph nodes. High-dose cabergoline administration resulted in a 30% decrease in serum PRL. Octreotide was administered as a continuous sc infusion with a profound analgesic effect on facial pain but with no effect on tumor progression. She was treated with a course of chemotherapy consisting of carboplatin and etoposide without any noticeable effect. The patient died 6 months following chemotherapy. Immunocytochemical analysis demonstrated positive nuclear staining for WAF-1, Rb protein, c-myc, and p53 both in the original and metastatic tumors. The metastases but not the primary tumor stained for c-jun. Metastatic prolactinoma remains a therapeutic challenge. It is associated with a variable proto-oncogene expression, which may be coincidental or causal. Cabergoline had no advantage over bromocriptine. Octreotide relieved facial pain but did not alter tumor progression. An effective therapy for metastatic prolactinoma remains to be identified. Publication Types: Case Reports PMID: 9284727 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 407: J Toxicol Environ Health. 1997 Aug 29;51(6):609-21. Effects of glutathione depletion on cadmium-induced metallothionein synthesis, cytotoxicity, and proto-oncogene expression in cultured rat myoblasts. Shimizu M, Hochadel JF, Waalkes MP. National Cancer Institute-FCRDC, Frederick, Maryland, USA. Cadmium (Cd) is a highly toxic metal and a known carcinogen. Although the carcinogenic mechanism of action is unknown, Cd will induce transcriptional activation of c-myc and c-jun. We have previously found that the extent of Cd-induced oncogene expression is limited by the presence of cellular metallothionein (MT) in rat L6 myoblasts. Glutathione (GSH) is thought to play an important role in protection against Cd before the onset of MT synthesis. Thus, this study examined the effects of GSH depletion on Cd-induced MT synthesis, cytotoxicity, and proto-oncogene expression in rat L6 myoblasts after pretreatment with L-buthionine sulfoximine (BSO), a potent inhibitor of gamma-glutamyl-cysteine synthetase, which effectively depletes GSH. Exposure of L6 cells to BSO (5 or 25 microM) resulted in a dose-dependent decrease in cellular GSH levels. GSH depletion had no effect on Cd- or zinc-induced MT synthesis. Although the depletion of GSH was not itself cytotoxic in L6 cells, BSO pretreatment, particularly at the higher dose (25 microM), resulted in a dose-dependent increase in the sensitivity to Cd cytotoxicity, as assessed by a tetrazolium-based dye (MTT) assay. Low levels of Cd (1 microM) slightly increased the expression of both c-myc and c-jun as assessed by increases in gene-specific mRNA levels, in accordance with previous studies. GSH depletion (5 muM BSO) likewise caused an increase in expression of c-myc and c-jun. However, combined GSH depletion and Cd exposure decreased levels of c-myc and c-jun transcription well below control levels. These results suggest that increased cytotoxicity resulting from exposure to Cd after BSO depletion of cellular GSH abrogates the oncogene activation observed after either treatment alone. Thus proto-oncogene expression induced by Cd appears to be dependent on the absence of over Cd-induced cytotoxicity. PMID: 9242231 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 408: J Biol Chem. 1997 Aug 22;272(34):21334-40. Critical cytoplasmic domains of human interleukin-9 receptor alpha chain in interleukin-9-mediated cell proliferation and signal transduction. Zhu YX, Sun HB, Tsang ML, McMahel J, Grigsby S, Yin T, Yang YC. Department of Medicine (Hematology/Oncology), Indianapolis, Indiana 46202, USA. Interleukin-9 receptor (IL-9R) complex consists of a ligand-specific alpha chain and IL-2R gamma chain. In this study, two regions in the cytoplasmic domain of human IL-9Ralpha were found to be important for IL-9-mediated cell growth. A membrane-proximal region that contains the BOX1 consensus sequence is required for IL-9-induced cell proliferation and tyrosine phosphorylation of Janus kinases (JAKs). Deletion of this region or internal deletion of the BOX1 motif abrogated IL-9-induced cell proliferation and signal transduction. However, substitution of the Pro-X-Pro in the BOX1 motif with Ala-X-Ala failed to abolish IL-9-induced cell proliferation but decreased IL-9-mediated tyrosine phosphorylation of JAK kinases, insulin receptor substrate-2, and signal transducer and activator of transcription 3 (STAT3) and expression of c-myc and junB. Another important region is downstream of the BOX1 motif and contains a STAT3 binding motif YLPQ. Deletion of this region significantly impaired IL-9-induced cell growth, activation of JAK kinases, insulin receptor substrate-2, and STAT3 and expression of early response genes. A point mutation changing YLPQ into YLPA greatly reduced IL-9-induced activation of STAT3 and expression of c-myc but did not affect cell proliferation. These results suggest that cooperation or cross-talk of signaling molecules associated with different domains of IL-9Ralpha other than STAT3 is essential for IL-9-mediated cell growth. PMID: 9261146 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 409: Cancer. 1997 Aug 15;80(4):668-75. Oncogene expression in gastroenteropancreatic neuroendocrine tumors: implications for pathogenesis. Wang DG, Johnston CF, Buchanan KD. Division of Metabolism and Endocrinology, School of Clinical Medicine, The Queen's University of Belfast, United Kingdom. BACKGROUND: Neuroendocrine tumors of the gastroenteropancreatic system include pancreatic islet cell and carcinoid tumors. These tumors comprise a functionally and biologically heterogeneous group of neoplasms that rarely show reliable histopathologic signs of malignancy. No etiologic factors are proven to be associated with them, and their exact ontogeny and carcinogenesis remain unknown. METHODS: Monoclonal antibodies were employed, along with microwave antigen retrieval and the avidin-biotin immunohistochemical method, to investigate the expression of c-myc, bcl-2, c-erb B-2, c-erb B-3, c-jun, and proliferating cell nuclear antigen (PCNA) in a retrospective series of 116 primary gastroenteropancreatic neuroendocrine tumors (GPNTs). The authors attempted to correlate this expression with the clinicopathologic outcome of the disease. RESULTS: Immunoreactivities for c-myc, bcl-2, c-erb B-2, c-erb B-3, and c-jun were detected in 100%, 45%, 24%, 7%, and 24% of pancreatic neuroendocrine tumors (PNTs), respectively. In carcinoid tumors, immunoreactivities were detected for c-myc (63%), bcl-2 (28%), c-erb B-2 (31%), c-erb B-3 (6%), and c-jun (23%). There were significantly higher incidences of c-myc, bcl-2, and c-erb B-2 immunoreactivities in carcinoid tumors of the rectum than in those of the appendix, and significantly higher incidences of bcl-2 and c-jun immunoreactivities in carcinoid tumors of the bronchus than in those of the appendix. Incidence of PCNA immunoreactivity was significantly higher in malignant than in benign PNTs and also significantly higher in carcinoid tumors of the jejunum and ileum than in those of the appendix. CONCLUSIONS: The oncogenes c-myc, bcl-2, c-erb B-2, and c-jun are frequently expressed in human GPNTs. The expression of these oncogenes may represent pathogenic events in the generation, malignant transformation, and progression of GPNTs. The immunohistochemical evaluation of cell kinetics in GPNTs by PCNA might be a useful adjunct to conventional diagnostic procedures. PMID: 9264349 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 410: Pflugers Arch. 1997 Sep;434(5):568-74. In vivo carbon monoxide exposure and hypoxic hypoxia stimulate immediate early gene expression. Gess B, Wolf K, Pfeifer M, Riegger GA, Kurtz A. Institut fur Physiologie, Universitat Regensburg, Germany. This study aimed to examine the influence of acute tissue hypoxygenation on the expression of immediate early genes in different rat tissues. To this end male Sprague-Dawley rats were exposed to 0.1% carbon monoxide for 0.5, 1 and 6 h or to 9% oxygen for 6 h and mRNA levels for c-jun, c-fos, c-myc and EGR-1 were assayed by RNase protection in hearts, kidneys, livers and lungs. We found that hypoxia increased c-jun mRNA levels between twofold (lung) and eightfold (liver) in all organs examined; c-fos mRNA increased between three-fold (lung) and 20-fold (heart); c-myc mRNA increased between twofold (lung) and sixfold (heart); and EGR-1 mRNA increased between twofold (lung) and sixfold (heart). Our findings suggest that acute tissue hypoxygenation is a general stimulus of the expression of immediate early genes in vivo. With regard to the sensitivity to hypoxia, organ differences appear to exist in that the lung is rather insensitive, whilst the heart is rather sensitive. PMID: 9242720 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 411: Int J Cancer. 1997 Aug 7;72(4):687-95. Leukemia inhibitory factor induces apoptosis and proliferation of human carcinoma cells through different oncogene pathways. Kamohara H, Sakamoto K, Ishiko T, Masuda Y, Abe T, Ogawa M. Department of Surgery II, Kumamoto University School of Medicine, Japan. Leukemia inhibitory factor (LIF) affects the growth of carcinoma cells, and we thus analyzed its underlying mechanisms. Carcinoma cells constitutively express LIF mRNA, and 23 lines (92.0%) and all (100%) of 25 lines express LIF receptor mRNAs of LIFRbeta and gp130, respectively. Exogenous addition of LIF promoted significant cell proliferation in 4 lines (MCF-7, ZR-75-1, Hs-700T and Panc-1) and suppressed cell growth in 3 lines (AZ-521, GBK-1 and HT-29). LIF significantly induced an immediate early response of genes c-fos and junB 3 hr after stimulation, but not of c-jun during the process of proliferation of MCF-7 and Hs-700T cells, with maximum levels at 30-60 min. The cell-cycle-related gene cyclin E was also induced in MCF-7 and Hs-700T cells, whereas cyclinA, cdk2, c-myc, c-myb and p53 mRNAs were not induced. On the other hand, LIF inhibited growth and increased the rate of cell death of AZ-521 and GBK-1 cells. LIF increased the number of TUNEL-positive cells in AZ-521 cells and DNA fragmentation in AZ-521 and GBK-1 cells. LIF induced apoptosis related genes c-myc and ICE during suppression of cell growth, but p53, p21, c-fos, cyclin A and cyclin E were not induced. Our results suggest that LIF is linked to cell proliferation and apoptosis in some human carcinoma cell lines. It is considered that this is related to differences in signal transduction and induction of oncogenes. PMID: 9259411 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 412: Mol Cell Biochem. 1997 Aug;173(1-2):59-69. Hydrogen peroxide-induced expression of the proto-oncogenes, c-jun, c-fos and c-myc in rabbit lens epithelial cells. Li DW, Spector A. Department of Ophthalmology, College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA. The involvement of H2O2 in cataract development has been established in both human patients and animal models. At the molecular level H2O2 has been observed to cause damage to DNA, protein and lipid. To explore the oxidative stress response of the lens system at the gene expression level, we have examined the effects of H2O2 on the mRNA change of the proto-oncogenes, c-jun, c-fos and c-myc in a rabbit lens cell line, N/N1003A. H2O2 treatment of the rabbit lens epithelial cells for 60 min induces quick up-regulation of both c-jun and c-fos mRNAs. The maximal induction is 38 fold for c-jun at 150 microM and 72 fold for c-fos at 250 microM H2O2. Treatment of N/N1003A cells with 50-250 microM H2O2 for 60 min leads to a 2-5 fold increase of the c-myc mRNA level. H2O2 also induces an up-regulation in transactivity of the activating protein-1 (AP-1) as shown with a reporter gene driven by a prolactin gene promoter with 4 copies of AP-1 binding sites inserted in the upstream of the promoter. Maximal induction occurs with 150 microM H2O2. In the same system, the antioxidants, N-acetyl-cysteine (NAC) and pyrrolidine dithiocarbamate (PDTC) at concentrations shown to up-regulate the mRNAs of both c-jun and c-fos, also enhance the transactivity of AP-1. NAC and PDTC have different effects in modulating the induction of AP-1 activity by H2O2 and TPA. These results reveal that oxidative stress regulates expression of various regulatory genes in lens systems, which likely affects cell proliferation, differentiation and viability and thus affect normal lens functions. PMID: 9278255 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 413: Pancreas. 1997 Aug;15(2):160-7. Expression of the protooncogene jun is induced in the rat pancreas by cerulein infusion. Ferrara C, Gress TM, Mueller-Pillasch F, Lutz MP, Weidenbach H, Poletti A, Lerch MM, Del Favero G, Adler G. Department of Gastroenterology, University of Padova, Italy. Cholecystokinin (CCK) can stimulate secretion and DNA synthesis in pancreatic acinar cells. Hyperstimulation with cerulein (a CCK analogue) induces acute edematous pancreatitis. To study the effects of in vivo pancreatic stimulation with cerulein, we analyzed the expression of the protooncogenes jun, myc, and fos on the mRNA and protein levels. RNA and protein were extracted from the pancreas of rats administered an infusion of cerulein, 10 micrograms/kg/h (Group A) or 0.25 microgram/kg/h (Group B), or saline (Group C) and sacrificed 2, 4, and 6 h after beginning the infusion and 0, 12, and 24 h and 2, 4, and 6 days after completing the infusion period. Transcript levels were studied using slot-blot analysis. Protein expression was studied using Western blot and immunohistochemistry. No changes were found for the expression of protooncogenes myc and fos on either the transcript or the protein levels. Higher jun mRNA levels were found in Group A than in Group B or C, particularly after 2 h of infusion and 12, 24, and 48 h after the end of a 12-h cerulein infusion. No significant difference was observed in Groups B and C. The jun protein behavior was similar in Groups A and B, revealing two peaks: one early during infusion and a second one after the end of a 12-h cerulein infusion. Jun protein was found mainly in the acinar cells. In conclusion, (1) acinar cells in the rat pancreas respond to cerulein stimulation by increasing the expression of jun; (2) in vivo high doses of cerulein increase the jun mRNA and jun protein levels, whereas low doses raise only the protein levels; and (3) myc and fos are apparently uninfluenced by cerulein administration. PMID: 9260201 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 414: J Cell Physiol. 1997 Aug;172(2):137-45. Induction in human osteoblastic cells (SaOS2) of the early response genes fos, jun, and myc by the amino terminal fragment (ATF) of urokinase. Rabbani SA, Gladu J, Mazar AP, Henkin J, Goltzman D. Department of Medicine, McGill University, Montreal, Quebec, Canada. Previous studies have demonstrated that overexpression of urinary plasminogen activator (uPA) in rat prostate cancer cells results in increased skeletal metastases, which are primarily of the osteoblastic variety. The osseous activation induced by the metastases appears to be mediated through the amino terminal fragment (ATF) of uPA, which lacks the catalytic domain and can act as a growth factor for osteoblasts. To explore further the mechanism of action of uPA in bone cells, we evaluated the effects of ATF on modulating the expression of various proto-oncogenes. Human-osteoblast-derived osteosarcoma cells, SaOS2, were treated with graded doses of ATF for 10-120 min, and effects on early response proto-oncogenes were monitored. ATF increased c-myc, c-jun, and c-fos gene expression in a time-dependent manner for up to 60 min, after which mRNA levels fell. The maximum induction was seen in c-fos gene expression, which was found to be dose dependent. This effect of ATF was localized to its growth-factorlike domain. Examination of the half life of these transcripts in the presence of the transcriptional inhibitor actinomycin D demonstrated that ATF does not alter the stability of c-fos mRNA in these bone cells. Nuclear run-off assays indicated that ATF effects were due to stimulation of c-fos gene transcription. An increase in c-fos protein levels was correlated with the augmentation of its mRNA in ATF-treated SaOS2 cells. Pretreatment of SaOS2 cells with the protein tyrosine kinase inhibitor herbimycin and recombinant soluble uPA receptor (uPAR) caused a significant reduction in the ability of ATF to induce c-fos expression. These results demonstrate a novel role for uPA in activating early response proto-oncogenes, in particular c-fos, which plays an important role in bone cell growth and differentiation and may be a key factor in the signal transduction pathway of ATF. PMID: 9258335 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 415: J Clin Endocrinol Metab. 1997 Aug;82(8):2596-600. Comparative messenger ribonucleic acid analysis of immediate early genes and sex steroid receptors in human leiomyoma and healthy myometrium. Lessl M, Klotzbuecher M, Schoen S, Reles A, Stockemann K, Fuhrmann U. Research Laboratories of Schering AG, Berlin, Germany. To shed light on the molecular mechanisms involved in the pathogenesis of uterine leiomyomas, transcript levels of the immediate early genes c-fos, c-myc, and c-jun and of the estrogen receptor (ER) and progesterone receptor (PR) were determined in tissue samples of human myometrium and leiomyoma. The messenger RNA (mRNA) content was analyzed by RT-PCR. mRNAs for c-fos, c-myc, c-jun, ER, and PR were detected in all 18 samples of leiomyoma and corresponding myometrial tissue collected in this study. Interestingly, in contrast to healthy tissues, we found a distinct and significant reduction of c-fos mRNA in the tumor. These data were substantiated by the finding of lowered c-Fos protein levels in leiomyomas tissues. Moreover, transcripts of c-jun and c-myc were less abundant in most of the leiomyomas than in the myometrium. This different expression of the protooncogenes in leiomyomas and myometrium was independent of the phase of the menstrual cycle in which samples were collected. In contrast to the reduced transcript levels observed for the immediate early genes, the ER and PR mRNA contents of the leiomyomas and myometrium did not differ. These results were confirmed by immunohistochemical studies for ER and PR protein. In conclusion, our data show that the deregulated expression of protooncogenes, especially of c-fos, is linked to the pathogenesis of leiomyomas. Confirmation of a potential role of downregulated c-fos levels for the benign character of these tumors requires further investigation. Additionally, the findings suggest that sex steroids do not influence the different expression patterns of c-fos, c-myc, and c-jun in leiomyomas, as compared with myometrium. PMID: 9253340 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 416: Cell Tissue Res. 1997 Aug;289(2):253-64. Multiple mechanisms contribute to myenteric plexus ablation induced by benzalkonium chloride in the guinea-pig ileum. Parr EJ, Sharkey KA. Neuroscience Research Group, Department of Physiology and Biophysics, The University of Calgary, 3330 Hospital Drive N.W., Calgary, Alberta, Canada T2N 4N1. Ablation of rat myenteric plexus with benzalkonium chloride has provided a model of intestinal aganglionosis, but the degenerative responses are not well understood. We examined the effects of this detergent on neurons and glia, including expression of c-Myc, c-Jun, JunB, and c-Fos, and on immunocytes in the guinea-pig ileum. Benzalkonium chloride (0.1%) or saline was applied to the serosal surface of distal ileum. Tissues were analyzed 2, 3, or 7 days later and compared with cyclosporine-treated and untreated animals. More than 90% of myenteric neurons were destroyed in ileal segments 3-7 days after benzalkonium-chloride treatment. Glia withdrew processes from around neurons after 2 days and were mostly gone after 3 days. Neuronal c-Myc began to disappear while c-Fos, c-Jun, and JunB were evident in some neuronal nuclei after 2 or 3 days. After 3 days, widespread apoptosis was evident in the myenteric plexus. Populations of T cells, B cells, and macrophage-like cells in untreated and saline-treated myenteric plexuses were substantially increased 3 and 7 days after benzalkonium-chloride treatment. Cyclosporine delayed significant neuronal loss. We conclude that a variety of degenerative mechanisms may be active in this model, including an immune response which may actively contribute to tissue destruction. PMID: 9211828 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 417: Cancer Res. 1997 Jul 15;57(14):3016-25. Human ornithine decarboxylase-overproducing NIH3T3 cells induce rapidly growing, highly vascularized tumors in nude mice. Auvinen M, Laine A, Paasinen-Sohns A, Kangas A, Kangas L, Saksela O, Andersson LC, Holtta E. Department of Pathology, University of Helsinki, Finland. Overexpression of human ornithine decarboxylase (ODC) under the control of strong promoters induces morphological transformation of immortalized NIH3T3 and Rat-1 fibroblasts [M. Auvinen et al., Nature (Lond.), 360: 355-358, 1992]. We demonstrate here that ODC-overproducing NIH3T3 cells are tumorigenic in nude mice, giving rise to rapidly growing, large fibrosarcomas at the site of inoculation. The tumors are capable of invading host fat and muscle tissues and are vascularized abundantly. To disclose the molecular mechanism(s) driving the tumorigenic, invasive, and angiogenic phenotype of the tumors, the ODC-overproducing cell lines and tumor tissues were analyzed for the expression of various potential regulators and mediators of cell proliferation, matrix degradation, and angiogenesis. The tumorigenicity of ODC transformants was associated with elevated polyamine levels and down-regulated growth factor receptors. The invasiveness of the ODC-induced tumors could not be attributed to overexpression of various known extracellular matrix-degrading proteases or matrix metalloproteinases. The induction of the tumor neovascularization proved not to be elicited by vascular endothelial growth factor or basic fibroblast growth factor. Instead, the ODC-overexpressing cells appeared to secrete a novel angiogenic factor(s) that was able to promote migration of bovine capillary endothelial cells in collagen gels and increase the proliferation of human endothelial cells in vitro. In parallel, ODC-transformed cells displayed down-regulation of thrombospondin-1 and -2, the negative regulators of angiogenesis. Thus, the induction of the angiogenic phenotype of the ODC transformants is likely due both to increased expression and secretion of the new angiogenesis-stimulating factor(s) and decreased production and release of the antiangiogenic thrombospondins. PMID: 9230217 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 418: J Biol Chem. 1997 Jul 4;272(27):16917-23. Conditional expression of the mitogen-activated protein kinase (MAPK) phosphatase MKP-1 preferentially inhibits p38 MAPK and stress-activated protein kinase in U937 cells. Franklin CC, Kraft AS. Department of Medicine, Division of Medical Oncology, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA. chris.franklin@uchsc.edu Phorbol ester tumor promoters, such as phorbol 12-myristate 13-acetate (PMA), are potent activators of extracellular signal-regulated kinase 2 (ERK2), stress-activated protein kinase (SAPK), and p38 mitogen-activated protein kinase (MAPK) in U937 human leukemic cells. These kinases are regulated by the reversible dual phosphorylation of conserved threonine and tyrosine residues. The dual specificity protein phosphatase MAPK phosphatase-1 (MKP-1) has been shown to dephosphorylate and inactivate ERK2, SAPK, and p38 MAPK in transient transfection studies. Here we demonstrate that PMA treatment induces MKP-1 protein expression in U937 cells, which is detectable within 30 min with maximal levels attained after 4 h. This time course coincides with the rapid inactivation of PMA-induced SAPK activity, but not ERK2 phosphorylation, which remains elevated for up to 6 h. To examine directly the role of MKP-1 in the regulation of these protein kinases in vivo, we established a U937 cell line that conditionally expresses MKP-1 from the human metallothionein IIa promoter. Conditional expression of MKP-1 inhibited PMA-induced ERK2, SAPK, and p38 MAPK activity. By titrating the levels of MKP-1 expression from the human metallothionein IIa promoter, however, it was found that p38 MAPK and SAPK were much more sensitive to inhibition by MKP-1 than ERK2. This differential substrate specificity of MKP-1 can be functionally extended to nuclear transcriptional events in that PMA-induced c-Jun transcriptional activity was more sensitive to inhibition by MKP-1 than either Elk-1 or c-Myc. Conditional expression of MKP-1 also abolished the induction of endogenous MKP-1 protein expression in response to PMA treatment. This negative feedback regulatory mechanism is likely due to MKP-1-mediated inhibition of ERK2, as studies utilizing the MEK1/2 inhibitor PD98059 suggest that ERK2 activation is required for PMA-induced MKP-1 expression. These findings suggest that ERK2-mediated induction of MKP-1 may play an important role in preferentially attenuating signaling through the p38 MAPK and SAPK signal transduction pathways. PMID: 9202001 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 419: Cell Growth Differ. 1997 Jul;8(7):731-42. Regulation of Myc-dependent apoptosis by p53, c-Jun N-terminal kinases/stress-activated protein kinases, and Mdm-2. Yu K, Ravera CP, Chen YN, McMahon G. Oncology Research Program, Preclinical Research, Sandoz Pharmaceuticals Corporation, East Hanover, New Jersey 07936, USA. Deregulated overexpression of c-Myc (Myc) confers susceptibility to apoptosis in several cell types, but the molecular regulation of these processes has not been well established. Here we have characterized several molecular changes that may modulate Myc-dependent apoptosis. Ectopic overexpression of Myc in both Rat1 fibroblasts and human osteosarcoma cells causes a dramatic increase of cellular p53 mRNA and protein, and this induction of p53 correlates with apoptosis triggered by withdrawal of serum. Stable transfection of a wild-type human p53 gene into Myc-transformed cells further potentiates apoptosis. Anticancer agents vinblastine and nocodazole also induce apoptosis in Myc-transformed Rat1 fibroblasts but are cytostatic to the same cells without Myc overexpression. We demonstrate that induction of Myc-dependent apoptosis in these cells is specifically associated with an activation of p46 c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK) activity, whereas this JNK/SAPK activation is absent in stress-treated cells without Myc overexpression. Moreover, overexpression of the Mdm-2 gene in Rat1-myc cells significantly inhibits apoptosis induced by low serum but has little effect on apoptosis triggered by chemotherapeutic drugs. Interestingly, differential inhibition by Mdm-2 paralleled differential activation of p46 JNK/SAPK. Thus, our data support a functional involvement of p53 in Myc-dependent apoptosis and implicate potential regulatory roles for JNK/SAPK and Mdm-2 pathways in the regulation of apoptosis in Myc-transformed tumor cells. PMID: 9218867 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 420: Endocrinology. 1997 Jul;138(7):2740-6. Transcriptional regulation of Sertoli cell immediate early genes by interleukin-6 and interferon-gamma is mediated through phosphorylation of STAT-3 and STAT-1 proteins. Jenab S, Morris PL. The Population Council, New York, New York 10021, USA. The immediate early genes are regulated by a variety of extracellular signals, including pleiotropic cytokines. The effects of the testicular cytokines, interleukin-6 (IL-6) and interferon-gamma (IFN-gamma), on signal transducers and activators of transcription 3 and 1 (STAT-3 and STAT-1) and on c-fos gene expression in primary Sertoli cells are suggestive of their roles in differential function. Using the tyrosine phosphorylation inhibitor, genistein, and electrophoretic mobility shift assay, we show that IL-6 and IFN-gamma induce nuclear factor STAT-3 and STAT-1 DNA-binding activity to the sis-inducible element of c-fos in a genistein-dependent pathway. Quantitative solution hybridization, Northern blot, and nuclear run-on analysis show that differential induction of c-fos, junB, and c-myc messenger RNA (mRNA) by these cytokines occur at transcriptional levels. IL-6 stimulates c-fos mRNA levels by 6-fold while increasing junB levels by 2-fold. IFN-gamma increases c-fos message 2-fold, but has no effect on junB mRNA levels. Furthermore, genistein treatment blocks the induction of c-fos and junB gene expression, demonstrating that tyrosine phosphorylation of STAT proteins is involved in the cytokine regulation of the Sertoli immediate early genes. H7, a serine/threonine phosphorylation inhibitor, also blocks c-fos gene induction by IL-6 and IFN-gamma, but does not affect the DNA-binding activities of STAT-3 and STAT-1. Finally, IL-6 treatment of Sertoli cells (3-6 h) increases the amounts of activating protein-1 binding to activating protein-1 element and c-myc transcription. PMID: 9202212 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 421: DNA Cell Biol. 1997 Jun;16(6):797-9. A simple improvement in expression cloning. Takemoto Y, Furuta M, Sato M, Hashimoto Y. Institute of Immunology, Syntex-Roche, Yamazaki Noda, Chiba, Japan. Expression cloning is an effective approach for isolating genes encoding proteins that associate with a target species. Several molecules have been isolated by expression cloning, including CRE-BP1 associating with Jun (Macgregor et al., 1990); Grb1, identical to p85 PI3-kinase, with the EGF receptor (Skolnik et al., 1991); and Max with Myc (Blackwood and Eisenman, 1991). Expression cloning involves induction of proteins from a lambda gt11 cDNA expression library and screening the proteins on nitrocellulose membranes using a peptide probe (Macgregor et al., 1990). With this method, we previously isolated an Lck tyrosine kinase-associated protein, LckBP1, which is identical to HS1 (Kitamura et al., 1989, 1995; Takemoto et al., 1995). In those experiments, we used a glutathione S-transferase (GST)-Lck SH3 domain fusion protein as a probe, followed by detection of the complex with anti-GST polyclonal antibody. Whereas the ease of obtaining the fusion construct and high-titer anti-GST polyclonal antibody represented clear advantages, the system suffered from high background and low sensitivity. Here we show that pretreatment of nitrocellulose filters with NaDodSO4 reduces background and, in turn, increases sensitivity. PMID: 9212173 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 422: Toxicol Appl Pharmacol. 1997 Jun;144(2):247-61. Cadmium, gene regulation, and cellular signalling in mammalian cells. Beyersmann D, Hechtenberg S. Department of Biology and Chemistry, University of Bremen, Germany. Effects of the carcinogenic metal cadmium on the regulation of mammalian gene expression are reviewed and discussed in the light of observations on interference with cellular signal transduction pathways. Cadmium ions are taken up through calcium channels of the plasma membrane of various cell types, and cadmium is accumulated intracellularly due to its binding to cytoplasmic and nuclear material. At elevated cytotoxic concentrations, cadmium inhibits the biosyntheses of DNA, RNA, and protein, and it induces lipid peroxidation, DNA strand breaks, and chromosome aberrations. Cadmium compounds as such are only weak mutagens and clastogens. However, cadmium at noncytotoxic doses interferes with DNA repair processes and enhances the genotoxicity of directly acting mutagens. Hence, the inhibition of repair and detoxifying enzymes by this metal may partially explain the observed weak genotoxic properties of this metal. Nongenotoxic mechanisms upregulating intracellular signalling pathways leading to increased mitogenesis are discussed as major mechanisms for the interpretation of the carcinogenic activity by chronic cadmium exposure. About 1 microM cadmium stimulates DNA synthesis and cell proliferation in various cell lines, whereas more elevated concentrations are inhibitory. Cadmium enhances the expression of several classes of genes at concentrations of a few microM. It stimulates the expression of immediate early genes (c-fos, c-jun, and c-myc), of the tumor suppressor gene p53, and of genes coding for the syntheses of protective molecules, including metallothioneins, glutathione, and stress (heat shock) proteins. The mechanisms underlying the modulation of gene activity by cadmium are discussed in terms of interference with cellular signalling at the levels of cell surface receptors, cellular calcium and zinc homeostases, protein phosphorylation, and modification of transcription factors. In considering the available evidence, the carcinogenic properties of cadmium are interpreted using a multifactorial approach involving indirect genotoxicity (interference with DNA repair) and the upregulation of mitogenic signalling pathways. Publication Types: Review Review, Tutorial PMID: 9194408 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 423: J Biol Chem. 1997 May 23;272(21):13816-22. Protein kinase C delta inhibits the proliferation of vascular smooth muscle cells by suppressing G1 cyclin expression. Fukumoto S, Nishizawa Y, Hosoi M, Koyama H, Yamakawa K, Ohno S, Morii H. Second Department of Internal Medicine, Osaka City University Medical School, Osaka 545, Japan. To elucidate the physiological role of protein kinase C (PKC) delta, a ubiquitously expressed isoform in vascular smooth muscle cells (VSMC), PKC delta was stably overexpressed in A7r5 cells, rat clonal VSMC. The [3H]thymidine incorporation in A7r5 overexpressed with PKC delta (DVs) was suppressed to 37.1 +/- 16.3% (mean +/- S.D.) of the level in control or A7r5 transfected with vector alone (EVs). The reduction of [3H]thymidine incorporation was strongly correlated with overexpressed PKC levels. Moreover, transient transfection of a dominant negative mutant of PKC delta restored the reduced proliferation in DVs. Flow cytometry analysis demonstrated that DVs were arrested in the G0/G1 phase of the cell cycle. Expression of cyclins D1 and E and retinoblastoma protein phosphorylation were reduced, while the protein levels of p27 were elevated in DVs as compared with EVs. There were no significant differences in the expression of c-fos, c-jun, c-myc, cyclin D2, D3, cyclin-dependent kinase 2, cyclin-dependent kinase 4, and p21 among the clones. We conclude that PKC delta inhibits the proliferation of VSMC by arresting cells in G1 via mainly inhibiting the expression of cyclin D1 and cyclin E. PMID: 9153238 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 424: Jpn J Cancer Res. 1997 May;88(5):435-42. S-methylcysteine and cysteine are inhibitors of induction of glutathione S-transferase placental form-positive foci during initiation and promotion phases of rat hepatocarcinogenesis. Takada N, Yano Y, Wanibuchi H, Otani S, Fukushima S. First Department of Pathology, Osaka City University Medical School. S-Methylcysteine (SMC) occurs in a variety of plants, including Allium sativum, Phaseolus vulgaris, and Cruciferae. In this study, we synthesized five organosulfur compounds (OSCs), SMC and four analogs, and examined their modifying effects on diethylnitrosamine-induced neoplasia of the liver in male F344 rats, using the medium-term bioassay system of Ito (Ito test) based on the two-step model of hepatocarcinogenesis. In addition, we investigated the modifying effects of SMC and cysteine on the initiation stage of rat hepatocarcinogenesis. Carcinogenic potential was scored by comparing the numbers and areas of induced glutathione S-transferase placental form (GST-P)-positive hepatocellular focl. All OSCs examined had a tendency to decrease the number of GST-P-positive foci when given in the promotion stage of the Ito test, and in particular SMC and cysteine exerted significant inhibitory effects. When given during the initiation stage, these two OSCs also significantly inhibited focus formation. Regarding the mechanism underlying the inhibitory effects of SMC and cysteine, measurement of ornithine decarboxylase in SMC- and cysteine-treated liver tissues after partial hepatectomy (PH) revealed a significantly reduced activity, and the proportion of hepatocytes positive for proliferating cell nuclear antigen was significantly decreased by SMC or cysteine administration. Moreover, examination of the expression of the early response proto-oncogenes, c-fos, c-jun, and c-myc, after PH demonstrated down-regulated induction of c-jun mRNA transcripts by SMC, sustained for an eight-hour period. Our results support the view that SMC and cysteine are chemopreventive agents for rat hepatocarcinogenesis and that their intake may be importance for cancer prevention. PMID: 9247599 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 425: Photochem Photobiol. 1997 May;65(5):908-14. Gene expression in skin tumors induced in hairless mice by chronic exposure to ultraviolet B irradiation. Sato H, Suzuki JS, Tanaka M, Ogiso M, Tohyama C, Kobayashi S. Kyoritsu College of Pharmacy, Tokyo, Japan. We investigated the expressions of c-Ha-ras, c-jun, c-fos, c-myc genes and p53 protein in the development of skin tumors induced by chronic exposure to UVB without a photosensitizer using hairless mice. When mice were exposed to UVB at a dose of 2 kJ/m2 three times a week, increased c-Ha-ras and c-myc transcripts were detected after only 5 weeks of exposure, while no tumor appeared on the exposed skin. The increase in gene expression continued until 25 weeks, when tumors, identified pathologically as mainly squamous cell carcinomas (SCC), developed in the dorsal skin. In these SCC, overexpression of c-fos mRNA was also observed along with the increases in c-Ha-ras and c-myc. A single dose of UVB (2 kJ/m2) applied to the backs of hairless mice transiently induced overexpression of the early event genes c-fos, c-jun and c-myc, but not c-Ha-ras, in the exposed area of skin. Accumulation of p53 protein was determined by Western blotting analysis or immunohistochemistry using monoclonal antibodies PAb 240 or 246, which recognize mutant or wild type, respectively. In the SCC, a mutant p53 protein accumulated in the cytoplasm and nucleus. After single-dose irradiation, the increased wild-type p53 protein was observed in the nuclei of epidermal cells. The present results suggest that overexpression of the c-fos, c-myc and c-Ha-ras genes, and the mutational changes in p53 protein might be associated with skin photocarcinogenesis. Moreover, overexpression of the c-Ha-ras and c-myc genes might be an early event in the development of UVB-induced skin tumors in mice. PMID: 9155265 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 426: Hepatology. 1997 May;25(5):1123-7. The effect of changes in hepatocyte membrane potential on immediate-early proto-oncogene expression following partial hepatectomy in rats. Minuk GY, Kren BT, Xu R, Zhang X, Burczynski F, Mulrooney NP, Fan G, Gong Y, Steer CJ. Department of Medicine, University of Manitoba, Winnipeg, Canada. The stimulus responsible for inducing hepatocytes to enter the cell cycle following partial hepatectomy (PHx) remains to be identified. One suggested candidate is the change in hepatocyte membrane potential (PD) that occurs immediately following PHx. To test this possibility, we monitored changes in hepatocyte PD and immediate-early proto-oncogene expression in rats pretreated with saline or gamma aminobutyric acid (GABA), an amino-acid neurotransmitter that hyperpolarizes isolated hepatocytes. Intraperitoneal injections of saline or GABA (500 microg/g body weight) were administered to adult, male Sprague-Dawley rats 1 hour prior to 70% PHx. Rats were sacrificed and the livers excised at various times until 180 minutes post-PHx for messenger RNA (mRNA) and protein analyses. In additional groups of saline- and GABA-treated rats, PD changes were recorded continuously from -260 to 180 minutes post-PHx. Serum GABA concentrations were determined by ion-exchange chromatography with orthopthaldehyde fluorescence detection. Hepatocyte PD's were recorded in situ by intracellular microelectrodes with an Axoprobe-1A amplifier. Steady-state levels of c-fos, c-jun, jun-B, and c-myc transcripts and proteins were documented by Northern blots of poly(A)-enriched RNA derived from resected livers and Western blots of total nuclear protein, respectively. Serum GABA concentrations remained unchanged in saline-treated controls but increased 10- to 20-fold above baseline values in GABA-treated rats. In saline-treated controls, hepatocyte depolarization occurred immediately and was maintained throughout the 180 minutes post-PHx period (PD pre-PHx, -36.8 +/- 5.1; 15 minutes post-PHx, -27.5 +/- 5.7; and 180 minutes post-PHx, -28.3 +/- 4.4 mV, mean +/- SD); whereas in GABA-treated rats, hepatocyte PD remained unchanged (-37.0 +/- 1.1; -36.4 +/- 3.1 and -39.2 +/- 2.7 mV, respectively). Despite abrogation of hepatocyte PD changes, proto-oncogene mRNA and protein levels in saline- and GABA-treated rats were either similar or, in the case of c-fos and c-jun, increased five- to sevenfold in GABA-treated rats. The results of this study indicate the following: 1) hepatocytes depolarize immediately post-PHx and remain depolarized throughout the priming phase of the cell cycle; 2) elevated serum GABA concentrations prevent PHx-induced hepatocyte depolarization; and 3) depolarization is not the stimulus responsible for priming hepatocytes into replicative competence. PMID: 9141428 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 427: FEBS Lett. 1997 Apr 7;406(1-2):17-22. On the involvement of calpains in the degradation of the tumor suppressor protein p53. Gonen H, Shkedy D, Barnoy S, Kosower NS, Ciechanover A. Department of Biochemistry, Faculty of Medicine, Technion-Israel Institute of Technology, Haifa. A crude fraction that contains ubiquitin-protein ligases contains also a proteolytic activity of approximately 100 kDa that cleaves p53 to several fragments. The protease does not require ATP and is inhibited in the crude extract by an endogenous approximately 250 kDa inhibitor. The proteinase can be inhibited by chelating the Ca2+ ions, by specific cysteine proteinase inhibitors and by peptide aldehyde derivatives that inhibit calpains. Purified calpain demonstrates an identical activity that can be inhibited by calpastatin, the specific protein inhibitor of the enzyme. Thus, it appears that the activity we have identified in the extract is catalyzed by calpain. The calpain in the extract degrades also N-myc, c-Fos and c-Jun, but not lysozyme. In crude extract, the calpain activity can be demonstrated only when the molar ratio of the calpain exceeds that of its native inhibitor. Recent experimental evidence implicates both the ubiquitin proteasome pathway and calpain in the degradation of the tumor suppressor, and it was proposed that the two pathways may play a role in targeting the protein under various conditions. The potential role of the two systems in this important metabolic process is discussed. PMID: 9109377 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 428: Ann Rheum Dis. 1997 Apr;56(4):262-7. Characterisation of fibroblast-like cells in pannus lesions of patients with rheumatoid arthritis sharing properties of fibroblasts and chondrocytes. Xue C, Takahashi M, Hasunuma T, Aono H, Yamamoto K, Yoshino S, Sumida T, Nishioka K. Department of Rheumatology and Immunology, St Marianna University School of Medicine, Kawasaki, Japan. OBJECTIVE: To better understand the characteristics of synoviocytes located in the rheumatoid arthritis (RA) pannus. METHODS: One cell line, termed PSC, was cloned from RA pannus lesions. Phenotypic analysis was done by contrast microscopy, indirect immunostaining, and safranin O staining. Transcription of several protooncogenes and matrix degrading enzymes was evaluated. The expression of mRNA for collagen II was detected by in situ hybridisation. The ability of anchorage independent growth was assessed by soft agarose culture. RESULTS: PSCs showed a high transcription of protooncogenes c-fos, c-myc and c-jun. They also expressed mRNA for matrix degrading enzymes, such as collagenase, cathepsin B, and cathepsin L. Anchorage independent growth assay demonstrated that PSCs formed colonies in soft agar culture. Phenotypic analysis showed that this fibroblast-like PSC was stained intensely with anti-vimentin and anti-fibroblast antibody. In situ reverse transcriptase assay showed that the cell line expressed type II collagen mRNA. CONCLUSION: Alternative fibroblast-like cells were identified in the pannus lesion of RA sharing properties of fibroblasts and chondrocytes. These findings suggest that this fibroblast-like cell derived from pannus lesions may contribute to the destruction of the cartilage in RA. PMID: 9166000 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 429: Immunol Invest. 1997 Apr;26(3):351-70. An RGD containing peptide from HIV-1 Tat-(65-80) modulates protooncogene expression in human bronchoalveolar carcinoma cell line, A549. el-Solh A, Kumar NM, Nair MP, Schwartz SA, Lwebuga-Mukasa JS. Department of Internal Medicine, SUNY at Buffalo School of Medicine and Biomedical Sciences, Buffalo General Hospital 14203, USA. Tat (transactivator of transcription) is essential for HIV-1 replication in vivo and in vitro. Tat-(65-80), an RGD containing domain, has been shown to regulate proliferative function of a variety of cell lines, including a human adenocarcinoma cell line, A549. The exact cellular and molecular mechanisms by which these effects are mediated, remain unknown. To evaluate the hypothesis that Tat-(65-80) modulates the expression of immediate early genes (IEG) c-jun, c-myc, c-fos and the tumor suppressor gene p53, serum starved A549 cells were incubated with Tat-(65-80) or heat-inactivated Tat-(65-80) at 10 ng/ml. Total cellular RNA was isolated from the cells at various time points (0-24 hours). In each case, 5 micrograms of RNA was reverse transcribed in 20 microliters of reaction volume. Equal amounts of cDNA were subjected to polymerase chain reaction (PCR) and analyzed by electrophoresis. The photographic negatives of the ethidium bromide stained gels were quantitated by densitometric scanning and normalized to corresponding beta-actin PCR products. Treatment with Tat-(65-80) showed a twofold induction of c-jun at 0.5 h. Peak expression occurred at 60 minutes and remained above baseline at 24 hours (h). c-myc was increased at 0.5 h, reached a twofold increase at 2 h and remained above baseline at 24 h. c-fos increased seven fold at 0.5 h and declined subsequently to baseline at 8 h. p-53 gene was reduced fivefold at 0.5 h and remained downregulated thereafter. These results show that Tat-(65-80) can modulate growth related genes in human lung epithelial cells. PMID: 9129988 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 430: Cell Growth Differ. 1997 Apr;8(4):425-34. Neonatal diethylstilbestrol treatment alters the estrogen-regulated expression of both cell proliferation and apoptosis-related proto-oncogenes (c-jun, c-fos, c-myc, bax, bcl-2, and bcl-x) in the hamster uterus. Zheng X, Hendry WJ 3rd. Department of Biological Sciences, Wichita State University, Kansas 67260-0026, USA. In the Syrian hamster, neonatal diethylstilbestrol (DES) treatment and then postpubertal estrogen stimulation induces hyperplasia plus apoptosis (preneoplastic responses) and ultimately neoplasia in the endometrial epithelial cell compartment. As part of a project to investigate the molecular and cellular mechanisms responsible for this phenomenon, expression of several proto-oncogenes (c-jun, c-fos, c-myc, bax, bcl-2 and bcl-x) was compared in estrogen-stimulated uteri from control versus neonatally DES-treated hamsters. According to Northern blot analysis of total uterine RNA, levels of the 3.2-kb c-jun and 2.4-kb c-myc transcripts were not altered by neonatal DES treatment. However, the 1.0 kb bax and 2.7 kb bcl-x transcript levels were significantly increased in the neonatally DES-exposed uteri. According to immunohistochemical analysis of paraformaldehyde-fixed and paraffin-embedded tissue sections, levels of c-Jun, c-Fos, c-Myc, Bax, and Bcl-x proteins were enhanced dramatically in both the luminal and glandular epithelial cells of neonatally DES-exposed uteri. In contrast, the immunostaining signal for Bcl-2 protein was decreased consistently in the epithelial cells of neonatally DES-exposed uteri. In conclusion, neonatal DES treatment induced persistent and epithelial cell-specific imbalances in the estrogen-regulated uterine expression of c-jun, c-fos, c-myc, bax, bcl-2, and bcl-x proto-oncogenes. These imbalances likely play a role in the molecular mechanism by which neonatal DES treatment induces altered estrogen responsiveness including hyperplasia, apoptosis, and ultimately neoplasia in the epithelial compartment of the hamster uterus. PMID: 9101088 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 431: Cell Growth Differ. 1997 Apr;8(4):371-80. Transformation by v-Jun prevents cell cycle exit and promotes apoptosis in the absence of serum growth factors. Clark W, Gillespie DA. Beatson Institute for Cancer Research, Cancer Research Campaign Beatson Laboratories, Bearsden, Glasgow, United Kingdom. To gain insight into the mechanism of action of the v-Jun oncoprotein, we compared the growth and cell cycle behavior of normal and v-Jun-transformed fibroblasts. We show that v-Jun induces marked alterations in cell cycle regulation in both the presence and absence of serum growth factors. During asynchronous growth, v-Jun-transformed fibroblasts divide more rapidly than their normal counterparts, owing to a reduction in the length of the G1 phase of the cell cycle. When deprived of serum mitogens, normal fibroblasts exit the cycle and enter a reversible state of quiescence (G0). In contrast, v-Jun-transformed fibroblasts continue to cycle and maintain increased levels of retinoblastoma tumor suppressor protein phosphorylation and elevated expression of cell cycle-dependent markers such as cyclin A, cyclin-dependent protein kinase 2 (CDK2), and CDC2. v-Jun-transformed fibroblasts nevertheless remain wholly dependent on growth factors for cell multiplication, because cell cycle progression in the absence of serum is accompanied by high rates of apoptotic cell death. We conclude that v-Jun shares the capacity of the Myc, E1A, and E2F oncoproteins to promote both cell cycle progression and apoptosis under conditions of mitogen depletion. PMID: 9101083 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 432: Hepatology. 1997 Apr;25(4):874-83. Hepatocarcinogenesis in woodchuck hepatitis virus/c-myc mice: sustained cell proliferation and biphasic activation of insulin-like growth factor II. Liu P, Terradillos O, Renard CA, Feldmann G, Buendia MA, Bernuau D. Laboratoire de Biologie Cellulaire, INSERM U 327, Faculte de Medecine X. Bichat, Paris, France. Transgenic mice carrying the c-myc oncogene under control of woodchuck hepatitis virus (WHV) DNA sequences invariably develop hepatocellular carcinoma (HCC), despite a temporally limited expression of the transgene in the neonatal liver. To better characterize the different steps of the tumorigenic process, we analyzed the liver expression of the c-myc transgene and several growth-related genes by in situ hybridization and Northern blotting. In parallel studies, proliferated changes were investigated by detection of bromodeoxy-uridine-positive S-phase nuclei and apoptosis was evaluated by in situ nick end-labeling of DNA. During the neonatal period, high levels of c-myc messenger RNAs (mRNAs) were detected in all hepatocytes, and the expression of insulin-like growth factor II (IGF II) was frequently enhanced, correlating with increased cell proliferation. Despite elevated expression of the p53 gene, no change in liver cell apoptosis was observed. After weaning, c-myc transgene expression decreased to undetectable levels in all hepatocytes, whereas proliferation decreased but remained notably higher than in age-matched controls. The expression of c-fos, c-jun, and c-H-ras was highly variable during the preneoplastic period and in the tumors, with no consistent increase compared with controls. Resurgence of c-myc transgene expression was evidenced in all cells from hyperplastic lesions and carcinomas, accompanied with frequent focal reactivation of IGF II. Thus the strong proliferative stimulus induced by the combined effects of c-myc and IGF II in the neonatal liver might initiate a process characterized by persistent, dysregulated hepatocyte proliferation, in turn greatly increasing the risk of hepatocellular transformation. PMID: 9096591 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 433: Pflugers Arch. 1997 Apr;433(6):827-31. Lack of control by immediate early response genes in the oxygen regulation of erythropoietin gene expression. Gess B, Wolf K, Kurtz A. Physiologisches Institut, Universitat Regensburg, Germany. This study sought to investigate the role of immediate early genes in the stimulation of erythropoietin (EPO) gene expression by hypoxia. To this end freshly isolated rat hepatocytes were exposed to either normoxia (20% oxygen) or to hypoxia (1% oxygen) and the mRNA levels of the early genes c-fos, c-jun, c-myc and EGR-1 were monitored together with EPO mRNA. Isolation of the cells from the livers strongly stimulated the expression of c-fos, c-jun, c-myc and of EGR-1, whilst EPO gene expression remained unchanged. Exposure of the isolated hepatocytes to hypoxia did not further change early gene expression when compared with cells kept under normoxic conditions. EPO mRNA increased time dependently with a delay of 1 h after onset of hypoxia. These findings suggest that even strong activation of early gene expression has no influence on EPO gene expression, whilst activation of EPO gene expression during hypoxia can happen without change of early gene expression. It appears, therefore, as if immediate early genes are not causally involved in the sequence of events by which hypoxia stimulates EPO gene expression. PMID: 9049176 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 434: Oncogene. 1997 Mar 6;14(9):1109-16. Suppression in transformed avian fibroblasts of a gene (CO6) encoding a membrane protein related to mammalian potassium channel regulatory subunits. Oberst C, Weiskirchen R, Hartl M, Bister K. Institute of Biochemistry, University of Innsbruck, Austria. Gene expression patterns in normal and v-myc-transformed quail embryo fibroblasts were compared by mRNA differential display. Displaying approximately 2500 mRNA species by reverse transcription/PCR, reamplification of 73 differential cDNA fragments and rescreening by Northern analysis led to the isolation of a clone, termed CO6, that hybridized to an mRNA species present only in the normal but not in the transformed fibroblasts. Further analyses revealed that the 0.95-kb CO6 mRNA was present in all normal quail and chicken embryo fibroblasts tested, but that it was undetectable in a variety of established quail cell lines transformed by the v-myc, v-myc/v-mil, v-jun/junD or v-src oncogenes or by a chemical carcinogen. Furthermore, CO6 mRNA was not detectable in fibroblasts newly transformed by retroviral constructs carrying v-myc or v-jun alleles or by the avian sarcoma virus ASV17. In fibroblasts transformed by a temperature-sensitive v-src mutant, expression of CO6 was strongly induced at the non-permissive temperature and reduced at the permissive temperature. Nucleotide sequence analysis of quail CO6 cDNA indicated that the corresponding gene encodes a 200-amino acid protein with 46 to 48% amino acid sequence identity to the regulatory beta subunits (K(VCa)beta) of the bovine, human and canine high conductance Ca2+-activated K+ channels. No sequence homology to other ion channel subunits or to any other proteins in the databases was found. Like the K(VCa)beta subunits, the CO6 protein contains two putative transmembrane segments. Based on the relationship to mammalian K(VCa)beta both in primary structure and domain topology, the CO6 protein may represent the regulatory subunit of a yet unidentified avian Ca2+-activated potassium channel or a related membrane protein possibly involved in the regulation of cell proliferation. PMID: 9070660 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 435: Expert Opin Investig Drugs. 1997 Mar;6(3):279-98. Phospholipase A2 inhibitors in development. Tibes U, Friebe WG. Department of Preclinical Research, Boehringer Mannheim GmbH, Germany. To date, three isoforms of phospholipase A2 (PLA2) have been identified. Of these, the two Ca2+-dependent isoforms, secretory (sPLA2) and cytosolic phospholipase A2 (cPLA2), are targets for new anti-inflammatory drugs. The catalytic mechanisms and functions of the third isoform, Ca2+-independent cytosolic phospholipase A2 (iPLA2), are unknown at present. sPLA2 and cPLA2 are both implicated in the release of arachidonic acid and prophlogistic lipid mediators. However, recent findings provide evidence that cPLA2 is the dominant isoform in various kinds of inflammation, such as T-cell-mediated experimental arthritis. A triple function of PLA2-derived lipid mediators has been suggested: causing immediate inflammatory signs, involvement in secondary processes, e.g., superoxide free radical (O2) generation, apoptosis, or tumour necrosis factor-alpha (TNF-alpha)-cytotoxicity, and controlling the expression and activation of pivotal proteins implicated in inflammation and cell development, e.g., cytokines, adhesion proteins, proteinases, NF-kappaB, fos/jun/AP-1, c-Myc, or p21ras. In the past, research predominantly focused on the development of sPLA2 inhibitors; however, present techniques enable discrimination of cPLA2, sPLA2, and iPLA2, and specific inhibitors of each of the three isoforms are likely to appear soon. Over the last decade, between 40 and 50 sPLA2 inhibitors have been described; and the list is growing. However, of these, few have the potential for clinical success, and those that do are predominantly active site-directed inhibitors, e.g., BMS-181162, LY311727, ARL-67974, FPL67047, SB-203347, Ro-23-9358, YM-26734, and IS-741. At present, there are no likely clinical candidates emerging from the ranks of cPLA2 and iPLA2 inhibitors in development. Indications for which PLA2 inhibitors are being pursued include, sepsis, acute pancreatitis, inflammatory skin and bowel diseases, asthma, and rheumatoid arthritis. The three main obstacles to the successful development of PLA2 inhibitors include, insufficient oral bioavailability, low affinity for the enzyme corresponding to low in vivo efficacy and insufficient selectivity. PMID: 15989628 [PubMed] --------------------------------------------------------------- 436: Placenta. 1997 Mar-Apr;18(2-3):163-8. The low proliferation rates of human amniotic cells are neither associated to deregulated proto-oncogenes' expression nor to the effect of IFN alpha 2. Oliveira JG, Kroon EG, Ferreira PC, Bonjardim CA. Departamento de Microbiologia, Instituto de Ciencias Biologicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil. Primary cultures of human amniotic membrane (PCHAM) cells display very low proliferation rates while their doubling times vary between 150 h and 210 h even after mitogenic stimuli. However, the pattern of proto-oncogenes (c-fos, c-myc and c-jun) expression in these cells, upon serum restimulation, resembled that of cell lines that display shorter population doubling times. Serum stimulation of quiescent PCHAM cells promoted a rapid and transient c-fos mRNA expression, which was detected within 10 min, reached maximal levels at 30 min and decreased to undetectable levels 2-3 h later. The levels of c-myc or c-jun mRNA increased within 10 min after serum restimulation, peaked at 3 h and decreased to intermediate levels thereafter. We also present evidence showing that IFN alpha 2 treatment of PCHAM cells had no effect on their population doubling times nor in c-fas, c-myc, or c-jun mRNA expression, under conditions in which induction of IFN-stimulated genes, such as 2'-5' oligo-adenylate synthetase (OAS) and 6-16 was observed. We conclude that the growth constraints observed with this cells are not directly associated with a negative cellular growth regulation exerted by IFN alpha 2, nor due to a deregulated proto-oncogenes' expression. PMID: 9089777 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 437: J Rheumatol. 1997 Mar;24(3):430-5. In situ expression of protooncogenes and Fas/Fas ligand in rheumatoid arthritis synovium. Asahara H, Hasunuma T, Kobata T, Inoue H, Muller-Ladner U, Gay S, Sumida T, Nishioka K. Division of Rheumatology and Immunology, Institute of Medical Science, St. Marianna University School of Medicine, Kawasaki, Japan. OBJECTIVE: To examine the relationship among the expression of protooncogenes such as c-fos and c-myc, Fas antigen, Fas ligand, and apoptosis in the synovial tissue of patients with rheumatoid arthritis (RA). METHODS: The expression of c-fos, c-myc, Fas antigen, and Fas ligand was examined in synovial tissues of 6 patients with RA and 4 with osteoarthritis (OA) using in situ reverse transcriptase (RT) assay and immunohistochemical staining. Apoptosis was detected by TUNEL method in situ. RESULTS: Expression of protooncogenes, c-fos, and c-myc was detected in all samples from patients with RA, but in only a few cells of OA synovium. 30 to 90% of cells in RA synovium positive for these protooncogenes also coexpressed Fas antigen. Fas positive cells in RA synovium underwent apoptosis to a significant degree. Fas ligand mRNA was detected only in mononuclear cells in RA synovium. CONCLUSION: The expression of protooncogenes is closely related to Fas mediated apoptosis in RA synoviocytes. PMID: 9058644 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 438: Hepatology. 1997 Mar;25(3):607-12. Effect of sialoadenectomy and epidermal growth factor administration on liver regeneration after partial hepatectomy. Lambotte L, Saliez A, Triest S, Maiter D, Baranski A, Barker A, Li B. Laboratory of Experimental Surgery, University of Louvain Medical School, Brussels, Belgium. Epidermal growth factor (EGF), a mitogen in vitro for hepatocytes, produces in various cell lines changes similar to those observed very rapidly in hepatocytes after partial hepatectomy (PH). These changes include ion movements, membrane hyperpolarization and proto-oncogene expression. A stimulatory effect of EGF on liver regeneration can therefore tentatively be associated with the events occurring within the first 3 hours after a PH, sometimes referred to as the "priming phase." To assess this hypothesis, we examined in Wistar rats the effect of EGF deprivation produced by sialoadenectomy (SX) performed before or after a PH of 70%. SX at the time of PH significantly decreased the 3H-thymidine uptake in the DNA 24 hours later (147 +/- 14 DPM per microgram of DNA, mean +/- SE) compared with a simple PH (322 +/- 16; P < .01), but also compared with results obtained when PH is combined with a sham sialoadenectomy (SSX) or in rats pair-fed with the sialoadenectomized rats. This incomplete inhibition was confirmed by a decreased rise in thymidine kinase (TK) activity and by reduced proliferating cell nuclear antigen (PCNA) labeling and mitotic indices 30 hours after PH. By contrast, SX did not inhibit the early expression of c-jun and c-fos, or of c-myc, 30 or 120 minutes after PH, respectively. A reduction of DNA synthesis was also obtained when SX was performed 3 hours after PH (127 +/- 15 DPM per microgram of DNA vs. 350 +/- 21 in SSX; P < .001) but not when SX was delayed until the 6th or the 17th hour after PH. It was sufficient to administer EGF (40 microg) from the third to the ninth hour to correct the reduction of [3H]thymidine uptake in rats sialoadenectomized before PH. These results indicate that the diminished EGF availability following SX decreases or at least delays liver regeneration, and that the effect of EGF on liver regeneration does not seem related to the early changes of proto-oncogene expression, but rather to events occuring later, at the time of reported internalization and binding of EGF to its nuclear receptors. PMID: 9049206 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 439: Oncogene. 1997 Feb 20;14(7):857-63. Increased expression of c-fos, c-jun and LRF-1 is not required for in vivo priming of hepatocytes by the mitogen TCPOBOP. Columbano A, Ledda-Columbano GM, Pibiri M, Piga R, Shinozuka H, De Luca V, Cerignoli F, Tripodi M. Istituto di Patologia Sperimentale, Universita di Cagliari, Italy. The notion that an increased expression of immediate early genes such as c-fos and c-jun is an absolute requirement for the G0-G1 transition of the hepatocytes has recently been challenged by the finding that rat liver cell proliferation induced by primary mitogens may occur in the absence of such changes (Columbano and Shinozuka, 1996). To further investigate the relationship between immediate early genes and hepatocyte proliferation, we have compared the hepatic levels of c-fos, c-jun and LRF-1 transcripts during mouse liver cell proliferation in two conditions: (i) direct hyperplasia induced by the non-genotoxic hepatocarcinogen 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene, and (ii) compensatory regeneration caused by a necrogenic dose of carbon tetrachloride. The results show striking differences in the activation of early genes. In spite of a rapid stimulation of S phase by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (approximately 8% of hepatocytes were BrdU-positive as early as 24 h after mitogen treatment versus 1% of labelled hepatocytes after 2/3 partial hepatectomy), no changes in the expression of c-fos, c-jun and LRF-1 could be observed. Moreover, no change in steady state mRNA hepatic levels of IGFBP-1 (a gene highly expressed in rat liver following partial hepatectomy), and only a slight increase in c-myc and PRL-1, was found after mitogen administration. On the contrary, a rapid, massive and transient increase in the hepatic mRNA levels of all these genes was observed during carbon tetrachloride induced regeneration. The results indicate that increased expression of immediate early genes may be dependent upon the nature of the proliferative stimulus, and it may not be a prerequisite in certain in vivo conditions such as proliferation induced in the absence of liver tissue damage. PMID: 9047393 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 440: Blood. 1997 Feb 15;89(4):1197-206. Characterization of cis-acting sequences and trans-acting signals regulating early growth response 1 and c-fos promoters through the granulocyte-macrophage colony-stimulating factor receptor in BA/F3 cells. Watanabe S, Kubota H, Sakamoto KM, Arai K. Department of Molecular and Developmental Biology, Institute of Medical Science, The University of Tokyo, Japan. Human granulocyte-macrophage colony-stimulating factor (hGM-CSF) activates a set of genes such as c-fos, jun, myc, and early growth response gene 1 (egr-1). Studies on BA/F3 cells that express hGM-CSF receptor (hGMR) showed that two different signaling pathways controlled by distinct regions within the beta subunit are involved in activation of c-fos/c-jun genes and in c-myc, respectively. However, the region(s) of the beta subunit responsible for activation of the egr-1 gene and other regulatory genes has not been identified. We describe here how egr-1 promoter is activated by hGMR through two regions of the beta subunit, with these regions being required for activation of the c-fos promoter. Coexpression of dominant negative (dn) Ras (N17ras) or dn JAK2 almost completely suppressed the activation of egr-1 and c-fos promoters. Deletion analysis of egr-1 promoter showed two cis-acting regions responsible for activation by hGM-CSF or mouse interleukin-3 (mIL-3), one between nucleotide positions (nt) -56 and -116, and the other between nt -235 and -480, which contains tandem repeats of the serum response element (SRE) sites. Similar experiments with the c-fos promoter showed that cis-acting regions containing the SRE/AP-1 sites is sufficient for activation by hGM-CSF. Based on these observations, we propose that signaling pathways activating egr-1 and c-fos promoters are controlled by SRE elements, either through the same or overlapping pathways that involve JAK2 and Ras. PMID: 9028942 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 441: J Biol Chem. 1997 Feb 7;272(6):3813-22. Detection and functional characterization of p180, a novel cell cycle regulated yeast transcription factor that binds retinoblastoma control elements. Cuevo RS, Garrett S, Horowitz JM. Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA. In recent years it has become apparent that the cellular machinery governing cell cycle progression and transcription control are often homologous in yeast and mammalian cells. We and others have previously shown that the SP family of mammalian transcription factors regulates the transcription of a number of genes whose activities are governed by the product of the retinoblastoma (Rb) susceptibility gene, including c-FOS, c-MYC, TGFbeta-1, IGF-II, and c-JUN. To determine whether a similar pathway of transcriptional regulation may function in yeast, we explored the possibility that transcription factors with nucleotide-binding specificities akin to those of the SP family are expressed in Saccharomyces cerevisiae and Schizosaccharomyces pombe. Here we report the detection of novel yeast proteins (S. cerevisiae, p180; S. pombe, p200) that specifically bind Rb-regulated promoter elements in vitro dependent on nucleotides that are also required for binding and trans-activation by SP family members in vivo. Our results indicate that the S. cerevisiae retinoblastoma control element-binding activity 1) requires zinc for association with DNA; 2) does not bind to SCB, MCB, or E2F sites in vitro; 3) is cell cycle-regulated in a SWI6-independent fashion; and 4) maximally stimulates retinoblastoma control element-mediated transcription in early- to mid-S phase. Taken together, these data suggest that p180 may regulate the transcription of a subset of yeast genes whose expression is coincident with the onset and/or progression of DNA replication. PMID: 9013640 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 442: Jpn J Cancer Res. 1997 Feb;88(2):143-51. Correlated expression of glutathione S-transferase-pi and c-Jun or other oncogene products in human squamous cell carcinomas of the head and neck: relevance to relapse after radiation therapy. Miura K, Suzuki S, Tanita J, Shinkawa H, Satoh K, Tsuchida S. Department of Otorhinolaryngology, Hirosaki University School of Medicine. The expression of glutathione S-transferase (GST)-pi and four oncogene products, c-Jun, c-Fos, c-H-Ras, and c-Myc, in human squamous cell carcinomas of the head and neck was investigated immunohistochemically before and after radiation therapy, to examine whether these oncogene products might be involved in GST-pi expression, and also to examine the relationship between their expression and therapeutic response. Clinical response to radiation was evaluated in terms of both tumor regression and relapse over two-year follow-up periods. The overall positive rates in 83 carcinoma specimens before therapy were 60.2% for GST-pi and 28.9-51.8% for the individual oncogene products, the positive rates for the oncogene products being higher in GST-pi-positive than in GST-pi-negative cancers. c-Jun was most highly correlated with GST-pi expression. Following radiation, the expression of GST-pi and the oncogene products was altered in about a half of 30 patients. Eleven of the 18 patients who exhibited prior positivity for GST-pi showed negative conversion, while 4 of the 12 patients with prior negativity demonstrated positive conversion. In most cases, changes in c-Jun staining coincided with those in GST-pi. Regarding clinical response to radiation therapy, the positive rates for GST-pi and c-Jun before radiation were higher in the residual cancer or relapse cases than in the group showing complete response without relapse. Examination of 26 patients with laryngeal cancer revealed that relapse occurred more frequently in cases exhibiting positive reactions for GST-pi, c-Jun, or c-H-Ras. These results suggest a direct link between c-Jun and GST-pi in head and neck cancers before and after radiation. Although GST-pi and the oncogene products can be influenced by radiation, GST-pi and c-H-Ras expression may be a risk factor for relapse of laryngeal cancer. PMID: 9119742 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 443: Biochem Mol Biol Int. 1997 Feb;41(2):279-84. Detection of apoptosis in the brain of the zitter rat with genetic spongiform encephalopathy. Ookohchi T, Ito H, Serikawa T, Sato K. Department of Molecular Biology, Faculty of Medicine, Tottori University, Yonago, Japan. Zitter rat develops genetic spongiform encephalopathy accompanied with whole-body tremors and flaccid paresis. To elucidate the mechanism of a neuronal cell death in the brain, we determined involvement of apoptosis in this rat. By Northern blot analysis, the elevation of mRNA levels were observed in c-jun, c-fos, c-myc and p53 genes which were induced by apoptotic signals: conversely, expression of bcl-2 was shown to be decreased in the zitter rat brain in contrast to the WTC control rat. Furthermore, TUNEL staining of fragmented DNA indicated apoptotic morphology in this brain. These results strongly suggested that the spongiform encephalopathy of the zitter rat was due to apoptosis in the brain cells. PMID: 9063567 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 444: AIDS Res Hum Retroviruses. 1997 Jan 20;13(2):151-9. Erratum in: AIDS Res Hum Retroviruses 1997 May 1;13(7):633. V3 loop of human immunodeficiency virus type 1 suppresses interleukin 2-induced T cell growth. Sakaida H, Murakami T, Kawamata S, Hattori T, Uchiyama T. Laboratory of AIDS Immunology, Research Center for Immunodeficiency Virus, Institute for Virus Research, Kyoto University, Japan. We tested the effect of three linear or two loop peptides derived from the V3 region of the HTLV-III BH10 clone or the SF2 strain of human immunodeficiency virus type 1 on IL-2-driven T cell proliferation. V3-BH10, which consists of 42 amino acids and has a loop structure, suppressed IL-2-driven proliferation of all IL-2-dependent cells [Kit225, ED-40515(+), KT-3, 7-day PHA-blasts, and fresh peripheral blood mononuclear cells] tested, whereas it did not suppress the cell growth of IL-2-independent cell lines (Hut102, Molt-4, and Jurkat). This suppressive effect was also seen in IL-2-driven cell growth of CD8-positive lymphocytes purified from 7-day PHA-blasts, indicating that CD4 molecules were not required for the suppression. The treatment with anti-V3 loop monoclonal antibody (902 antibody) completely abolished the suppressive effect of V3-BH10. In addition, V3-BH10 generated the arrest of Kit225 cells and also purified CD8-positive lymphocytes in G1 phase in the presence of IL-2. Neither chromatin condensation nor DNA fragmentation was detected in Kit225 cells cultured with V3-BH10 and IL-2. V3-BH10 neither blocked radiolabeled IL-2 binding to IL-2 receptors nor affected tyrosyl phosphorylation of several cellular proteins (p120, p98, p96, p54, and p38), which is immediately induced by IL-2 stimulation. However, V3-BH10 enhanced IL-2-induced mRNA expression of c-fos but not c-myc or junB. Thus, the binding of V3 loop of gp120 to the cell surface molecule(s) appears to affect intracellular IL-2 signaling, which leads to the suppression of IL-2-induced T cell growth. PMID: 9007200 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 445: Cancer Lett. 1997 Jan 15;112(1):65-9. Selective hyperexpression of c-jun oncoprotein by glass fiber- and silica-transformed BALB/c-3T3 cells. Gao H, Brick J, Ong S, Miller M, Whong WZ, Ong T. West Virginia University, Morgantown 26506, USA. Mining and mineral processing are important industries in the United States. A large number of workers are potentially exposed to silica during mining and to glass fibers during manufacturing. There is a concern regarding lung cancer risk among workers exposed to silica and glass fibers. Our previous studies showed that both glass fibers and silica induced transformation of BALB/c-3T3 cells. In order to explore the relationship between silica and glass fiber-induced cell transformation and oncoprotein expression, the protein products of seven proto-oncogenes (c-K-ras, c-H-ras, c-sis, c-myc, c-myb, c-erb B1 and c-jun) and one tumor suppressor gene (p53) were examined in BALB/c-3T3 cells transformed by glass fibers or silica using immunoblotting with specific monoclonal or polyclonal antibodies. The results showed that all transformants, including eight induced by glass fibers and eight by silica (Min-U-Sil 5), were positive for c-jun protein expression; the level of c-jun protein was elevated 8-21-fold in these transformants. Other protooncogene proteins in transformed cells were either not detectable or not different from non-transformed cells. These results suggest that the overexpression of c-jun is common in BALB/c-3T3 transformed cells induced by glass fibers or silica. It seems, therefore, that the expression of c-jun may play an important role in the transformation process. PMID: 9029170 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 446: Cancer Lett. 1997 Jan 15;112(1):57-63. Induction of lung carcinogenesis in AKR-mice by N-nitrosodiethylamine/phenobarbitone, associated with high expression of c-myc and c-jun oncoproteins. Giri RK, Baral RN, Das BR. Molecular Biology Division, Institute of Life Sciences, Sahid Nagar, Bhubaneswar, India. Lung carcinogenesis was induced in AKR mice using N-nitrosodiethylamine (NDEA). Tumors were detected in 46.8% of mice provided with 100 ppm NDEA in drinking water. The incidence of tumors was increased to 64.2% when the same carcinogenesis was promoted by phenobarbitone (PB). Lung tumor bearing mice showed no tumors in other organs. Characteristic features of these lung tumors are: (i) appearance of tumors within a short period of time i.e. less than 75 days; (ii) no increase in the number and size of tumors with the increase in dose and duration of treatment of carcinogen; (iii) the same histological type was maintained in more than 80% of tumors. Animals that received treatment for 75-125 days showed no significant advancement in the stage of carcinogenesis in comparison to the 50-75 days treatment period. Moreover, mice which received treatment for 125-150 days, did not have any neoplastic lesions in lungs, but they consisted of liver tumors generally. Expression of oncoproteins, c-myc and c-jun, was detected in all lung tumors but the expression of c-myc protein was more than that of c-jun and both of these oncoproteins were enhanced by the promoter, PB. Highest level of expression of c-myc and c-jun was detected within the period of 50-75 days, whereafter it was decreased significantly within the period of 75-125 days and 125-150 days of treatment. Thus, the results indicate that c-myc/c-jun might be involved in the development of lung cancer in AKR mice, but may not have any role in the maintenance of the malignant phenotype of lungs. PMID: 9029169 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 447: Oncol Res. 1997;9(4):205-11. Induction of proto-oncogenes during 3-deazaadenosine-stimulated differentiation of 3T3-L1 fibroblasts to adipocytes: mimicry of insulin action. Zeng G, Dave JR, Chiang PK. Department of Biochemistry and Molecular Biophysics, Medical College of Virginia, Richmond 23298-0614, USA. 3-Deazaadenosine (DZA) mimicked the molecular action of insulin in the induction of 3T3-L1 fibroblasts to differentiate into adipocytes. The molecular effects of DZA were compared with insulin, which served as a positive control, on the expression of proto-oncogenes during the initial stage of differentiation of 3T3-L1 fibroblasts. Treatment of confluent 3T3-L1 fibroblasts with DZA or insulin produced a rapid but-transient expression of mRNA for proto-oncogenes c-fos and c-jun within 30-60 min. The mRNA of c-myc increased for 2 h and then decreased 4 h after treatment. Electrophoretic mobility shift assays showed a heightened increase in the appearance of transcription factors AP-1 and AP-2. The increase was detectable as early as 1 h after the treatment with either DZA or insulin and was maintained for 6 h. 3T3-L1 cells stably transfected with the promotor of c-fos linked to a CAT reporter gene showed an increase in CAT activity in response to DZA in a time- and dose-dependent manner. In cells stably transfected with antisense c-fos, neither DZA nor insulin was able to induce a differentiation response. The early transcription of c-fos, c-jun, and c-myc proto-oncogenes and the increased expression of transcription factors AP-1 and AP-2, induced by DZA and insulin, appear to be crucial events in the differentiation of the 3T3-L1 fibroblasts to adipocytes. PMID: 9268991 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 448: J Cancer Res Clin Oncol. 1997;123(5):267-71. Effect of the cytostatic butyric acid pro-drug, pivaloyloxymethyl butyrate, on the tumorigenicity of cancer cells. Aviram A, Rephaeli A, Shaklai M, Nudelman A, Ben-Dror I, Maron L, Rabizadeh E. Institute of Hematology, Rabin Medical Center, Tel Aviv University, Israel. Previously we have shown that pivaloyloxymethyl butyrate (AN-9), a pro-drug of butyric acid (BA), is a differentiation-inducing agent in a variety of cells. In this report, we demonstrate that AN-9 is a cytostatic but not cytotoxic agent in a myelomonocytic cell line (WEHI); thus, the cells were growth-arrested and differentiated. These late changes in the cells were preceded by changes in the expression of the early regulatory genes, c-myc and c-jun. Although initiation of all these events had already occurred after 1 h exposure to AN-9, the tumorigenicity of these cells tested in Balb/c mice was not affected. A marked reduction in the tumorigenicity of AN-9-treated cells was observed after 4 h of exposure. Exposure of the highly metastatic subclone of Lewis lung carcinoma (3LLD122) to AN-9 resulted in a very pronounced effect on the tumorigenicity of these cells tested in C57BL mice. Unlike WEHI cells, the tumorigenicity of 3LLD122 was almost completely diminished after 1 h of exposure. In both cell types a 10-fold higher concentration of BA did not affect the tumorigenicity of the cells as did AN-9. PMID: 9201249 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 449: J Steroid Biochem Mol Biol. 1997 Jan;60(1-2):99-104. Immunocytochemical localization of C-myc and C-jun oncoproteins in hamster kidney and estrogen-induced kidney tumors. Bhat HK, Springer I, Rajaraman S, Liehr JG. Department of Pharmacology, University of Texas Medical Branch, Galveston 77555-1031, U.S.A. The chronic administration of 17beta-estradiol to male Syrian hamsters for 6-7 months induces kidney tumors which express high levels of c-fos, c-myc and c-jun mRNA compared to surrounding tissue or untreated controls. In this study, we have investigated, by immunocytochemical methods, the cellular localization of c-myc and c-jun oncoproteins in estrogen-dependent kidney tumors, in kidney tissue of hamsters treated with 17beta-estradiol for 6 months and in the kidneys of age-matched controls. The c-myc oncoprotein was strongly expressed in tumors, in smooth muscle layers of arteries and in parietal epithelial cells of the glomerulus. In age-matched untreated kidneys, there was little or no staining in the glomerulus, arteries or kidney tubular cells. The c-jun oncoprotein was detected in kidney tumors and in the tubular epithelium of surrounding tissue. The immunoreactivity for c-jun oncoprotein was highest in the tumor, intermediate in estrogen-treated kidney tissue and lowest in kidney tubular cells of controls. It is concluded that the high expression of c-myc in estrogen-induced kidney tumors, in the smooth muscle layer of arteries, and in glomerular parietal epithelial cells in the kidneys of 17beta-estradiol-treated hamsters, but poor expression in control kidneys indicate an involvement of this oncoprotein in the tumorigenic process. In contrast, c-jun is expressed in untreated, in 17beta-estradiol-treated kidneys and in tumors, and may not serve as a prognostic marker in the transformation of these cells to the malignant phenotype. PMID: 9182863 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 450: Curr Opin Oncol. 1997 Jan;9(1):79-87. Role of oncogenes in resistance and killing by cancer therapeutic agents. el-Deiry WS. Howard Hughes Medical Institute, Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104, USA. Chemotherapeutic drug resistance is a major clinical problem and cause for failure in the therapy of human cancer. One of the goals of molecular oncology is to identify the underlying mechanisms, with the hope that more effective therapies can be developed. Several mechanisms have been suggested to contribute to chemoresistance: 1) amplification or overexpression of the P-glycoprotein family of membrane transporters (eg, MDR1, MRP, LRP) which decrease the intracellular accumulation of chemotherapy; 2) changes in cellular proteins involved in detoxification (eg, glutathione S-transferase pi, metallothioneins, human MutT homologue, bleomycin hydrolase, dihydrofolate reductase) or activation of the chemotherapeutic drugs (DT-diaphorase, nicotinamide adenine dinucleotide phosphate:cytochrome P-450 reductase); 3) changes in molecules involved in DNA repair (eg, O6-methylguanine-DNA methyltransferase, DNA topoisomerase II, hMLH1, p21WAF1/CIP1; 4) activation of oncogenes such as Her-2/neu, bcl-2, bcl-XL, c-myc, ras, c-jun, c-fos, MDM2, p210 BCR-abl, or mutant p53. An overview of these resistance mechanisms is presented, with a particular focus on the role of oncogenes. Some current strategies attempting to reverse their effects are discussed. Publication Types: Review Review, Tutorial PMID: 9090498 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 451: J Submicrosc Cytol Pathol. 1997 Jan;29(1):1-17. Estrogen receptor, growth factor receptor and protooncogene protein activities and possible signal transduction crosstalk in estrogen dependent and independent breast cancer cell lines. Mohamood AS, Gyles P, Balan KV, Hollis VW, Eckberg WR, Asseffa A, Han Z, Wyche JH, Anderson WA. Department of Biology, Howard University, Washington, D.C., USA. Binding of estrogen to its receptor (ER) activates early genes that drive responsive cells through the proliferative phase. Earlier studies to evaluate the expression of protooncogenes, growth factors, growth factor receptor and steroid hormone receptor gene activities in the rat uterine system indicated complex pathways that involve significant 'crosstalk' between ER-systems and signal transduction pathways (Bhattacharyya et al., 1994). To analyze the interactions between these factors, we examined two well characterized estrogen dependent (MCF-7) and estrogen independent (MDA-MB-231) human breast cancer cell lines. Antibodies to estrogen receptor, epidermal growth factor receptor, c-Fos, c-Jun, and Ras proteins, protein kinases involved in receptor tyrosine kinase signal transduction pathway, MEK1 and phosphotyrosine were utilized in immunocytochemical localization experiments to evaluate temporal expression of these factors in response to estrogen treatment. ER, which was diminished in MCF-7 cells grown in estrogen-stripped medium, increased 9-fold in estrogen-reconstituted medium by 120 min. Fos and Jun appeared at nuclear and perinuclear cytoplasmic sites within 60 min after estrogen treatment in MCF-7 cells. Fos/Jun proteins were prominent in MDA-MB-231 cells, especially in association with actin filaments. Immunolabeling studies revealed no EGF-r in MCF-7 cells, while MDA-MB-231 cells contained intense EGF-r labeling in the plasma membrane. Ras protein was prominent in the cytoplasm and at the cell surface within 60 min after treatment of MCF-7 cells with estrogen. Ras was intense in MDA cells. Similarly, MCF-7 and MDA cells contained high concentrations of MEK1 and phosphotyrosine (pTyr) containing proteins in their cytoplasm and immunolabeling remained high as long as MCF-7 cells were grown in medium containing estrogen. It is speculated that MEK1 (cytoplasmic) functioning through Fos/Jun or Myc/Max (nuclear) may regulate the activity of AP-1 transcription factor. In all cases however, MEK1 and pTyr protein labeling was more intense in the highly metastatic and hormone independent MDA-MB-231 breast cancer cells. Results revealed signal transduction pathway proteins in ER+ estrogen dependent cells suggesting possible crosstalk between both receptor pathways during the proliferative phase of MCF-7 cells. PMID: 9066137 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 452: Cancer Lett. 1997 Jan 1;111(1-2):225-31. Adriamycin induces apoptosis in rat thymocytes. Azmi S, Bhatia L, Khanna N, Dhawan D, Singh N. Department of Biochemistry, A.I.I.M.S., Ansari Nagar, New Delhi, India. Apoptosis is a controlled form of cell death accompanied by distinct morphological and biochemical changes. In this study the nature of cytotoxicity induced by adriamycin (ADM) in rat thymocytes was evaluated. Morphological and biochemical changes characteristic of apoptosis were found to precede adriamycin-induced cell death. Our findings demonstrate the involvement of c-Myc, c-Jun, antioxidant enzymes CuZn superoxide dismutase and catalase, and perhaps poly ADP ribosylation in ADM-induced cell death. PMID: 9022151 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 453: Mol Reprod Dev. 1997 Jan;46(1):11-8. Regulation of cell cycle entry and G1 progression by CSF-1. Roussel MF. Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA. Proliferation, differentiation, and survival of monocytes, macrophages, and their immediate progenitors is regulated by the macrophage colony-stimulating factor (CSF-1). CSF-1 initiates a mitogenic response by binding to its receptor (CSF-1R), thereby activating the receptor's intrinsic tyrosine kinase activity and initiating signaling via multiple effector-mediated pathways. CSF-1 is required throughout G1 to ensure entry of bone marrow-derived macrophages into S phase, and persistent CSF-1R kinase activity is necessary to the expression of both immediate early (e.g., c-fos, c-jun, and c-myc) and delayed early (e.g., D-type cyclins) response genes. Ectopic expression of human CSF-1R in different mouse cell lines, including fibroblasts, IL-3-dependent myeloid cells, and early pre-B cells, confers CSF-1 responsiveness by replacing the cells' requirements for other mitogenic growth factors. NIH-3T3 fibroblasts engineered to express a human CSF-1 receptor point mutant (CSF-1R [Y809F]) fail to proliferate in response to CSF-1 and remain arrested in the early G1 phase of the cell cycle. Despite CSF-1-dependent transcription of fos and jun family members, c-myc, D-type, and E-type G1 cyclin mRNAs are not expressed in the latter cells in response to growth factor stimulation. However, enforced expression of c-myc or D-type cyclins, but not cyclin E, resensitizes cells bearing CSF-1R (Y809F) to the mitogenic effects of CSF-1, enabling them to proliferate continuously in liquid culture and to form colonies in agar in response to the growth factor. D-type cyclin mutants defective in binding to the retinoblastoma protein (pRB) were unable to rescue mutant receptor signaling, suggesting that the ability of D-type cyclin-dependent kinases to cancel pRB's growth-suppressive function is necessary for CSF-1-induced G1 exit. By contrast, cyclin E must function in a different pathway. Cells rescued by c-myc were prevented from entering S phase by microinjection of antibodies to cyclin D1. Conversely, cyclin D1-rescued cells were inhibited from forming CSF-1-dependent colonies in agar when challenged with either a dominant-negative c-myc mutant or mad, a transcription factor which competes with myc for max, its requisite heterodimeric partner. Thus, although the expression of c-myc and D-type cyclins is rate limiting for G1 phase progression, their functions are interdependent, with both activities being required for mitogenicity. Publication Types: Review Review, Tutorial PMID: 8981358 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 454: Nutr Cancer. 1997;27(1):92-101. RRR-alpha-tocopheryl succinate inhibits EL4 thymic lymphoma cell growth by inducing apoptosis and DNA synthesis arrest. Yu W, Sanders BG, Kline K. Genetics Institute, University of Texas at Austin 78712-1097, USA. RRR-alpha-tocopheryl succinate (vitamin E succinate, VES) treatment of murine EL4 T lymphoma cells induced the cells to undergo apoptosis. After 48 hours of VES treatment at 20 micrograms/ml, 95% of cells were apoptotic. Evidence for the induction of apoptosis by VES treatments is based on staining of DNA for detection of chromatin condensation/fragmentation, two-color flow-cytometric analyses of DNA content, and end-labeled DNA and electrophoretic analyses for detection of DNA ladder formation. VES-treated EL4 cells were blocked in the G1 cell cycle phase; however, apoptotic cells came from all cell cycle phases. Analyses of mRNA expression of genes involved in apoptosis revealed decreased c-myc and increased bcl-2, c-fos, and c-jun mRNAs within three to six hours after treatment. Western analyses showed increased c-Jun, c-Fos, and Bcl-2 protein levels. Electrophoretic mobility shift assays showed increased AP-1 binding at 6, 12, and 24 hours after treatment and decreased c-Myc binding after 12 and 24 hours of VES treatment. Treatments of EL4 cells with VES+RRR-alpha-to-copherol reduced apoptosis without effecting DNA synthesis arrest. Treatments of EL4 cells with VES+rac-6-hydroxyl-2, 5,7,8-tetramethyl-chroman-2-carboxylic acid, butylated hydroxytoluene, or butylated hydroxyanisole had no effect on apoptosis or DNA synthesis arrest caused by VES treatments. Analyses of bcl-2, c-myc, c-jun, and c-fos mRNA levels in cells receiving VES + RRR-alpha-tocopherol treatments showed no change from cells receiving VES treatments alone, implying that these changes are correlated with VES treatments but are not causal for apoptosis. However, treatments with VES + RRR-alpha-tocopherol decreased AP-1 binding to consensus DNA oligomer, suggesting AP-1 involvement in apoptosis induced by VES treatments. PMID: 8970189 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 455: Mol Cell Biochem. 1996 Dec 6;165(1):55-63. Inhibition of rat parotid gland growth response induced by chronic isoproterenol following treatment with quinolone antibiotics. Kelentey B, Kerr M, Tao Z, Purushotham KR, Humphreys-Beher MG, Zelles T. Clinic of Stomatology, University of Debrecen, Hungary. While antibiotics are broadly used in dental and medical therapy, little attention has been directed towards the potential toxic side effects of antibiotics on tissue regeneration. Here we examined the effect of a quinolone antibiotic, pefloxacin (Rhone Poulenc) on rat parotid gland responses to chronic isoproterenol treatment. Groups of rats received injections of isoproterenol to induce glandular growth, saline (controls), pefloxacin, or isoproterenol and pefloxacin in combination. Parotid gland weight decreased significantly after pefloxacin treatment for 7 days as well as inhibiting glandular enlargement provoked by isoproterenol. The same trend was observed for the rates of DNA synthesis, with the incorporation of [3H]-thymidine in isoproterenol/pefloxacin-treated rats reduced to 49% of isoproterenol treatment alone levels. Saline-treated animals were 42% of the rate of [3H]-thymidine incorporation into DNA observed in isoproterenol treated rats. While isoproterenol treatment increased steady-state mRNA levels for fos, jun, myc, src, c-erbB-2, ras and topo II, inclusion of pefloxacin with the isoproterenol regimen blocked these increases. Pefloxacin treatment by itself did not alter proto-oncogene mRNA levels in the parotid gland. Glandular amylase activity was decreased in the pefloxacin treated group, while the combination of isoproterenol with pefloxacin did not decrease glandular amylase levels to the extent of that observed with beta-agonist treatment alone. In acute experiments, pefloxacin significantly decreased the volume of saliva secreted by the parotid gland. These results suggest that quinolone-based antibiotics disturb the secretory function of the parotid gland and can inhibit cell proliferation and regeneration. PMID: 8974081 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 456: Oncogene. 1996 Dec 5;13(11):2421-30. Activation of JNK/SAPK pathway is not directly inhibitory for cell cycle progression in NIH3T3 cells. Shu J, Hitomi M, Stacey D. The Cleveland Clinic Foundation Research Institute, Department of Molecular Biology, Ohio 44195, USA. In this study the induction of stress activated protein kinase (SAPK) activity by protein synthesis inhibitors was shown not to inhibit cellular proliferation. Anisomycin induced strong SAPK activity at non-inhibitory concentrations for either protein or DNA synthesis, while the other two inhibitors, emetine and cycloheximide, blocked cell cycle progression without strong SAPK induction. With all three inhibitors, the induction of SAPK activity was always accompanied by protein synthesis inhibition to some extent. Stimulation of mRNA expression of the genes c-jun, c-fos and c-myc correlated well with SAPK induction, but not with cell cycle inhibition. With concentrations of each inhibitor able to block DNA synthesis, no induction of message for the cyclin dependent kinase inhibitor waf-1 was observed; while induction of gadd45 message indicated that the cells might be responding to growth-arrest or DNA damage. The inability of microinjected E2F/DP1 transcription factor proteins to overcome the inhibition of DNA synthesis induced by protein synthesis inhibitors indicate that blockage of an early event in cell cycle progression had occurred. These results indicate that the SAPK induction by protein synthesis inhibitors has no proliferative consequences. PMID: 8957084 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 457: Cancer Lett. 1996 Dec 3;109(1-2):121-7. Differential expression of c-jun and c-myc in N-nitroso diethylamine-induced hepatic oncogenesis in AKR mice. Giri RK, Das BR. Molecular Biology Division, Institute of Life Sciences, Sahid Nagar, Bhubaneswar, India. Expression of c-jun, c-myc, c-fos and c-Ha-ras was examined and correlated with the various stages of N-nitrosodiethylamine (NDEA)-induced hepatocarcinogenesis in male AKR mice. The treated groups were given NDEA 100 ppm, in drinking water for 120 days. The histopathology along with the expression of oncogenes were studied at different durations of treatment such as after 1 day, 3 days, 6 days, 9 days, 15 days, 20 days 30 days, 60 days, 90 days and 120 days of treatment. The results of histological investigation indicate mild hyperplasia and anisonucleosis at 30 days of treatment and prominent pathological features from 60 days onwards until the appearance of hepatocarcinoma at 120 days even without the development of any preneoplastic or neoplastic nodule. The results of the Northern blot hybridization clearly indicate an increased expression of c-jun from 15 days onwards. This overexpression of c-jun at such an early stage indicates its association with the events earlier than the neoplastic changes. However, the persistent overexpression of c-jun at all durations of treatment indicates its association with the events during the later stage of hepatocarcinogenesis, whereas c-myc overexpression starts from 30 days of treatment and persists until the end of the experiment, i.e. 120 days of treatment. However, the results of densitometric quantification indicate that the extent of increase expression of c-myc is less than that of c-jun expression until 1 month of treatment, after which the induction of c-myc exceeds the expression of c-jun. Thus, the overexpression of c-myc from 2 months onwards might be playing a critical role in maintenance of the malignant phenotype. On the other hand, the expressions of c-fos and c-Ha-ras do not have any alteration during NDEA treatment. Thus, our data demonstrate the involvement of c-jun and c-myc in NDEA-induced hepatocarcinogenesis in AKR mice. PMID: 9020911 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 458: Eur J Cancer. 1996 Dec;32A(13):2334-41. Examination of multidrug resistance in cell lines and primary breast tumours by flow cytometry. Brotherick I, Shenton BK, Egan M, Cunliffe WE, Browell DA, Lunt LG, Young JR, Higgs MJ. Department of Surgery, Medical School, University of Newcastle upon Tyne, UK. The aim of this study was to measure multidrug resistance (MDR) by flow cytometry and quantify the expression of P-glycoprotein (using antibody) glutathione transferase (using alpha-GSTpi antibody) in alpha-JSB-1 and alpha-GSTpi of a series of cell lines and primary breast cancers, and to assess the relationship between these MDR proteins and a selection of oncogene and prognostic markers in breast cancer. Flow cytometry was performed using permeabilised cells stained with fluorescent antibodies using well-established methods. Antibody staining was confirmed for JSB1, but not GSTpi by use of known positive and negative controls. No correlation was seen when comparing the number of molecules of alpha-JSB-1 with alpha-GSTpi (P = 0.1, r2 = 0.4, n = 14) using a selection of cell lines. Examination of 45 breast tumours for expression of JSB-1 and GSTpi revealed a significant association between these two antibodies (P < 0.00001, r2 = 0.5, n = 45). On examining the breast tumours, alpha-JSB-1 showed a positive association with c-erbB-2 (P = 0.003), c-myc (P = 0.0004) and c-jun (P = 0.02) but not ER or EGF-R expression. alpha-GSTpi showed a positive association with c-erbB-2 (P = 0.03) and c-myc (P = 0.0004) but not ER, EGF-R or c-jun. Flow cytometric MDR levels were not related to tumour grade or axillary node status. In solid tumours, a relationship between the two antibodies used has been clearly demonstrated, however, specificity of alpha-GSTpi is questioned. Both antibodies show an association with c-erbB-2, which is associated with poor prognosis and with c-myc which is involved in cell cycling and differentiation. Monitoring MDR markers (Pgp) using this methodology may be useful for evaluation of prognosis in breast cancer. PMID: 9038618 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 459: Prostaglandins Leukot Essent Fatty Acids. 1996 Dec;55(6):403-11. The induction of apoptosis in human cervical carcinoma (HeLa) cells by gamma-linolenic acid. de Kock M, Lottering ML, Grobler CJ, Viljoen TC, le Roux M, Seegers JC. Department of Physiology, Medical University of Southern Africa. A high concentration (50 micrograms/ml) of gamma-linolenic acid (GLA) induced morphological lesions typical of apoptosis, as well as DNA fragmentation, in HeLa cells. A lower concentration of GLA (20 micrograms/ml), caused an increased proliferating cell nuclear antigen (PCNA) labelling, with 92.7% cells positive, compared to 27.7% at a concentration of 50 micrograms/ml GLA. In correlation with these results, the number of cells with degraded DNA below the G0/G1 peak increased significantly in the 50 micrograms/ml GLA-treated cells, but increased only slightly in cells exposed to the lower level of GLA. The high levels of PCNA induced by 20 micrograms/ml GLA, in both G1 and S phases, may indicate a state of DNA repair synthesis, whilst at the higher concentration of GLA, most of the cells became apoptotic. Since apoptosis is associated with the deregulation of c-Myc expression, and as the Raf-1-MAP kinase cascade activates the expression of c-Myc and c-Jun, we investigated the effects of 20 and 50 micrograms/ml GLA on the Raf-1, c-Myc and c-Jun levels, and on the activity of MAP kinase. The results showed that 50 micrograms/ml GLA lowered the activity of MAP kinase. As expected with the decreased MAP kinase activity in the cells exposed to the higher level GLA, the c-Jun levels were also lowered. The levels of c-Myc, however, were increased. It is therefore possible that the deregulated expression of c-Myc in the HeLa cells exposed to the high level of GLA (50 micrograms/ml) may contribute to the induction of apoptosis in HeLa cells. PMID: 9014218 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 460: J Cell Sci. 1996 Dec;109 ( Pt 12):2855-63. Integrin-dependent induction of early growth response genes in capillary endothelial cells. Dike LE, Ingber DE. Department of Surgery, Children's Hospital, Boston, MA 02115, USA. Studies were carried out to explore how extracellular matrix molecules, such as fibronectin (FN), promote capillary endothelial (CE) cell growth. When G0-synchronized cells were plated on FN-coated dishes, expression of the immediate-early mRNAs, c-fos, c-myc and c-jun, was rapidly induced, even in the absence of serum or soluble growth factors. Moreover, plating cells on different FN densities (5-200 micrograms/150 mm dish), resulted in a dose-dependent increase in the steady state levels of these mRNAs. Addition of FGF potentiated gene activation and was required for maximal DNA synthesis, however, the overall steady-state level of gene induction was dictated primarily by the density of immobilized FN. Expression of junB also was induced when suspended cells bound RGD-peptide coated microbeads that promote integrin clustering, but not when the suspended cells bound beads coated with other receptor ligands (e.g. acetylated low density protein) or when they were stimulated by soluble FN or FGF in the absence of substrate adhesion. c-Jun exhibited a similar requirement for gene induction except that it also was partially induced by binding to soluble FN alone. In contrast, c-fos expression was induced by all stimuli tested. Interestingly, inhibition of Na+/H+ exchange using hexamethylene-amiloride prevented most of the FN-induced increase in c-jun expression whereas it was relatively ineffective when cells were simultaneously stimulated by both FN and FGF. These data demonstrate that cell adhesion to extracellular matrix and associated integrin binding can directly activate signaling cascades in quiescent CE cells that lead to induction of immediate-early genes associated with the G0/G1 transition and thereby, stimulate these cells to reenter the growth cycle. PMID: 9013333 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 461: Isr J Med Sci. 1996 Dec;32(12):1186-91. Esterase inhibitors diminish the modulation of gene expression by butyric acid derivative, pivaloyloxymethyl butyrate (AN-9). Rabizadeh E, Shaklai M, Eisenbach L, Nudelman A, Rephaeli A. Institute of Hematology, Rabin Medical Center, Petah Tikva, Israel. Pivaloyloxymethyl butyrate (AN-9) belongs to the family of acyloxyalkyl ester prodrugs of carboxylic acids which undergo intracellular hydrolysis to yield butyric acid (BA). We have previously shown that AN-9 and BA reduce the level of c-myc and enhance c-jun transcripts in HL-60 cells, and that the differentiation of these cells, induced by AN-9, is dependent on the presence of intracellular esterases. In this study we show that esterase inhibitors abolish the changes induced by AN-9 on c-myc and c-jun expression. In contrast, esterase inhibitors do not change the effects of BA on c-myc or c-jun. Interestingly, these inhibitors affect the modulation induced by both AN-9 and BA on the retinoblastoma tumor suppressor gene. These data suggest that AN-9 is indeed a prodrug of BA and that prior intracellular hydrolysis by esterases is material for AN-9 activity. PMID: 9007151 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 462: Carcinogenesis. 1996 Dec;17(12):2711-7. Changes in methyl-sensitive restriction sites of liver DNA from hamsters chronically exposed to hydrazine sulfate. Zheng H, Shank RC. Department of Community and Environmental Medicine, University of California, Irvine 92697-1825, USA. Hydrazine sulfate is a genotoxic hepatocarcinogen for the hamster. A study was conducted to follow changes in DNA maintenance methylation in selected genes in liver DNA during the 21-month induction of liver adenomas and hepatocellular carcinomas by demonstrating changes in restriction fragment length polymorphism. Male Syrian golden hamsters were exposed to hydrazine sulfate in the drinking water at three concentrations (170, 340 and 510 mg/l) shown previously to result in a dose-dependent induction of liver tumors. Liver DNA from animals exposed to the high concentration for 6, 12, 16, 20 and 21 months and animals exposed to the low or mid concentration for 21 months was digested with EcoRI, MspI, HindIII or BamHI, or a combination of one of these endonucleases and a methyl-sensitive restriction enzyme, HpaII or HhaI. The DNA digests were subjected to Southern analysis using a c-DNA probe for one of the following genes: DNA methyltransferase (DMT), c-Ha-ras, c-jun, c-fos, and c-myc proto-oncogenes, p53 tumor suppressor gene or gamma-glutamyltranspeptidase. Alteration in DNA restriction by methyl-sensitive endonucleases was detected in four (DMT, c-Ha-ras, p53 and c-jun) of the seven genes examined and as early as 6 months in animals exposed to the highest concentration of hydrazine sulfate; alteration of recognition sites in c-Ha-ras was also detected in DNA from animals exposed for 21 months to the intermediate concentration of hydrazine sulfate. Early changes in recognition sites, presumed to indicate altered methylation status of DNA cytosine and/or guanine mutations, were seen using c-DNA probes for DMT, c-Ha-ras and c-jun; in the p53 tumor suppressor gene alteration of such sites was a late event relevant to appearance of liver adenomas and hepatocellular carcinomas. Evidence for hypomethylation in the p53 and c-jun genes and hypermethylation of the c-Ha-ras and DMT genes is provided. This study supports the induction of site-specific hypomethylation and hypermethylation during the course of hydrazine carcinogenesis. PMID: 9006110 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 463: Mol Biol Cell. 1996 Dec;7(12):2045-56. Structure of pp32, an acidic nuclear protein which inhibits oncogene-induced formation of transformed foci. Chen TH, Brody JR, Romantsev FE, Yu JG, Kayler AE, Voneiff E, Kuhajda FP, Pasternack GR. Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. pp32 is a nuclear protein found highly expressed in normal tissues in those cells capable of self-renewal and in neoplastic cells. We report the cloning of cDNAs encoding human and murine pp32. The clones encode a 28.6-kDa protein; approximately two-thirds of the N-terminal predicts an amphipathic alpha helix containing two possible nuclear localization signals and a potential leucine zipper motif. The C-terminal third is exceptionally acidic, comprised of approximately 70% aspartic and glutamic acid residues; the predicted pI of human pp32 is 3.81. Human and murine pp32 cDNAs are 88% identical; the predicted proteins are 89% identical and 95% similar. Although the structure of pp32 is suggestive of a transcription factor, pp32 did not significantly modulate transcription of a reporter construct when fused to the Gal4 DNA-binding domain. In contrast, in cotransfection experiments, pp32 inhibited the ability of a broad assortment of oncogene pairs to transform rat embryo fibroblasts, including ras + myc, ras + jun, ras + E1a, ras + mutant p53, and E6 + E7. In related experiments, pp32 inhibited the ability of Rat 1a-myc cells to grow in soft agar, whereas it failed to affect ras-induced focus formation in NIH3T3 cells. These results suggest that pp32 may play a key role in self-renewing cell populations where it may act in the nucleus to limit their sensitivity to transformation. PMID: 8970164 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 464: Oncogene. 1996 Nov 7;13(9):1859-66. ECA39 is regulated by c-Myc in human and by a Jun/Fos homolog, Gcn4, in yeast. Ben-Yosef T, Yanuka O, Benvenisty N. Department of Genetics, Institute of Life Sciences, The Hebrew University of Jerusalem, Israel. myc oncogenes are transcription factors regulating the level of expression of other genes. Using a subtraction/coexpression strategy, a murine genetic target for Myc regulation was isolated. To further characterize this target gene, named ECA39, we have recently isolated the human, nematode and budding yeast homologs of the mouse gene. The recognition site for Myc binding, located 3' to the start site of transcription in the mouse gene, is conserved in the human homolog. Transfection experiments demonstrated that the Myc binding site of the human gene, mediates activation of a reporter gene in response to over-expression of c-myc. The activation was better executed when the c-Myc binding element was positioned downstream to the promoter, which is the usual position of the c-Myc DNA binding element in its genetic targets. The tissue specific expression of human ECA39 during embryogenesis is similar to that of the mouse homolog. Moreover, ECA39 is expressed in c-myc induced human tumors. It is expressed in Burkitt's lymphoma (where c-myc is translocated and activated) but not in non Burkitt's B-cell lymphoma or in T-cell lymphoma. Thus, it seems that ECA39 is a target for c-myc oncogenesis in humans. In yeast, where c-myc is absent, the ECA39 sequences lack the c-Myc binding element. However, the promoter region of the yeast ECA39 harbors several Gcn4 binding elements. Moreover, ECA39 is markedly down regulated in cells deleted for gcn4, and deletion of Gcn4 binding elements down regulated the transcription from ECA39 promoter. We thus suggest that ECA39 is a target for c-Myc regulation in mammals, while in yeast the regulator is not c-Myc but the c-Jun/c-Fos homolog - Gcn4. PMID: 8934531 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 465: Anticancer Res. 1996 Nov-Dec;16(6B):3659-64. The effect of etoposide (VP-16) on mouse L fibroblasts: modulation of stress response, growth and apoptosis genes. Sacchi CM, Schiaffonati L. Dipartimento di Medicina ed Oncologia Sperimentale, Universita degli Studi, Torino, Italy. Molecular events were studied in mouse L cells treated with etoposide (VP-16) a drug widely used in cancer therapy. Modulation of the expression of stress response genes belonging to the hsp70 family (hsp70, hsc73, grp78), growth- and cycle-related genes (c-myc, c-fos, c-jun, histone H3) and apoptosis-related genes (p53, TRPM-2, tTG) was monitored at different time points in the cells that remained adherent to the substrate up to 96 hours after exposure to VP-16. The steady state level of mRNA was determined by Northern blot analysis and hybridization with specific probes, and the relative rate of gene transcription was monitored by run on transcription with isolated nuclei. Our results indicate that protracted VP-16 treatment of 1. cells induces, within 24 hours, the arrest of DNA synthesis, repression of growth-related genes and transient induction of the tTG gene. Altogether these molecular events may contribute to the cytotoxic effect of VP-16. However in cells surviving a longer exposure to the drug, the expression of growth-related genes resumes, even if a blockade in DNA replication persists, and expression of the grp78 gene significantly increases. These data suggest that under continuous treatment with VP-16 a fraction of L cells showing increased resistance to the drug may emerge. PMID: 9042238 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 466: Anticancer Res. 1996 Nov-Dec;16(6B):3483-9. Deoxyadenosine-resistant mouse leukemia L1210 cell lines with alterations in early response genes and p53. Cory JG, He AW, Cory AH. Department of Biochemistry, East Carolina University School of Medicine, Greenville, NC 27858, USA. L1210 cell lines selected for resistance to deoxyadenosine exhibit altered steady-state levels of the mRNA for the early response genes and p53. In the deoxyadenosine-resistant cell lines (Y8 and ED2), the levels of the mRNAs for p53 and c-jun were markedly decreased while the steady-state levels for mRNAs for c-myc, c-fos and jun B were elevated in the Y-8 and ED2 cell lines. The levels of the mRNAs for PCNA and c-myb were the same in the wild type and mutant cell lines. The levels of the mRNAs for krox-24 were extremely low in the wild type and mutant cell lines. Cycloheximide (CHX) treatment of the cells resulted in the increase in the mRNA levels for c-jun, jun B, krox 24 and p53 in the Y-8 and ED2 cell lines. The time courses and the extents of the increases in the mRNA levels following CHX treatment were not the same for all of these mRNAs. The level of p53 RNA increased with no lag following CHX treatment while the levels of the mRNAs for c-myc, c-jun and krox-24 increased after a one-hour lag period. The level of the mRNA for p53 and c-myc increased 20- and 7-fold, respectively while the mRNA level for knox-24 increased 80-fold following CHX treatment. The Y8 and ED2 cell lines that lack steady-state levels of p53 show decreased sensitivity to cisplatin and increased frequency of gene amplification as measured by PALA resistance in a manner similar to other cell lines lacking p53. On the other hand, the ED2 and Y8 cell lines do not show a G1-block in response to PALA treatment. The cell lines appear to offer an experimental system in which to study the interactions between/among these early response genes and the p53-dependent and independent pathways. PMID: 9042210 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 467: Endocr Res. 1996 Nov;22(4):373-83. Regulation of growth by ACTH in the Y-1 line of mouse adrenocortical cells. Armelin HA, Lotfi CF, Lepique AP. Departmento de Bioquimica, Universidade de Sao Paulo, Brasil. Y-1 adrenal cells were cell cycle arrested by serum starvation to characterize a G0-->G1-->S transition in these cells. Cycle arrested Y-1 cells start to enter S phase 8h after serum feeding, reaching more than 90% cells synthesizing DNA by 24h. ACTH displays a dual effect in the G0-->G1-->S transition: 2h ACTH treatment stimulates DNA synthesis initiation, but longer treatments inhibit S phase entry. This dual effect of ACTH is similar to the antagonistic actions of PMA (phorbol-12-miristate-13-acetate) on the G0-->G1-->S transition. However ACTH and PMA are likely to have different mechanisms of action. ACTH inhibitory effect requires PKA, whereas PMA inhibitory effect is not dependent on PKA. ACTH induces the proto-oncogenes c-fos and c-jun, but inhibits the expression of the c-myc proto-oncogene. PMA, on the other hand, induces equally well c-fos, c-jun and c-myc. We hypothesize that ACTH promotes G0-->G1 transition by induction of c-fos and c-jun and blocks G1-->S transition by c-myc inhibition. PMID: 8969886 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 468: J Immunol. 1996 Oct 15;157(8):3235-41. IL-12-induced activation of NK and T cells occurs in the absence of immediate-early activation gene expression. Azzoni L, Kanakaraj P, Zatsepina O, Perussia B. Kimmel Cancer Institute, Jefferson Medical College, Philadelphia, PA 19107, USA. The responses of lymphocytes to IL-2 and IL-12, involving proliferation, differentiation, and cytokine production, are only partially overlapping, and may depend on induced differential expression of specific sets of genes. Using reverse-transcription PCR differential display, we isolated an mRNA species expressed in IL-2- but not IL-12-stimulated NK cells. This was identified as the mRNA encoding the transcription factor egr-1, which is expressed with fast kinetics in T and NK cells upon IL-2, but not IL-12, stimulation. Analysis of the accumulation of mRNA-encoding members of the AP-1 transcription factor family demonstrated that c-fos and junB are also expressed upon stimulation of NK and T cells with IL-2, but not IL-12, whereas expression of c-jun and junD is not modified by either cytokine. Accordingly, increased AP-1 DNA-binding activity and AP-1-dependent transcriptional activity were detected exclusively in IL-2-stimulated cells. Analysis of the expression of genes reported to regulate cytokine-induced proliferation demonstrated that both IL-2 and IL-12 induce c-myc mRNA accumulation in NK and T cells, whereas only IL-2 induces bcl-2 expression. Our data provide the first demonstration that IL-12-mediated activation of T and NK cells does not involve expression of members of the immediate-early activation genes family (egr-1, c-fos, and junB), AP-1 transcriptional activity, or bcl-2 expression. This indicates that functional differences observed in IL-2- and IL-12-stimulated cells may depend, at least in part, on differential gene regulation. PMID: 8871617 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 469: J Biol Chem. 1996 Oct 4;271(40):24753-60. Increased hepatic cell proliferation and lung abnormalities in mice deficient in CCAAT/enhancer binding protein alpha. Flodby P, Barlow C, Kylefjord H, Ahrlund-Richter L, Xanthopoulos KG. Karolinska Institute, Department of Biosciences at Novum S-141 57 Huddinge, Sweden. CCAAT/enhancer binding protein alpha (C/EBPalpha) is a transcription factor that has been implicated in the regulation of cell-specific gene expression mainly in hepatocytes and adipocytes but also in several other terminally differentiated cells. It has been previously demonstrated that the C/EBPalpha protein is functionally indispensable, as inactivation of the C/EBPalpha gene by homologous recombination in mice results in the death of animals homozygous for the mutation shortly after birth (Wang, N., Finegold, M. J., Bradley, A., Ou, C. N., Abdelsayed, S. V., Wilde, M. D., Taylor, L. R., Wilson, D. R., and Darlington, G. J. (1995) Science 269, 1108-1112). Here we show that C/EBPalpha -1-mice have defects in the control of hepatic growth and lung development. The liver architecture is disturbed, with acinar formation, in a pattern suggestive of either regenerating liver or pseudoglandular hepatocellular carcinoma. Pulmonary histology shows hyperproliferation of type II pneumocytes and disturbed alveolar architecture. At the molecular level, accumulation of glycogen and lipids in the liver and adipose tissues is impaired, and the mutant animals are severely hypoglycemic. Levels of c-myc and c-jun RNA are specifically induced by several fold in the livers of the C/EBPalpha -/- animals, indicating an active proliferative stage. Furthermore, immunohistologic detection with an antibody to proliferating cell nuclear antigen/cyclin shows a 5-10 times higher frequency of positively stained hepatocytes in C/EBPalpha -/- liver. These results suggest a critical role for C/EBPalpha in vivo for the acquisition of terminally differentiated functions in liver including the maintenance of physiologic energy homeostasis. PMID: 8798745 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 470: Cancer Lett. 1996 Oct 1;107(1):97-103. Calcium ionophore A 23187 induces apoptotic cell death in rat thymocytes. Azmi S, Dhawan D, Singh N. Department of Biochemistry, A.I.I.M.S., Ansari Nagar, New Delhi, India. Apoptotic cell death was induced in rat thymocytes on exposure to calcium ionophore A 23187 (100 micron(s)) for 24 h as observed from morphological changes and DNA fragmentation into oligonucleosomal ladder. The cell death was independent of de novo syntheses of protein. However, the involvement of c-Myc, c-Jun, poly ADPR polymerase and antioxidant enzymes CuZn SOD and catalase was observed. PMID: 8913272 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 471: Br J Rheumatol. 1996 Oct;35(10):933-42. Oncoprotein expression in human synovial tissue: an immunohistochemical study of different types of arthritis. Roivainen A, Soderstrom KO, Pirila L, Aro H, Kortekangas P, Merilahti-Palo R, Yli-Jama T, Toivanen A, Toivanen P. Turku Immunology Centre, Department of Medical Microbiology, Turku University, Finland. Based on the fact that synovial lining cells have some properties of transformed-appearing cells, we have examined the expression of Myc, Myb, Fos, Jun and Ras oncoproteins in synovial tissues from patients with different types of arthritis. Formalin-fixed and paraffin-embedded sections of synovial tissue from 12 patients with rheumatoid arthritis (RA), 14 with reactive arthritis (ReA), nine with other seronegative arthritis (OSA), seven with bacterial arthritis (BA), eight with probable bacterial arthritis (PBA) and eight with osteoarthritis (OA) were studied using the immunoperoxidase staining technique. The oncoproteins studied were expressed both in the synovial lining layer and in the sublining layer, consisting of lymphocytes, other inflammatory cells and blood vessels. Among the six disease entities, RA and OA appeared to be the most distinct, whereas the results obtained for ReA and OSA, and on the other hand for BA and PBA, closely resembled each other. The expression of Myc, Myb, Fos and Jun was significantly correlated both to the degree of synovial hypercellularity and the synovial lymphocytic infiltration. For Ras, such a correlation could not be seen. We conclude that we find no evidence of a cell lineage-specific or a disease-specific abnormality of proto-oncogene products in RA, and the expression of these oncoproteins is consistent with inflammation rather than with any primary abnormality of cell growth. PMID: 8883430 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 472: Hepatology. 1996 Oct;24(4):849-54. Role of the pituitary in tumor promotion with ethinyl estradiol in rat liver. Hallstrom IP, Liao DZ, Assefaw-Redda Y, Ohlson LC, Sahlin L, Eneroth P, Eriksson LC, Gustafsson JA, Blanck A. Department of Medical Nutrition, NOVUM, Huddinge University Hospital, Sweden. Synthetic estrogens act as tumor promoters in rat liver. Because estrogen treatment markedly increases the secretion of pituitary prolactin, also shown to be a tumor promoter in rat liver, the possibility of a pituitary influence in estrogen promotion was investigated in Wistar rats. In diethylnitrosamine (DEN)-initiated hypophysectomized (hx) female rats, 24 weeks of ethinyl estradiol (EE) administration (500 microg/kg/d, intraperitoneally) did not increase the number of hepatocyte nodules and did not induce hepatocellular carcinoma (HCC) in a 2-year study. Very few placental forms of glutathione-S-transferase (GST-P)-positive foci were observed at the end of EE administration. Estrogen receptor (ER) messenger RNA (mRNA) levels in hx females were 20% of the levels in intact females. EE administration (range, 160-210 microg/kg/d, subcutaneous release pellets) to DEN-initiated intact males and females increased the number and size of hepatocyte foci. A significant increase in HCC frequency was observed in EE-treated females compared with females receiving sham-release pellets, and the latency period for HCC induction was decreased by EE in both males and females. Inhibition of prolactin (PRL) secretion by bromocriptine (Brc) (ParlodelLAR, slow intramuscular release vehicles) during EE treatment decreased the number of foci without affecting their size and markedly prolonged the latency period in both sexes. EE treatment also significantly increased the expression of c-myc, and c-jun, enhanced the levels of growth hormone receptor (GHr) mRNA in females and the levels of ER mRNA in males and "feminized" the expression of the GH-regulated genes cytochrome P450 (CYP), 2C11, CYP 2C12, and GHr in male liver. Brc administration decreased the mRNA levels of the female-predominant CYP 2C12 in EE-treated males but otherwise had no effects. In conclusion, a decreased promotive effect of EE was obtained by decreasing the PRL levels, indicating that estrogens exert at least part of their promotion effects indirectly, by increasing the levels of pituitary PRL. PMID: 8855187 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 473: J Am Coll Cardiol. 1996 Oct;28(4):803-12. The renin-angiotensin system and vascular hypertrophy. Rosendorff C. Department of Medicine, Mount Sinai School of Medicine, New York, New York, USA. In addition to its vasoconstrictor and aldosterone-stimulating action, angiotensin II also drives cell growth and replication in the cardiovascular system, which may result in myocardial hypertrophy and hypertrophy or hyperplasia of conduit and resistance vessels in certain subjects. These actions are mediated through angiotensin II receptors (subtype AT1), which activate the G protein, phospholipase C, diacylglycerol and inositol trisphosphate pathway, to increase the expression of certain protooncogenes (c-fos, c-myc and c-jun) and growth factors (platelet-derived growth factor-A-chain, transforming growth factor-beta 1 and basic fibroblast growth factor). The cellular responses to angiotensin II in vascular smooth muscle have been shown in different hypertensive vessels to be either hypertrophy alone, hypertrophy and DNA synthesis without cell division (polyploidy) or DNA synthesis with cell division (hyperplasia). In genetic hypertension, the altered structure of small arteries is due to either cellular hyperplasia or remodeling, whereas in renovascular hypertension there is hypertrophy of vascular smooth muscle cells. Angiotensin II also increases synthesis of some matrix components, activates blood monocytes and is thrombogenic. Angiotensin-converting enzyme (ACE) inhibitors prevent or reverse vascular hypertrophy in animal models of hypertension; this seems to be a class effect, shared to some extent with calcium channel blocking agents. In human hypertension, ACE inhibitors reduce the increased media/lumen ratio of large and small arteries in hypertension and increase arterial compliance. These properties are also shared by losartan, the first of the new class of angiotensin II receptor (AT1) antagonists. The clinical implications of these findings need to be tested through rigorous and prospective clinical trials. Publication Types: Review Review, Tutorial PMID: 8837552 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 474: Brain Res Mol Brain Res. 1996 Sep 5;41(1-2):74-80. Transient expression of the mitogen-activated protein kinase phosphatase MKP-1 (3CH134/ERP1) in the rat brain after limbic epilepsy. Gass P, Eckhardt A, Schroder H, Bravo R, Herdegen T. German Cancer Research Center, Heidelberg, Germany. The immediate early gene-encoded enzyme, MAP kinase phosphatase 1 (MKP-1), is thought to be a key element in controlling cellular signalling pathways activated by MAP kinases. Since MAP kinase have been demonstrated to participate in neuronal stimulus-transcription coupling following seizure activity, the present study investigated the induction of MKP-1 in the rat brain after limbic epilepsy. MKP-1 expression was studied with a polyclonal antiserum by Western blots, immunocytochemistry and immuno-electron microscopy at different time periods between 1 and 24 h after kainic acid-induced limbic seizures. MKP-1 induction was identified in dentate granule cells of the hippocampus but not in pyramidal neurons, furthermore in neurons of the outer layers of the neocortex, as well as in neurons of the lateral nucleus of the bed of the stria terminalis. Immuno-electron microscopy demonstrated that MKP-1 was localized in the neuronal nucleus, where the substrate of MKP-1, activated MAP kinases, are also found. In view of the restricted areas of MKP-1 expression and the widespread areas of altered MAP kinases activity it can be concluded that in the majority of CNS populations other mechanisms than MKP-1 induction are responsible for the shut-off of MAP kinases following seizure activity. MKP-1 may contribute in the specific subpopulations where it is induced to the post-translational control of inducible transcription factors of the fos, jun and myc family. PMID: 8883936 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 475: Braz J Med Biol Res. 1996 Sep;29(9):1133-40. Polyomavirus-induced malignant transformation: comparative analysis of wild type and mutant middle T-overexpressing cell lines. Armelin MC, Oliveira ML. Instituto de Quimica, Universidade de Sao Paulo, Brasil. mcsarmel@quim.iq.usp.br Polyomavirus, a DNA tumor virus, expresses three viral oncoproteins (large, middle and small T antigens), causes malignant transformation in cell culture and induces multiple tumors in vivo. The middle T (MT) antigen seems to play an essential role in transformation and tumorigenicity. The observation that MT-overexpressing cell lines are able to grow in the absence of PDGF (platelet-derived growth factor) led several laboratories to study the mechanism underlying MT-induced growth deregulation and the signal transduction pathway used by this viral oncoprotein. A number of cellular proteins were shown to be common to both the normal PDGF mitogenic pathway and the MT transforming pathway. The expression of some PDGF primary response genes (fos, jun, myc, JE, KC) was shown to be rendered constitutive by MT overexpression. Using MT mutants, important domains for binding and activation of cytoplasmic proteins were mapped. Wild type and mutant MT cell lines are used in our laboratory to analyze the expression and activity of the PDGF early response genes during cell transformation and correlate them with activation of specific cytoplasmic proteins. In addition to abrogating the PDGF requirement for growth, activation of cellular proteins caused by MT results in cell lines that have an altered morphology and are able to form colonies in agarose. These changes may be due to alterations in connexin 43 and other cell surface proteins. PMID: 9181056 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 476: J Viral Hepat. 1996 Sep;3(5):227-38. Long-term productive episomal hepatitis B virus replication in primary cultures of adult human hepatocytes infected in vitro. Rumin S, Gripon P, Le Seyec J, Corral-Debrinski M, Guguen-Guillouzo C. Inserm U49, Unite de Recherches Hepatologiques, Hopital Pontchaillou, Rennes, France. We have previously increased the efficiency of hepatitis B virus (HBV) infection of human hepatocytes in vitro by using polyethylene glycol. After further documenting by neutralization experiments, this in vitro infection, we used this model to define new culture conditions that would maintain stable episomal replication for several weeks. We found that in the presence of 10% porcine serum and 2% dimethyl sulphoxide (DMSO), high-density cultures survived more than 3 months, while addition of hydrocortisone instead of DMSO resulted in survival of less than 1 month. HBV episomal replication was maintained without any evidence of viral integration into the host genome. The maintenance of HBV replication was demonstrated by: first, stability of the covalently-closed-circular DNA in the nucleus and relaxed circular and single-stranded replicative intermediates in the cytoplasm; second, detection of two major transcripts of 3.5 and 2.1-2.4 kb corresponding to the pregenomic and surface genes respectively; and third, continuous secretion of mature viral particles in the supernatant of infected cells. We showed that under these culture conditions, hepatocytes were blocked in the G1 phase of the cell cycle and did not spontaneously proliferate. Upon hepatocyte growth factor (HGF) stimulation, however, the ability of hepatocytes to divide was demonstrated and was compared in infected and non-infected cells. No change in proliferative capacity and no variation in c-myc and c-jun levels could be found. Hepatocyte survival was not modified in infected cells, confirming that HBV is not cytopathic for normal human hepatocytes. These new culture conditions represent substantial progress in the study of HBV-host cell interactions. PMID: 8914002 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 477: Invest Ophthalmol Vis Sci. 1996 Sep;37(10):2068-80. Differential regulation of cytokine and receptor transcript expression in human corneal and limbal fibroblasts by epidermal growth factor, transforming growth factor-alpha, platelet-derived growth factor B, and interleukin-1 beta. Li DQ, Tseng SC. Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Florida 33101, USA. PURPOSE: To explore further the significance of three patterns of cytokine dialogues that have been characterized between human corneal and limbal epithelial cells and fibroblasts. METHODS: Northern hybridization of the transcript expression of type I cytokine receptors (EGFR, IL-1R, and PDGFR-beta), type II cytokines (bFGF, LIF, and TGF-beta 1), and type III cytokines (HGF and KGF) by human corneal and limbal fibroblasts was conducted under the modulation of TGF-alpha, PDGF-BB, IL-1 beta, and EGF (type I cytokines). The mechanism of upregulation by IL-1 beta was studied further with respect to proto-oncogene expression and under the treatment of cycloheximide and actinomycin D. RESULTS: Results showed that EGF upregulated LIF and HGF but downregulated KGF and M-CSF. Unlike EGF, TGF-alpha upregulated additional EGFR, PDGFR-beta, bFGF, and TGF-beta 1, suggesting that although they share the same EGFR, TGF-alpha, which is produced by epithelium, is more effective in activating fibroblasts than EGF, which is present in tears. The upregulation of PDGF-BB was similar to that of TGF-alpha, except that it further stimulated IL-8, supporting their synergistic roles in promoting wound healing. Uniquely, IL-1 beta upregulated KGF expression by limbal fibroblasts more than corneal fibroblasts and IL-8 and M-CSF expression, but it downregulated PDGFR-beta. In IL-1 beta, the upregulation of cytokines and receptors was preceded by the upregulation of c-fos, c-jun, and c-myc, and it was inhibited by actinomycin D. Its upregulation of LIF was superinduced, but the upregulation of bFGF and KGF was inhibited, and that of the rest was not affected by cycloheximide. CONCLUSIONS: These findings suggest that epithelial cells under stress or injury (producing IL-1) might preferentially activate limbal epithelial stem cells indirectly by fibroblasts and simultaneously might promote inflammation during wound healing. PMID: 8814146 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 478: J Gen Virol. 1996 Sep;77 ( Pt 9):2173-82. Extracellular simian virus 40 induces an ERK/MAP kinase-independent signalling pathway that activates primary response genes and promotes virus entry. Dangoria NS, Breau WC, Anderson HA, Cishek DM, Norkin LC. Department of Microbiology, University of Massachusetts, Amherst 01003, USA. Simian virus 40 (SV40) binding to growth-arrested cells activated an intracellular signalling pathway that induced the up-regulation of the primary response genes c-myc, c-jun and c-sis within 30 min and of JE within 90 min. The up-regulation of the primary response genes occurred in the presence of cycloheximide and when UV-inactivated SV40 was adsorbed to cells. SV40 binding did not activate Raf or mitogen-activated protein kinase (MAP/ERK1), or mobilize intracellular Ca2+. The SV40-induced up-regulation of c-myc and c-jun was blocked by the tyrosine kinase inhibitor, genistein, and by the protein kinase C (PKC) inhibitor, calphostin C, but not by expression of the MAP kinase-specific phosphatase, MKP-1. These results suggest that the SV40-induced signalling pathway includes the activities of a tyrosine kinase and a Ca(2+)-independent isoform of PKC, but not of Raf or MAP kinase. Finally, SV40 infectious entry into cells was specifically and reversibly blocked by genistein. PMID: 8811017 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 479: Cell Struct Funct. 1996 Aug;21(4):259-69. The phorbol ester TPA regulates collagen gene expression at the transcriptional level. Stuiver I, Hendrix MJ, Shimizu Y, Shimizu N. Department of Molecular and Cellular Biology, University of Arizona, Tucson 85721, USA. Previously, we reported that growth activation of quiescent 3T3-L1 cells by TPA led to a rapid increase of pro-alpha 2 (I) collagen mRNA and protein, while induction of pro-alpha 2 (I) was not observed in VT-1 cells, a line non-mitogenic in the presence of TPA (26). Here, we further examine the expression of pro-alpha 2 (I) collagen during mitogenic stimulation at the molecular level. In addition to pro-alpha 2 (I) mRNA, TPA treatment increased mRNA production of other collagen family members, pro-alpha 1 (I) and pro-alpha 1 (III) although in reduced amounts relative to pro-alpha 2 (I). In contrast to pro-alpha 2 (I), the mRNA expression profiles of several protooncogenes were regulated in both VT-1 and 3T3-L1 cells. Consistent with increased mRNA levels, TPA treated 3T3-L1 cells produced a matrix abundant in collagen type I protein. In vitro nuclear "run-on" transcription assays demonstrated a 4-fold increase in pro-alpha 2 (I) mRNA that was maximal within 10 min of TPA treatment. Using a chloramphenicol-acetyl transferase (CAT) assay, we identified a TPA sensitive domain within the promoter of the COL1A2 gene. These results establish COL1A2 as an early growth responsive gene, and that its regulation is PKC dependent. Additionally, the increased expression of protooncogenes and transin during TPA stimulation of non-mitogenic VT-1 cells indicated that the regulation of these genes is independent of PKC, indicating the existence of multiple regulatory mechanisms amongst early response genes. PMID: 8906362 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 480: Nephrol Dial Transplant. 1996 Aug;11(8):1528-31. Alteration of growth-related proto-oncogene expression in diabetic glomeruli by a specific endothelin receptor A antagonist. Nakamura T, Ebihara I, Tomino Y, Koide H. Department of Medicine, Juntendo University School of Medicine, Tokyo, Japan. BACKGROUND: The early phase of glomerular growth in diabetic rats is accompanied by increased c-fos and c-jun expression. The renal endothelin (ET)-1 mRNA levels increased with the progression of diabetic nephropathy. This study was designed to assess whether glomerular mRNA expression of growth-related proto-oncogenes in diabetic rat glomeruli is affected by a specific ET receptor A antagonist, FR139317. METHODS: Diabetes was produced by streptozotocin (STZ) injection. Animals were divided into four groups: (1) untreated diabetic rats (n = 25); (2) FR139317-treated diabetic rats (n = 20); (3) control non-diabetic rats (n = 20); and (4) FR139317-treated controls (n = 20). FR139317 treatment was continued for 24 weeks. c-myc, c-fos, c-jun and proliferating cell nuclear antigen (PCNA) mRNA expression in rat glomeruli from each group (n = 7) at 4, 12 and 24 weeks after STZ or saline injection was evaluated. RESULTS: Glomerular mRNA levels for c-myc, c-fos, c-jun and PCNA in group 1 increased with the progression of diabetic nephropathy (24 weeks: c-myc, 4.6-fold; c-fos, 3.8-fold; c-jun, 4.2-fold; and PCNA, 4.4-fold). FR139317 treatment attenuated increases in glomerular mRNA levels of these genes (24 weeks, c-myc, 0.4-fold; c-fos, 0.4-fold; c-jun, 0.5-fold; and PCNA, 0.4-fold) in group 2. However, FR139317 did not affect mRNA levels in control glomeruli. CONCLUSIONS: These data suggest that FR139317 may protect against glomerular injury of diabetes in rats by reducing mRNA levels of growth-related genes. PMID: 8856205 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 481: Cell Growth Differ. 1996 Aug;7(8):1039-50. Terminally differentiated skeletal myotubes are not confined to G0 but can enter G1 upon growth factor stimulation. Tiainen M, Pajalunga D, Ferrantelli F, Soddu S, Salvatori G, Sacchi A, Crescenzi M. Molecular Oncogenesis Laboratory Regina Elena Cancer Center, Rome, Italy. Terminally differentiated cells are specialized cells unable to proliferate that constitute most of the mammalian body. Despite their abundance, little information exists on the characteristics of cell cycle control in these cells and the molecular mechanisms that prevent their proliferation. They are generally believed to be irreversibly restricted to the G0 state. In this report, we define some features of a paradigmatic terminally differentiated system, the skeletal muscle, by studying its responses to various mitogenic stimuli. We show that forced expression of a number of cell cycle-regulatory genes, including erbB-2, v-ras, v-myc, B-myb, ld-1, and E2F-1, alone or in combinations, cannot induce terminally differentiated skeletal muscle cells (myotubes) to synthesize DNA. However, serum-stimulated myotubes display a typical immediate-early response, including the up-regulation of c-fos, c-jun, c-myc, and ld-1. They also elevate the expression of cyclin D1 after 4 hours of serum treatment. All these events take place in myotubes in a way that is indistinguishable from that of quiescent, undifferentiated myoblasts reactivated by serum. Moreover, pretreatment with serum shortens the time required by E1A to induce DNA synthesis, confirming that myotubes can partially traverse G1. Serum growth factors do not activate late-G1 genes in myotubes, suggesting that the block that prevents terminally differentiated cells from proliferating acts in mid-G1. Our results show that terminally differentiated cells are not confined to G0 but can partially reenter G1 in response to growth factors; they contribute to a much-needed definition of terminal differentiation. The important differences in the control of the cell cycle between terminally differentiated and senescent cells are discussed. PMID: 8853900 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 482: Circulation. 1996 Aug 1;94(3):547-54. Retraction in: Wu D, Yang CM, Lau YT, Chen JC. Circulation. 1998 Jul 7;98(1):94. Mechanism of catecholamine-induced proliferation of vascular smooth muscle cells. Yu SM, Tsai SY, Guh JH, Ko FN, Teng CM, Ou JT. Department of Pharmacology, Chang Gung College of Medicine and Technology, Kwei-San, Tao-Yuan, Taiwan, Republic of China. BACKGROUND: Catecholamines have been shown to aggravate atherosclerosis in animals and humans, and abnormal proliferation of vascular smooth muscle cells (VSMC) is a key event in the early stage of atherosclerosis. Catecholamines may be involved in such cell growth. Therefore, a series of experiments using cultured VSMC was performed to elucidate their possible mitogenic effect. METHODS AND RESULTS: We examined the mitogenic effect of catecholamines using rat aortic smooth muscle cells (VSMC) by measuring [3H]thymidine incorporation, checking with flow cytometry, and counting the cell number directly. Furthermore, the catecholamine-activated signal transduction pathway was assessed by measurement of the formation of inositol 1, 4, 5-triphosphate, intracellular Ca2+ concentration, mitogen-activated protein kinase (MAPK) activity, and mitogenic gene expression. Norepinephrine (NE) and phenylephrine stimulated [3H]thymidine incorporation and cell growth. Clonidine and isoproterenol showed little of such effects. Prazosin was more effective than either yohimbine or propranolol in suppressing the mitogenic effect of NE, indicating that catecholamine-induced VSMC proliferation is mediated by alpha 1-adrenoceptors. The alpha 1-adrenoceptor activation was coupled to pertussis toxin-insensitive Gq-protein and triggered phosphoinositide hydrolysis with subsequent activation of protein kinase C and MAPK in VSMC. In response to NE, both 42- and 44-kD MAPK were activated and tyrosine was phosphorylated. alpha 1-Adrenoceptor stimulation with NE also caused accumulation of c-fos, c-jun, and c-myc mRNA. Chloroethylclonidine completely blocked the alpha 1-adrenoceptor-mediated mitogenesis. CONCLUSIONS: The effect of catecholamines appears to be mediated via the activation of the chloroethylclonidine-sensitive alpha 1-adrenoceptors that triggers the phosphoinositide hydrolysis and activates the MAPK pathway, leading to DNA synthesis and cell proliferation. Publication Types: Retracted Publication PMID: 8759101 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 483: FASEB J. 1996 Aug;10(10):1118-28. Liver regeneration versus direct hyperplasia. Columbano A, Shinozuka H. Istituto di Patologia Sperimentale, University of Cagliari, Italy. Liver cell growth can be induced in two distinct patterns: compensatory regeneration and direct hyperplasia. In the former, DNA synthesis is preceded by a loss of liver cells such as seen after partial resection of the liver or cell necrosis, whereas in direct hyperplasia, DNA synthesis is stimulated without cell loss. During the past decade, considerable advances have been made in understanding molecular mechanisms of the compensatory regeneration. There is increasing evidence that hepatocyte proliferation induced by some primary mitogens is mediated by patterns of growth factor modulation and signal transduction different from those of compensatory regeneration. Indeed, whereas activation of transcription factors such as NF-kappa B and increased expression of immediate early genes such as c-fos, c-jun, egr-1, and c-myc are induced during compensatory regeneration, such changes are not observed during hyperplasia induced by certain primary mitogens. In addition, although experimental evidence suggests a critical role for growth factors such as hepatocyte growth factor and transforming growth factor-alpha for the progression into cell cycle of competent hepatocytes in compensatory regeneration, these growth factors do not appear to play a major role in direct hyperplasia. One class of primary mitogens may trigger their actions through tumor necrosis factor-alpha, and the other by activation of nuclear hormone receptors. The differences in molecular events observed between liver regeneration and direct hyperplasia may affect differently the initiation step of chemical hepatocarcinogenesis. Whereas the former supports initiation by chemicals, the latter does not. A similar lack of effect on promotion of carcinogen-altered cells has also been observed after acute treatment with some primary mitogens. Definition of the mechanisms by which primary mitogens stimulate liver cell proliferation may elucidate the nature of the signals responsible for triggering the entry into cell cycle. Furthermore, due to their low toxicity, primary liver mitogens could have significant clinical applications in gene transfer and liver transplantation. Publication Types: Review Review, Tutorial PMID: 8751714 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 484: J Cell Physiol. 1996 Aug;168(2):255-63. C-myc expression affects proliferation but not terminal differentiation or survival of explanted erythroid progenitor cells. Bondurant MC, Yamashita T, Muta K, Krantz SB, Koury MJ. Department of Medicine, Department of Veterans Affairs Medical Center, Nashville, Tennessee, USA. The expression of c-myc was analyzed in murine and human erythroblasts throughout their differentiation in vitro into reticulocytes. The murine cells were splenic erythroblasts from animals infected with the anemia strain of Friend virus (FVA cells). In FVA cells cultured without EPO, the c-myc mRNA and protein levels decrease sharply within 3 to 4 h, showing that continual EPO stimulation is required to maintain c-myc expression. When cultured with EPO, the c-myc mRNA level of FVA cells is raised within 30 min of exposure. The c-myc mRNA and protein reach maxima at 1 to 3 h, then decline slowly to very low levels by 18 h. In contrast, c-fos and c-jun mRNA levels are not regulated by EPO in FVA cells. The human cells analyzed were colony-forming units-erythroid, CFU-E, derived in vitro by the culture of peripheral blood burst-forming units-erythroid (BFU-E). When grown in EPO and insulin-like growth factor 1 (IGF-1) these cells differentiate into reticulocytes over 6 days rather than the 2 days required for murine cells, but the c-myc mRNA kinetics and response to EPO parallel those of mouse cells at similar stages of differentiation. Both IGF-1 and c-kit ligand (SCF) cause an additive increase in c-myc mRNA in human CFU-E in conjunction with EPO. These additive effects suggest that EPO, IGF-1, and SCF affect c-myc mRNA accumulation by distinct mechanisms. Addition of an antisense oligonucleotide to c-myc in cultures of human CFU-E specifically inhibited cell proliferation but did not affect erythroid cell differentiation or apoptosis. When human cells were grown in high SCF concentrations, an environment which enhances proliferation and retards differentiation, antisense oligonucleotide to c-myc strongly inhibited proliferation, but such inhibition did not induce differentiation. This latter result indicates that differentiation requires signals other than depression of c-Myc and resultant depression of proliferation. PMID: 8707861 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 485: Genes Cells. 1996 Jul;1(7):687-705. D-type cyclin expression is decreased and p21 and p27 CDK inhibitor expression is increased when tsBN462 CCG1/TAFII250 mutant cells arrest in G1 at the restrictive temperature. Sekiguchi T, Noguchi E, Hayashida T, Nakashima T, Toyoshima H, Nishimoto T, Hunter T. Molecular Biology and Virology Laboratory, Salk Institute, La Jolla, CA 92037, USA. BACKGROUND: The tsBN462 temperature-sensitive mutant hamster cell line exhibits cell cycle arrest and apoptosis at the restrictive temperature of 39.5 degrees C, due to a point mutation in the CCG1/TAFII250 gene, which encodes a component of the general transcription factor TFIID. RESULTS: We now report that CCG1/TAFII250 persisted as a complex with TBP and associated proteins (TAFs) in tsBN462 cells at the restrictive temperature. FACScan analysis revealed that the tsBN462 mutation resulted in a failure to progress out of G0 into G1. Using two-dimensional gel electrophoresis we observed a decrease in the synthesis of several proteins, starting in the middle of the G1 phase, becoming very pronounced during late G1. The expression of the immediate early genes c-fos, c-jun and c-myc was normally induced by serum treatment of quiescent cells at the restrictive temperature, whereas expression of cyclins A, D1 and D3 was reduced. Expression of the cyclin-dependent kinase (CDK) inhibitor proteins p21 and p27 was enhanced. Consistent with the decreased cyclin D and increased p21/p27 expression, we found that phosphorylation of Rb was decreased at 39.5 degrees C. Cyclin A-, E- and Cdk2-associated histone H1 kinase activity was reduced concomitantly with the increase in p21 protein. CONCLUSION: Decreased cyclin/Cdk kinase activity and decreased Rb phosphorylation are possible causes of G1 cell cycle arrest in tsBN462 cells at the restrictive temperature. PMID: 9078394 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 486: Braz J Med Biol Res. 1996 Jul;29(7):911-9. Molecular genetic approach to cell proliferation control and neoplasia. Armelin MC, Oliveira ML, Mercado JM, Sasahara RM, Valentini SR, Carvalho LH. Instituto de Quimica, Universidade de Sao Paulo, Brasil. mcsarmel@quim.iq.usp.br A number of gene products involved in the control of cell proliferation fall into one of two classes: oncogenes and tumor suppressor genes. The same gene products have also been associated with malignant growth (tumors) caused by radiation, chemicals and tumor viruses. Here we describe our attempts to elucidate the molecular mechanisms underlying polyomavirus-induced cell transformation and the anti-tumor activity of glucocorticoid hormones. Wild type and mutant polyomavirus middle T (MT) overexpressing cell lines, generated with retroviral vector constructs, were used to investigate the role played by peptide growth factor primary response genes (fos, jun, myc, JE, KC) in viral transformation and to map the transduction pathway of the mitogenic signal of MT. Overexpression of MT leads to increased AP-1 (Fos/Jun) transcriptional complex activity. Transformation defective mutant analysis allowed the identification of sites in the MT molecule that are crucial for this activity. Two different approaches were used to investigate the molecular basis for glucocorticoids anti-tumor activity, namely: blind cloning of cDNAs and analysis of growth control genes in C6 glioma cell variants that are either hypersensitive (C6/ST1) or unresponsive to glucocorticoids (C6/P7). Four different glucocorticoid-regulated cDNA sequences were isolated using differential hybridization. A number of differentially expressed sequences were isolated from glucocorticoid-treated C6/ST1 cells by differential display (DDRT-PCR) and are currently being characterized. Expression of known growth control genes in C6/ST1 cells allowed the identification of important candidates for glucocorticoid hormone targets. Publication Types: Review Review, Tutorial PMID: 9070380 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 487: Neuroendocrinology. 1996 Jul;64(1):20-4. Regulation of galanin by dexamethasone in the rat anterior pituitary and the uterus. Vrontakis M, Torsello A, Leite V, Vuille JC, Zhang H. Department of Physiology and Anatomy, University of Manitoba, Canada. Galanin is a 29 amino acid neuropeptide widely distributed throughout the mammalian nervous and endocrine system. We have previously reported that estrogen dramatically increases galanin gene expression and protein synthesis in the anterior pituitary (AP), while the expression in the uterus (UT) of the same animals is transient and similar to the induction of protooncogenes (c-fos, c-jun, c-myc). In order to examine if this pattern of induction is specific to estrogen administration, we investigated the effect of glucocorticoids, another steroid, on the gene expression of galanin in the AP and in the UT of ovariectomized female rats and in the AP of male rats. Using Northern blot analysis, the AP and the UT showed almost undetectable levels of galanin mRNA, but in vivo treatment of female rats with 1 mg/kg body weight of dexamethasone (DEX) led to a significant increase of galanin mRNA levels in both AP and UT. Similarly, DEX (0.1-5 mg/kg i.p.) significantly stimulated galanin mRNA levels in the AP of the male rats. In both males and females the peak of induction was at 9 h after injection that is different from the 3-hour peak after estrogen administration. Daily injection of DEX for up to 7 days sustained the levels of galanin mRNA in both the AP and the UT, in contrast to the transient induction of galanin in the UT after estrogen administration. No change was noted in the galanin protein content of AP (control = 30 +/- 3.5 ng/mg protein; DEX treated = 38 +/- 4.2 ng/mg protein). Interestingly, in the UT of ovariectomized rats the combination of DEX and DES (diethylstilbestrol) treatment for 2 days resulted in a synergistic stimulation of galanin mRNA. In summary, these data demonstrate a tissue- and steroid-specific regulation of the galanin gene in AP and UT and suggest that DEX regulates the galanin gene possibly through a pathway different from estrogen. PMID: 8811662 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 488: Anticancer Res. 1996 Jul-Aug;16(4A):1707-17. Molecular genetics of malignant insulinoma. Pavelic K, Hrascan R, Kapitanovic S, Vranes Z, Cabrijan T, Spaventi S, Korsic M, Krizanac S, Li YQ, Stambrook P, Gluckman JL, Pavelic ZP. Department of Molecular Medicine, Ruder Boskovic Institute, Zagreb, Croatia. Malignant insulinoma is an rare form of cancer with poor prognosis and a reported 5-year survival of 35%. Relatively little is known about the etiology of this disease or of the oncogenes and tumor suppressor genes that participate in its genesis and progression. To address this issue, several protooncogenes, including K-ras, N-ras, erbB-2, erbB-3,c-myc, c-fos, c-jun were examined. Also analyzed was the expression of the growth factors TGF-alpha, EGF, and insulin as well as the EGF receptor (EGF-R), p53 and the putative anti-metastasis gene nm23-H1. These were examined in malignant insulinomas, benign insulinomas, pancreatic B cell hyperplasias and in normal endocrine pancreas. Normal endocrine pancreas showed moderate immunoreaction for c-myc and a strong reaction for insulin. All other parameters were negative. Benign pancreatic B cell hyperplasias were slightly or moderately positive for N-ras and TGF-alpha, and were weakly positive for EGF-R. They were strongly positive for c-myc and insulin. In malignant insulinomas there was strong immunoreaction for c-myc, TGF-alpha, N-ras, K-ras and p53. Insulin reaction was moderate or strong. Molecular genetic studies have been performed for the presence of activating point mutations in codon 12 of the c-K-ras oncogene. Mutations were detected using primer-mediated, mutant-enriched, polymerase chain reaction-restriction fragment length polymorphism analysis and were further characterized by allele-specific oligonucleotide hybridization. Four out of six patients with malignant insulinoma and two out of eight patients with benign insulinoma harbored K-ras point mutations at codon 12. All patients with mutated K-ras oncogene also had elevated levels of p53 protein as well as c-myc and TGF-alpha. In one extremely malignant case we found concomitant mutation at codon 12 of K-ras and codon 61 of the N-ras gene. Our data are consistent with the idea that malignant progression is accompanied by the progressive accumulation of multiple genetic lesions and suggest that activation of myc, TGF-alpha and ras genes may be early events in the development of insulinoma. PMID: 8712689 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 489: J Vasc Res. 1996 Jul-Aug;33(4):266-87. Postnatal growth of the heart and its blood vessels. Hudlicka O, Brown MD. Department of Physiology, University of Birmingham, Edgbaston, UK. Although rapid growth of the heart during early postnatal development ceases with maturation of the organism, the potential for cardiomyocyte growth is not lost and may be observed even in senescent hearts. Rapid developmental heart growth is accompanied by a proportional growth of capillaries but not always of larger vessels, and thus coronary vascular resistance gradually increases. Growth of adult hearts can be enhanced by thyroid hormones, catecholamines and the renin-angiotensin system hormones, but these do not always stimulate growth of coronary vessels. Likewise, chronic exposure to hypoxia leads to growth, mainly of the right ventricle and its vessels but without vascular growth elsewhere in the heart. On the other hand, ischaemia is a potent stimulus for the release of various growth factors involved in the development of collateral circulation. Heart hypertrophy develops in response to training, pressure or volume overload. Training usually leads to growth of larger coronary vessels but little growth of capillaries, except in young animals. However, growth of the capillary bed, but not the resistance vasculature capacity, can be induced by either increased coronary blood flow, bradycardia (electrically or pharmacologically induced) or increased inotropism, all of which are involved in the training stimulus. Thus, what actually promotes growth of larger vessels as opposed to capillaries in training is unclear. Pressure overload hypertrophy is mediated by both the renin-angiotensin system and the response of cardiomyocytes to stretch; both lead to activation of early oncogenes (c-fos, c-jun, c-myc) and angiotensin II activates several protein kinases involved in cell growth. In this condition, growth of larger vessels is inadequate, although some capillary growth may occur. Volume overload leads to cardiomyocyte hypertrophy and hyperplasia and some increase in vascular supply. Deficits in capillary supply in pressure or volume overload hypertrophy can be reversed by chronic administration of ACE inhibitors, dipyridamole, the bradycardic drug alinidine or pacing-induced bradycardia respectively, but in neither case is training effective. Mechanical and humoral factors are involved in growth of cardiomyocytes and vessels. For cardiomyocytes, stretch is most important, activating oncogenes, protein kinases and possibly the inositol phosphate pathway, but not ion channels, with regulation by the balance of angiotensin II, TGF-beta 1 and IGF-1, but not FGFs. For vessels, growth is stimulated by stretch and shear stress, possibly with involvement of VEGF. Increased shear stress disrupts the glycocalyx on the luminal side of vessels and releases plasminogen activator and metalloproteinases which disrupt the basement membrane and enable endothelial cell migration and proliferation. It also causes rearrangement of the endothelial cytoskeleton and transmission of mechanical signals to the abluminal side disturbing extracellular matrix and causing distortion of capillary basement membrane. Stretch acting from the abluminal side has a similar effect resulting also in basement membrane disruption and endothelial cell proliferation. Publication Types: Review PMID: 8695752 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 490: J Toxicol Environ Health. 1996 Jul;48(4):359-77. Induction of c-myc and c-jun proto-oncogene expression in rat L6 myoblasts by cadmium is inhibited by zinc preinduction of the metallothionein gene. Abshire MK, Buzard GS, Shiraishi N, Waalkes MP. Inorganic Carcinogenesis Section, National Cancer Institute, Frederick Cancer Research and Development Center, Maryland 21702, USA. Certain proto-oncogenes transfer growth regulatory signals from the cell surface to the nucleus. These genes often show activation soon after cells are exposed to mitogenic stimulation but can also be activated as a nonmitogenic stress response. Cadmium (Cd) is a carcinogenic metal in humans and rodents and, though its mechanism of action is unknown, it could involve activation of such proto-oncogenes. Metallothionein (MT), a metal-inducible protein that binds Cd, can protect against many aspects of Cd toxicity, including genotoxicity and possibly carcinogenesis. Thus, the effects of Cd on expression of c-myc and c-jun in rat L6 myoblasts, and the effect of preactivation of the MT gene by Zn treatment on such oncogene expression, were studied. MT protein levels were determined by the Cd-heme assay, and MT, c-myc, and c-jun mRNA levels were measured using oligonucleotide hybridization and standardized to beta-actin levels. Cd (5 microM CdCl2, 0-30 h) stimulated both c-myc and c-jun mRNA expression. An initial peak of activation of c-myc expression occurred 2 h after initiation of Cd exposure, and levels remained elevated throughout the assessment period. Zn pretreatment markedly reduced the activation of c-myc expression by Cd compared to cells not receiving Zn pretreatment. Cd treatment increased c-jun mRNA levels by up to 3.5-fold. Again, Zn pretreatment markedly reduced Cd-induced activation of c-jun expression as minimal increases occurred with Cd exposures of < or = 1 h, but otherwise the Zn pretreatment prevented activation of c-jun. The Zn pretreatment elevated MT protein levels > 5-fold over control at the point of Cd exposure, but Cd exposure did not further elevate these Zn-induced MT levels. Similarly, Zn pretreatment did not result in increased relative MT mRNA levels above Cd exposure alone at various time points after Cd exposure. Therefore, Zn pretreatment, possibly by providing elevated MT protein levels at the point of Cd exposure, inhibited the Cd-induced c-myc and c-jun proto-oncogene expression. The extent of Cd-induced proto-oncogene activation thus may be limited by the presence of cellular MT. PMID: 8691507 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 491: Int J Radiat Oncol Biol Phys. 1996 Jul 1;35(4):709-19. Ionizing radiation effects on the KG1a primitive hematopoietic cell line. Clave E, Carosella ED, Gluckman E, Dubray B, Socie G. Unite de Recherche sur la Biologie des Cellules Souches, Hopital Saint-Louis, Paris, France. PURPOSE: Better understanding of radiation-induced effects on the hematopoietic system is important in both the context of therapeutic intervention and accidental exposure. However, direct study of these effects on the hematopoietic stem cell pool is hampered by the small number of accessible cells. We, thus, studied radiation-induced effects on the KG1a stem cell line. METHODS AND MATERIALS: We confirmed and extended the immunophenotype of KG1a with monoclonal antibodies, established a radiation survival curve, and quantified mRNAs by Northern blotting 30 min after 1, 2, and 3 Gy of ionizing radiation (IR) and followed for up to 48 h after a 3 Gy dose. Cell cycle status and apoptosis were assessed by fluorescent-activated cell sorter (FACS) analysis, cell morphology, and DNA fragmentation. RESULTS: KG1a was found to be CD34+, CD7+, Thy1 low, CD38 low, lineage negative (neg), C-KITneg and HLA-DRneg, a phenotype consistent with a primitive hematopoietic origin. This immunophenotype was not altered by x-ray irradiation. The D0 value was 1.75 Gy. We showed a time-dependent variation of c-jun mRNA expression with an early and transient dose-dependent induction followed by a second increase at 24 and 48 h: a biphasic dose-dependent variation of bcl-2 expression 30 min after irradiation with a reduction of mRNA level at 1 Gy, and a normalization at higher doses and stable levels of mRNA for c-fos, c-myc, G-CSF, GM-CSF, IL-6, TNF-alpha, TGF-beta, and MIP-1 alpha genes. Cell cycle analysis showed the absence of G1/S phase arrest, a point consistent with the absence of detection of P53 mRNA by Northern blot analysis. The dose-dependent G2/M phase arrest was not followed by significant apoptotic cell death. CONCLUSION: Taken together, this data indicates that radiation-induced cell death of KG1a, a cell line that has a relatively high D0 value, does not seem to be the result of the apoptotic pathway but occurs subsequent to a G2/M phase arrest. PMID: 8690637 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 492: Mol Carcinog. 1996 Jul;16(3):139-48. Dissociation of ras oncogene-induced gene expression and anchorage-independent growth in a series of somatic cell mutants. Feinleib JL, Krauss RS. Department of Biochemistry, Mount Sinai School of Medicine, New York, New York 10029, USA. The mechanism or mechanisms by which ras oncogenes induce morphological transformation and anchorage-independent growth are poorly understood but are thought to involve stable alterations in gene expression. We previously described a genetically dominant, mutant rat fibroblast cell line (ER-1-2) that is resistant to ras-induced anchorage-independent growth. We now describe a cell line derived from ER-1-2 cells, termed ER-1-2T, that has apparently sustained a second, dominant mutation that conferred on these cells the ability to form colonies in soft agar. Analysis of these and control cell lines demonstrated that deregulation of many of the genes commonly associated with the transformed phenotype could be dissociated from anchorage-independent growth. After infection with a ras-expressing retrovirus, both control and ER-1-2 cell lines constitutively expressed elevated levels of the c-jun, junB, fosB, c-myc, collagenase, ornithine decarboxylase, osteopontin, stromelysin, cathepsin L, and insulin-like growth factor 1 genes. These data indicate that signaling events downstream of ras were largely intact in ER-1-2 cells and that the defect in these cells lies either on a pathway separate from those that control stable, ras-mediated expression of these genes or at a point in the cell-division cycle distinct from those that control expression of the genes. In contrast, only c-jun, junB, c-myc, and ornithine decarboxylase were expressed at a significantly elevated level in ER-1-2T cells. Thus, deregulated expression of the genes analyzed was not sufficient for anchorage-independent growth. Furthermore, deregulation of most of them was also not necessary. PMID: 8688149 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 493: Cancer. 1996 Jun 15;77(12):2581-7. Oncogene expression in carotid body tumors. Wang DG, Barros D'Sa AA, Johnston CF, Buchanan KD. Division of Metabolism and Endocrinology, School of Clinical Medicine, The Queen's University of Belfast, United Kingdom. BACKGROUND: The genetic etiology of carotid body tumors is suggested by the familial occurrence of the neoplasm. Environmental influences are also implied by the fact that the tumor is more common in those living at high altitudes. However, the development of sporadic tumors occurring at sea level, which account for the majority of cases, remains unknown. METHODS: The clinical and pathologic features of 13 carotid body tumors excised in 13 patients were reviewed. Two patients had bilateral tumors, one with a strong family history, and two patients had recurrent carotid body tumors. All tumors were benign except for one that had local lymph gland metastases. All patients were followed up for a period ranging from 1 to 17 years. Each tumor was examined for the oncoproteins c-myc, bcl-2 c-erbB-2, c-erbB-3 and c-jun and for the proliferating cell nuclear antigen (PCNA) by immunohistochemistry. RESULTS: c-myc immunoreactivity was observed in all tumors and, in 12 of 13 cases, was present in more than 10% of tumor cells, bcl-2 immunoreactivity was found in 11 cases with 6 tumors exhibiting more than 10% immunopositive cells, c-jun expression was found in 5 cases with 3 tumors containing more than 10% immunopositive cells. Only two tumors were positive for c-erb-B2 immunoreactivity with a cytoplasmic staining pattern. One tumor was positive for c-erb-B3. CONCLUSIONS: The oncogenes c-myc, bcl-2 and c-jun, are abnormally expressed in some carotid body tumors. Their expression may contribute to the genesis of carotid body tumors. PMID: 8640709 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 494: Biochim Biophys Acta. 1996 Jun 11;1301(3):273-87. Ceramide signalling and the immune response. Ballou LR, Laulederkind SJ, Rosloniec EF, Raghow R. Department of Medicine, University of Tennessee, Memphis 38104, USA. lballou@utmem1.utmem.edu Ceramide, produced through either the induction of SM hydrolysis or synthesized de novo transduces signals mediating differentiation, growth, growth arrest, apoptosis, cytokine biosynthesis and secretion, and a variety of other cellular functions. A generalized ceramide signal transduction scheme is shown in Fig. 2 in which ceramide is generated through the activation of distinct SMases residing in separate subcellular compartments in response to specific stimuli. Clearly, specificity of cellular responses to ceramide depends upon many factors which include the nature of the stimulus, co-stimulatory signals and the cell type involved. Ceramide derived from neutral SMase activation is thought to be involved in modulating CAPK and MAP kinases, PLA2 (arachidonic acid mobilization), and CAPP while ceramide generated through acid SMase activation appears to be primarily involved in NF-kappa B activation. While there is no apparent cross-talk between these two ceramide-mediated signalling pathways, there is likely to be significant cross-talk between ceramide signalling and other signal transduction pathways (e.g., the PKC and MAP kinase pathways). Other downstream targets for ceramide action include Cox, IL-6 and IL-2 gene expression, PKC zeta, Vav, Rb, c-Myc, c-Fos, c-Jun and other transcriptional regulators. Many, if not all, of these ceramide-mediated signalling events have been identified in the various cells comprising the immune system and are integral to the optimal functioning of the immune system. Although the role of the SM pathway and the generation of ceramide in T and B lymphocytes have only recently been recognized, it is clear from these studies that signal transduction through SM and ceramide can strongly affect the immune response, either directly through cell signalling events, or indirectly through cytokines produced by other cells as the result of signalling through the SM pathway. An overview of the signalling mechanisms coupling ceramide to the modulation of the immune response is depicted in Fig. 3 and shows how ceramide may play pivotal roles in regulating a number of complex processes. The SM pathway represents a potentially valuable focal point for therapeutic control of immune responses, perhaps for either enhancement of the activity of T cells in the elimination of tumors, or the down-regulation of lymphocyte function in instances of autoimmune disease. The recent explosion of knowledge regarding ceramide signalling notwithstanding, a number of critical questions need to be answered before a comprehensive, mechanistic understanding can be formulated relative to the incredibly varied effects of ceramide on cell function. For example, (i) how is a structurally simple molecule like ceramide able to mediate so many different, and sometimes paradoxical, physiological responses ranging from cell proliferation and differentiation to inhibition of cell growth and apoptosis, (ii) what are the molecular identities and modes of activation of the various SMase isoforms, (iii) what determines the distribution of the unique isoforms of SMase in cells of different lineages or at different stages of differentiation, (iv) what is the relative contribution of ceramide generated through SM hydrolysis versus de novo synthesis, and (v) by what means does ceramide interact with specific intracellular targets? Although a number of ceramide-activatable kinases, phosphatases, and their protein substrates have been identified, a more extensive search for additional cellular targets will be indispensable in determining the phosphorylation cascades linking the activation of the SM pathway to the regulation of nuclear events. Clearly, cross-talk between ceramide-induced signal transduction cascades and other signalling pathways adds to the inherent difficulty in distinguishing the specific effects of complex, intertwining signalling pathways. Publication Types: Review PMID: 8664339 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 495: Glycobiology. 1996 Jun;6(4):399-406. Ganglioside GM3 inhibition of EGF receptor mediated signal transduction. Rebbaa A, Hurh J, Yamamoto H, Kersey DS, Bremer EG. Chicago Institute for Neurosurgery and Neuroresearch, IL 60614, USA. Ganglioside GM3 is a membrane component that has been described to modulate cell growth through inhibition of EGF receptor associated tyrosine kinase. In order to determine if the inhibition of cell growth by this ganglioside is specifically mediated through EGF receptor signaling, the effects of GM3 on key enzymes implicated in EGF signaling were determined and compared to another inhibitor of the EGF receptor kinase. Treatment of A1S cells in culture by GM3 or a tyrosine kinase inhibitor, leflunomide, led to the inhibition of MAP kinase and PI3 kinase activities. There was no detectable effect on phosphotyrosine phosphatases. In a cell free system, however, GM3 had no effect on the activity of these signaling intermediates. Leflunomide was able to directly inhibit MAP kinase activity. GM3 and leflunomide were also found to act differently on the expression of the early immediate genes. The expression of c-fos and c-jun was inhibited by both GM3 and leflunomide. The expression of c-myc, however, was only inhibited by leflunomide. These findings suggest that the action of GM3 on cell growth and signaling is specifically mediated by EGF receptor and that this ganglioside does not act directly on the intracellular intermediates of EGF receptor signaling. In addition, soluble small molecule tyrosine kinase inhibitors such as leflunomide can directly affect the activity of MAP kinases and possibly other signaling intermediates. The direct effects of leflunomide on signaling intermediates may explain the differential effects of leflunomide and GM3 on gene expression and cell growth. PMID: 8842703 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 496: J Dermatol Sci. 1996 Jun;12(2):147-55. Wavelength-specific induction of immediate early genes by ultraviolet radiation. Ariizumi K, Bergstresser PR, Takashima A. Department of Dermatology, University of Texas Southwestern Medical Center, Dallas 75235-9069, USA. Exposure of skin in vivo to ultraviolet B (UVB) or ultraviolet A (UVA) radiation produces a variety of distinct clinical manifestations. In the present study, we characterized the immediate early genes that are activated in an epidermoid carcinoma cell line (A431) when exposed to UVB (FS20 sunlamp) or UVA radiation (window glass-filtered black light). We observed that: (a) c-jun mRNA expression is upregulated predominantly by UVB; (b) fra-1 and c-myc are downregulated by UVB, whereas both are upregulated by UVA; (c) fra-2 and AP-2 are downregulated modestly by UVB, (d) c-fos is unaffected, and (e) optimal regulation of each gene is achieved at environmentally relevant fluences (25-100 J/m2 for UVB and 2500-10 000 J/m2 for UVA). Thus, distinct sets of genes are activated (or repressed) by UVB and UVA irradiation. Treatment with organic hydrogen peroxides mimicked UVB radiation in upregulating c-jun expression, suggesting the participation of reactive oxygen intermediates in the UVB-signaling pathway. We propose that wavelength-specific regulation of nuclear mediator genes accounts for the development of at least some of the wavelength-specific cutaneous manifestations of ultraviolet radiation. PMID: 8814547 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 497: Carcinogenesis. 1996 Jun;17(6):1349-56. In vitro exposure to cadmium in rat L6 myoblasts can result in both enhancement and suppression of malignant progression in vivo. Abshire MK, Devor DE, Diwan BA, Shaughnessy JD Jr, Waalkes MP. Inorganic Carcinogenesis Section, Laboratory of Comparative Carcinogenesis, Frederick Cancer Research and Development Center, MD 21702, USA. Cadmium (Cd), a carcinogenic metal in humans and rodents, has been shown to transform cells in vitro. Cd in certain instances can also be anti-carcinogenic. The effects of Cd have been studied in different mammalian cell culture systems, where it has been shown to increase expression of several proto-oncogenes. In the present study the ability of Cd to affect malignant transformation was systematically investigated in L6 cells. Cells were grown in monolayer culture with concentrations of either 0 or 0.5 microM CdCl2 in the medium. Cell cultures treated with Cd for 9 weeks showed growth of large colonies in soft agar, while untreated control cells did not. When injected s.c. into athymic nude mice the 9 week Cd-treated cells gave rise to large, highly malignant sarcomas, resulting in high host mortality (9 dead/9 injected, 100%) by 7 weeks. Mice injected with untreated control cells also developed tumors, but of significantly smaller size and growth rate and associated with a lower host mortality (4/10, 40%, P or = 1.0 x 10(-6) M were equally inhibitory to the clonogenic growth of U937 and TAM67-expressing cells. In contrast, lower concentrations of ara-C (i.e., < 5.0 x 10(-7) M) were significantly less inhibitory to mutant U937 cell colony formation than to their parental counterparts. The reduced sensitivity of TAM67-expressing cells to low concentrations of ara-C could not be attributed to biochemical or cytokinetic factors, since the two cell lines were indistinguishable with respect to 1-beta-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP) formation, ara-CTP:dCTP ratios, and S-phase fraction. However, a significantly lower percentage of TAM67-expressing cells exposed to submicromolar concentrations of ara-C exhibited features associated with a differentiated monocytoid phenotype (i.e., increased plastic adherence and CD11b expression) compared to their parental counterparts. Lower concentrations of ara-C were also significantly less effective in decreasing the percentage of S-phase cells and in down-regulating c-myc mRNA levels in the mutant line, events associated with induction of leukemic cell differentiation. Finally, ara-C-induced up-regulation of c-jun message and protein was markedly attenuated in TAM67-expressing cells, findings consistent with a c-jun dominant-negative model. Collectively, these findings suggest that dysregulation of c-jun in U937 cells antagonizes low-dose ara-C-mediated cellular maturation but does not prevent higher concentration of this agent from triggering apoptosis. They also raise the possibility that separate aspects of the antiproliferative actions of ara-C may be differentially regulated by c-jun. PMID: 8732670 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 506: Clin Exp Metastasis. 1996 May;14(3):209-14. Microvessel density, expression of proto-oncogenes, resistance-related proteins and incidence of metastases in primary ovarian carcinomas. Volm M, Koomagi R, Kaufmann M, Mattern J, Stammler G. German Cancer Research Center, University of Heidelberg, Germany. Relationships between the incidence of metastatic spread and microvessel density, expression of proto-oncogene products, or expression of resistance-related proteins were investigated in human ovarian carcinomas by immunohistochemistry. Ovarian carcinomas with a high microvessel density showed a significantly increased formation of metastases (P = 0.005). Tumors with positive immunoreactivity of c-jun and c-myc products had a higher metastatic spread; however, these results were not statistically significant. A marginally significant correlation existed between the expression of erbB1 (EGFR) and metastatic spread (P = 0.05). No significant relationship was found between the expression of the resistance-related proteins P-glycoprotein or glutathione S-transferase-pi and the incidence of metastases. Furthermore, no correlation was detected between expression of the heat shock protein 70 and the occurrence of metastases. PMID: 8674274 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 507: Exp Hematol. 1996 May;24(6):682-9. Reversible differentiation of human monoblastic leukemia U937 cells by ML-9, an inhibitor of myosin light chain kinase. Yamamoto-Yamaguchi Y, Makishima M, Kanatani Y, Kasukabe T, Honma Y. Department of Chemotherapy, Saitama Cancer Center Research Institute, Japan. Human monoblastic leukemia U937 cells are induced to differentiate into monocytes and macrophages by various agents. We have shown that 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-9), an inhibitor of myosin light chain kinase, induces differentiation of monocytoid leukemia cell lines U937 and THP-1 but not of myeloblastic leukemic ML-1 cell or erythroleukemia K562 cells. In the present study, we further analyzed the effect of ML-9 in comparison with that of 1 alpha, 25-dihydroxyvitamin D3 (VD3) a typical inducer of monocytic differentiation. ML-9 induced nitroblue tetrazolium (NBT)-reducing activity of U937 cell more rapidly than VD3: This differentiation marker was induced significantly after incubation with ML-9 and VD3 for 4 hours and 1 day, respectively. ML-9 also induced alpha-naphthyl acetate esterase (ANAE) activity, another monocytic differentiation marker, more rapidly than VD3. The maximum levels of these markers induced by ML-9 were comparable to those induced by VD3, but after removal of ML-9 from the medium by washing the cells, the expressions of theses markers decreased within 4 hours and reached basal levels in 1 day, indicating that ML-9's induction of expression of differentiation-associated phenotypes was reversible. The growth inhibition of U937 cells by ML-9 was also reversible. Similar effects were observed in another line of human monoblastic cells, THP-1. ML-9 had little or no effect on the morphology of U937 cells but increased the expression of monocyte-macrophage lineage-associated surface antigen, CD14, to some extent. Irreversible terminal differentiation induced by VD3 is associated with down regulation of the expression of c-myc and upregulation of the expression of c-fos and c-jun, but ML-9 did not affect the expression of these oncogenes appreciably. ML-9-induced differentiation was also reversible when the cells were cultured with cultured with ML-9 plus an anti-cancer drug such as 1-beta-D-arabino-furanosylcytosine or daunomycin. it became irreversible, however, upon simultaneous treatment with dexamethasone and transforming growth factor-beta 1 (TGF-beta 1), which did not induce differentiation of U937 cells but caused growth arrest of the cells in the G0/G1 phase of the cell cycle. These results suggest that ML-9 should be useful for studying the mechanisms of monocytic differentiation. PMID: 8635523 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 508: Mol Cell Biol. 1996 May;16(5):2283-94. Role of EGR1 in regulation of stimulus-dependent CD44 transcription in B lymphocytes. Maltzman JS, Carman JA, Monroe JG. Department of Pathology and Laboratory Medicine, University of Pennsylvania, School of Medicine, Philadelphia, 19104, USA. The immediate-early gene egr-1 encodes a transcription factor (EGR1) that links B-cell antigen receptor (BCR) signals to downstream activation events through the regulation of previously unidentified target genes. Here we identify the gene encoding the lymphocyte homing and migration protein CD44 as a target of EGR1 regulation in B cells. BCR-induced increases in CD44 mRNA expression and transcription levels are shown to occur in EGR1-expressing but not in nonexpressing subclones of the B-cell line WEHI-231. Kinetics of egr-1 transcription and the appearance of nuclear EGR1 protein precede CD44 induction and occur within 30 min after stimulation in the EGR1-expressing subclone. A single EGR1 binding motif is demonstrated at bp -301 of the human CD44 promoter. Cotransfection of a CD44 promoter-chloramphenicol acetyltransferase reporter construct with an egr-1 expression vector resulted in a 6.5- to 8.5-fold induction of transcriptional activity relative to an empty expression vector. The EGR1 binding motif was shown to be necessary for stimulus-induced expression of a CD44 promoter-chloramphenicol acetyltransferase reporter construct in nontransformed B lymphocytes and was required for transactivation by an EGR1 expression vector in a B-cell line. These studies identify EGR1 as an intermediary linking BCR-derived signals to the induction of CD44. The relevance of these molecular events to BCR signal transduction and antigen-stimulated B-cell-mediated immune responses is discussed. PMID: 8628295 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 509: Mol Cell Biol. 1996 May;16(5):1881-8. Stepwise transformation of rat embryo fibroblasts: c-Jun, JunB, or JunD can cooperate with Ras for focus formation, but a c-Jun-containing heterodimer is required for immortalization. Vandel L, Montreau N, Vial E, Pfarr CM, Binetruy B, Castellazzi M. Unite de Virologie Humaine, INSERM-U412, Ecole Normale Superieure, Paris, France. Among the Jun family of transcription factors, only c-Jun displays full transforming potential in cooperation with activated c-Ha-Ras in primary rat embryo fibroblasts. c-Jun in combination with Ras can both induce foci of transformed cells from rat embryo fibroblast monolayers and promote the establishment of these foci as tumoral cell lines. JunB can also cooperate with Ras to induce foci but is unable to promote immortalization. We report here that JunD, in cooperation with Ras, induces foci with an efficiency similar to that of JunB. Artificial Jun/eb1 derivatives from each of the three Jun proteins were also analyzed. These constructs carry a heterologous homodimerization domain from the viral EB1 transcription factor and are thought to form only homodimers in the cell. We show here that these Jun/eb1 chimeras are potent transactivators of AP1 sites and that they can cooperate with c-Ha-Ras to induce foci. However, among all the Ras-Jun and Ras-Jun/eb1 combinations tested, only foci from Ras-c-Jun can be efficiently expanded and maintained as long-term growing cultures. Therefore, we suggest that a heterodimer containing c-Jun might be required for in vitro establishment of these primary mammalian cells. PMID: 8628254 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 510: Hepatology. 1996 May;23(5):1012-9. Cell cycle progression proteins (cyclins), oncogene expression, and signal transduction during the proliferative response of human hepatocytes to hepatocyte growth factor. Gomez-Lechon MJ, Guillen I, Ponsoda X, Fabra R, Trullenque R, Nakamura T, Castell JV. Unidad de Hepatologia Experimental, Centro de Investigacion, Hospital La Fe, Valencia, Spain. Human hepatocytes stimulated with human recombinant hepatocyte growth factor (h-rHGF) (10 ng/mL) displayed a characteristic lag period before entering into the S phase. The duration of this delay was dependent on the timing of h-rHGF addition to cultures. The highest peak of DNA synthesis was observed at 120 hours of culture when hepatocytes were stimulated with h-rHGF at 72 hours of culture. This was accompanied by an early peak of c-jun and c-fos synthesis (3 hours after addition of h-rHGF) followed by c-myc (6 hours) and increased expression of cyclins A, B, D, and E (12 hours after h-rHGF). A significant dose-dependent increase in inositol 1,4,5-P3 was observed within 45 seconds after stimulation with the factor. This was followed by an immediate increase in the cytosolic-free calcium. Cyclic adenosine monophosphate (cAMP) levels did not change after stimulation with the factor. Tyrosine phosphorylation seems to be an early event in the course of the stimulatory effect of h-rHGF on DNA synthesis of hepatocytes; genistein, a tyrosine kinase inhibitor, impaired the stimulatory effect of h-rHGF on DNA synthesis dose dependently. On the other hand, the action of the factor was negatively regulated by protein kinase C activation, as shown by the increased stimulatory effect of h-rHGF on DNA synthesis upon inhibition of protein kinase C by H7. PMID: 8621126 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 511: Biochim Biophys Acta. 1996 Apr 24;1311(2):85-92. Neurotrophin-3 increases the DNA-binding activities of several transcription factors in a mouse osteoblastic cell line. Iwata E, Nakanishi T, Ogawa N, Ohyama K, Murakami T, Takigawa M. Department of Neuroscience, Okayama University Medical School, Japan. In the mouse osteoblastic cell line MC3T3-E1, the signaling responses of several DNA-binding proteins induced by the treatment of neurotrophin-3 were examined using electrophoretic mobility shift assay. Neurotrophin-3 increased binding activities in nuclear extracts of MC3T3-E1 cells to TPA-responsive element (TRE), cyclic AMP-responsive element (CRE) and serum-responsive element (SRE), but not binding activity in the nuclear extracts to c-Myc binding DNA element. Competition experiments revealed that the binding activity to TRE in the nuclear extracts of neurotrophin-3-treated MC3T3-E1 cells was entirely inhibited by the both unlabeled TRE and CRE probes. On the other hand, the binding activity to CRE was abolished by the unlabeled CRE probe but not by the same amount of unlabeled TRE probe. Moreover, immunodepletion/supershift assay using antibodies directed to Fos, Jun and CREB proteins, showed that the binding activities to TRE and CRE in the nuclear extracts were derived in part from these proteins. PMID: 8630334 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 512: Oncogene. 1996 Apr 18;12(8):1625-33. Regulation of epidermal growth factor receptor by activated H-ras and V-myc oncogenes in mouse Balb/3T3 cells: possible roles of AP-1. Okimoto T, Kohno K, Kuwano M, Gopas J, Kung HF, Ono M. Department of Biochemistry, Kyushu University, School of Medicine, Fukuoka, Japan. We previously reported that introduction of H-ras oncogene decreases the epidermal growth factor (EGF) binding activity to cell surface EGF receptor in mouse Balb/3T3. In this study, we have further isolated four H-ras transfectants, four v-myc transfectants and three both H-ras and v-myc (H-ras/v-myc) transfectants of mouse Balb/3T3 cells. In comparison with introduction of v-myc alone or both H-ras and v-myc oncogene, introduction of H-ras alone resulted in a loss of [125I]EGF binding activity to the cell surface EGF receptor. RT-PCR analysis also showed much lower levels of EGF receptor gene expression in H-ras transfectants compared to that of parental untransformed cells (Balb-Neo1), v-myc and H-ras/v-myc transfectants. Our results demonstrated the activated binding of a transcription factor, Stat1 p84/p91, which directly interacts with EGF receptor, to c-sis-inducible element (SIE) in both v-myc and H-rs/v-myc transfectants, but not in H-ras transfectants. Among transcription factors which we have analysed, activator protein 1 (AP-1) but not SP-1 was modulated by H-ras. Gel shift assays demonstrated the mobility pattern of TPA-responsive element (TRE) binding complex with AP-1 derived from H-ras transfectants migrated faster than those from Balb-Neo1, v-myc and H-ras/v-myc. Expression of c-Jun and Fra-1 was increased more than threefold in H-ras transfectants compared with Balb-Neo1, v-myc and H-ras/v-myc transfectants, but that of c-Fos, Jun B and SP-1 was unchanged. Both transient and permanent expression of H-ras enhanced AP-1 activity in mouse cells, but further co-introduction of dominant negative c-jun mutant encoding a transcriptionally inactive product inhibited the H-ras dependent AP-1 induction. Transfection of the dominant negative c-jun mutant also restored down-regulation of EGF binding by activated H-ras oncogene. Down-regulation of EGf receptor by activated H-ras and the possible involvement of a transcription factor, AP-1 will be discussed. PMID: 8622882 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 513: Dig Dis Sci. 1996 Apr;41(4):660-9. Expression of protooncogene-encoded mRNA by colonic epithelial cells in inflammatory bowel disease. Alexander RJ, Panja A, Kaplan-Liss E, Mayer L, Raicht RF. Department of Veterans Affairs Medical Center, New York, New York 10010, USA. Protooncogenes are cell cycle-related genes that are involved in cell growth of proliferation. Alterations in the level of expression of these genes, or expression of aberrant gene productions, have been observed in tumors and precancerous conditions. To determine if expression of these genes is altered in patients with inflammatory bowel disease (IBD) --who are at risk for development of colon cancer--we assayed transcripts of 15 protooncogenes in colonic epithelial cells of IBD patients and controls. Nine of these genes (H-ras, c-myc, c-fos, c-jun, junB, N-myc, c-abl, c-yes, and p53) were expressed in epithelial cells, whereas two (RB1 and N-ras) were not. expression of four other genes (c-src, K-ras, c-raf, and c-myb) was observed, but the intensity of these bands was too low for densitometric analysis. The steady-state levels of transcripts of H-ras and five nuclear protooncogenes (c-myc, c-fos, c-jun, junB, and N-myc) were lower in epithelial cells from involved or uninvolved IBD samples than in normal epithelial cells from either sporadic colon cancer or diverticulitis patients. The level of c-fos mRNA was two- to threefold higher in involved than in uninvolved areas of the colons of two ulcerative colitis (UC) patients, but not in one Crohn's disease (CD) patient. Message abundance of c-abl transcripts was two- to threefold lower in UC epithelial cells than in either the CD or control samples. The steady-state level of c-yes-encoded mRNA was considerably higher in IBD patients resected for colon cancer than in patients resected for active chronic IBD or in controls. The level of p53 message was constant in these samples. Increased levels of c-fos mRNA in involved UC relative to uninvolved UC may be related to the disease process. Decreased expression of c-abl transcript in UC may be a diagnostic marker for UC and may be related to the rate of cell turnover in these diseases. Enhanced expression of c-yes in IBD patients with tumors compared to active chronic IBD and controls suggests that expression of this gene may be a marker for development of colon cancer in IBD. PMID: 8674385 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 514: J Clin Invest. 1996 Apr 1;97(7):1577-88. 1,25(OH)2 vitamin D3, and retinoic acid antagonize endothelin-stimulated hypertrophy of neonatal rat cardiac myocytes. Wu J, Garami M, Cheng T, Gardner DG. Department of Medicine, University of California, San Francisco, 94143, USA. 1,25(OH)2 Vitamin D3 (VD3) and retinoic acid (RA) function as ligands for nuclear receptors which regulate transcription. Though the cardiovascular system is not thought to represent a classical target for these ligands, it is clear that both cardiac myocytes and vascular smooth muscle cells respond to these agents with changes in growth characteristics and gene expression. In this study we demonstrate that each of these ligands suppresses many of the phenotypic correlates of endothelin-induced hypertrophy in a cultured neonatal rat cardiac ventriculocyte model. Each of these agents reduced endothelin-stimulated ANP secretion in a dose-dependent fashion and the two in combination proved to be more effective than either agent used alone (VD3: 49%; RA:52%; VD3 + RA:80% inhibition). RA, at concentrations known to activate the retinoid X receptor, and, to a lesser extent, VD3 effected a reduction in atrial natriuretic peptide, brain natriuretic peptide, and alpha-skeletal actin mRNA levels. Similar inhibition (VD3:30%; RA:33%; VD3 + RA:59% inhibition) was demonstrated when cells transfected with reporter constructs harboring the relevant promoter sequences were treated with VD3 and/or RA for 48 h. These effects were not accompanied by alterations in endothelin-induced c-fos, c-jun, or c-myc gene expression, suggesting either that the inhibitory locus responsible for the reduction in the mRNA levels lies distal to the activation of the immediate early gene response or that the two are not mechanistically coupled. Both VD3 and RA also reduced [3H]leucine incorporation (VD3:30%; RA:33%; VD3 + RA:45% inhibition) in endothelin-stimulated ventriculocytes and, once again, the combination of the two was more effective than either agent used in isolation. Finally, 1,25(OH)2 vitamin D3 abrogated the increase in cell size seen after endothelin treatment. These findings suggest that the liganded vitamin D and retinoid receptors are capable of modulating the hypertrophic process in vitro and that agents acting through these or similar signaling pathways may be of value in probing the molecular mechanisms underlying hypertrophy. PMID: 8601621 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 515: Eur J Neurosci. 1996 Mar;8(3):589-97. Clusterin (SGP-2) induction in rat astroglial cells exposed to prion protein fragment 106-126. Chiesa R, Angeretti N, Lucca E, Salmona M, Tagliavini F, Bugiani O, Forloni G. Alzheimer Neurobiology Unit, Istituto di Ricerche Farmacologiche Mario Negri, Milan, Italy. Prion-related encephalopathies are characterized by the accumulation of an abnormal prion protein isoform (PrPSc) associated with neuronal degeneration and astrogliosis. The synthetic peptide homologous to PrP fragment 106-126 (PrP 106-126) induced in vitro neuronal apoptosis and glial proliferation. We used Northern blot analysis and the RNA polymerase chain reaction to assess the expression of several genes associated with programmed cell death and proliferation. Blots of total RNA extracted from neuronal and astroglial cells exposed to PrP 106-126 for between 1 h and 7 days were hybridized with probes recognizing c-fos, c-jun, c-myc, p53, hsp-70 and bcl-2 mRNA. Except for a slight decrease in bcl-2 mRNA in neuronal cells, no change in other transcripts was evident. Since clusterin (apolipoprotein J) mRNA levels are increased in prion-related encephalopathies and clusterin immunoreactivity has been located in association with PrPSc in Gerstmann-Straussler-Scheinker brain, the expression of clusterin was determined in neuronal and astroglial cells chronically exposed to PrP 106-126. Although the induction of clusterin has been involved in the apoptotic mechanism in other experimental conditions, its expression was unchanged in PrP 106-126-treated neurons, while a three-fold induction of clusterin mRNA was observed in astrocytes exposed to PrP 106-126. To investigate whether the clusterin up-regulation was simply associated with the astroglial proliferative stimulus of PrP 106-126 or was specifically induced by the peptide, we measured clusterin expression in astrocytes cultured in fetal calf serum-free medium and exposed to PrP 106-126 or fetal calf serum restoration. In this condition the PrP peptide, like fetal calf serum, increased the glial proliferation rate, but only PrP 106-126 doubled clusterin mRNA. The selectivity of this effect indicates that PrPSc is directly involved in the clusterin up-regulation seen in prion-related encephalopathies and is associated with astroglial cells. PMID: 8963451 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 516: FASEB J. 1996 Mar;10(4):413-27. Liver regeneration 4: transcriptional control of liver regeneration. Taub R. Department of Genetics and Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104-6145, USA. Determining what factors are responsible for initiating regeneration following partial hepatectomy or toxic damage, and how the liver maintains differentiated functions while the hepatocytes are undergoing cellular proliferation are central issues in understanding the molecular bases of liver regeneration. Examination of the transcriptional milieu in the regenerating liver provides clues to the answers to these questions. Growth factor-generated intracellular signals that trigger liver regeneration result in activation via posttranslational modifications of latent, normally inactive transcription factors that preexist in the liver. Two transcription factors that are activated by this mechanism include posthepatectomy factor/nuclear factor-kappa B) and Stat3. Because cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin-l (IL-1), and IL-6 can induce these factors in the liver, the finding of activated Stat3 and PHF/NF-kappa B suggests that these cytokines may play a role in some aspects of growth regulation during liver regeneration. Rapidly induced transcription factors, Stat3, PHF/NF-kappa B, and others are responsible for activation of the primary growth response or immediate-early genes, which play a role in regulating later phases of cell growth in regenerating liver and other mitogen-activated cells. Immediate-early genes encode many members of diverse transcription factor families including the Jun-Fos-LRF-1, nuclear receptor, and myc families to name a few. In this way a transcriptional cascade is established during the G1 phase of liver regeneration. Coexisting with these induced factors are liver-specific transcription factors such as the CAAT enhancer binding proteins and hepatocyte nuclear factors, which may interact with growth-induced factors to help the liver maintain metabolic homeostasis during regeneration. As a result the liver is able to accomplish the goals of reestablishing its mass while it maintains its functional capacity during regeneration. Publication Types: Review Review, Tutorial PMID: 8647340 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 517: Cell Immunol. 1996 Feb 25;168(1):1-12. MHC class I molecules are implicated in costimulatory signals during TCR/CD3-induced activation. Amirayan N, Machy P. Centre d'Immunologie INSERM-CNRS de Marseille-Luminy, Marseille, France. Mouse MHC class I-specific mAbs recognizing the alpha 1/alpha 2, but not those directed against the alpha 3 domain of the molecule, inhibited RNA, protein, and DNA synthesis of splenic T cells in response to stimulation through the TCR/CD3 complex. Similar inhibition was seen with LFA-1-specific mAbs under the same stimulation conditions. The effect of class I- and LFA-1-specific mAbs reflected a decrease of both IL-2 and IFN-gamma synthesis and IL-2 receptor alpha chain induction. IL-2, IL-2 receptor alpha chain, IFN-gamma, c-fos, c-jun, and c-myc mRNAs were not detected. Activation of AP-1 (c-Fos and c-Jun proteins) and NF-kappa B transcription factors were also inhibited. Inhibition was observed both after treatment of cells in culture and after intravenous injection of Abs in mice. Although bulk phosphorylation was inhibited, early tyrosine phosphorylation and calcium ion influx were normally induced. Protein phosphatase inhibitors did not reverse this inhibition, ruling out an enhanced activation of these enzymes in the observed inhibition. Cell surface expression of one of early PKC activation marker, CD69 was also inhibited. Phorbol esters that directly activate PKC prevented inhibition. Thus, class I molecules are implicated in signal transduction involved at an early stage for T cell activation in a manner that suggests their implication in accessory signal transmission that contributes to the regulation of PKC activity. PMID: 8599831 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 518: J Biol Chem. 1996 Feb 9;271(6):3229-37. Signal transduction pathways regulated by mitogen-activated/extracellular response kinase kinase kinase induce cell death. Johnson NL, Gardner AM, Diener KM, Lange-Carter CA, Gleavy J, Jarpe MB, Minden A, Karin M, Zon LI, Johnson GL. Division of Basic Sciences, National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado 80206, USA. Mitogen-activated/extracellular response kinase kinase (MEK) kinase (MEKK) is a serine-threonine kinase that regulates sequential protein phosphorylation pathways, leading to the activation of mitogen-activated protein kinases (MAPK), including members of the Jun kinase (JNK)/stress-activated protein kinase (SAPK) family. In Swiss 3T3 and REF52 fibroblasts, activated MEKK induces cell death involving cytoplasmic shrinkage, nuclear condensation, and DNA fragmentation characteristic of apoptosis. Expression of activated MEKK enhanced the apoptotic response to ultraviolet irradiation, indicating that MEKK-regulated pathways sensitize cells to apoptotic stimuli. Inducible expression of activated MEKK stimulated the transactivation of c-Myc and Elk-1. Activated Raf, the serine-threonine protein kinase that activates the ERK members of the MAPK family, stimulated Elk-1 transactivation but not c-Myc; expression of activated Raf does not induce any of the cellular changes associated with MEKK-mediated cell death. Thus, MEKK selectively regulates signal transduction pathways that contribute to the apoptotic response. PMID: 8621725 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 519: J Clin Endocrinol Metab. 1996 Feb;81(2):519-23. Insulin regulation of multiple ribonucleic acid species in human skeletal muscle in insulin-sensitive and insulin-resistant subjects. Thompson DB, de Gregorio M, Sommercorn J. Clinical Diabetes and Nutrition Section, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Phoenix, Arizona 85016, USA. In vivo short term (2 h) insulin-regulated gene expression was examined in skeletal muscle of persons with differing insulin sensitivities. Nine genes were analyzed by a S1 nuclease protection assay with multiple probes (multiple S1 nuclease protection assay) to allow the simultaneous examination of RNA abundances from the multiple genes. In insulin-sensitive individuals, 5 of these 9 genes were insulin responsive. RNA from the proto-oncogenes c-Ha-ras, c-myc, and c-src transiently increased 2- to 4-fold within 30 min of insulin infusion. In addition, the RNA abundance of myf-5, a muscle specific differentiation factor, increased 3-fold with a time course similar to that of c-Ha-ras, c-myc, and c-src. In contrast, type 1 protein phosphatase alpha (PPP1A) RNA levels decreased by 50% within 30 min. In insulin-resistant individuals, the RNA levels of c-Ha-ras and myf-5 did not increase, whereas c-src RNA did increase within 30 min of insulin infusion. RNA encoding c-myc transiently increased in both groups; however, this response was lower in insulin-resistant individuals than in insulin-sensitive individuals in a pattern similar to c-Ha-ras and myf-5. PPP1A RNA levels slightly increased in insulin-resistant individuals. In both insulin-sensitive and insulin-resistant persons, RNA quantities of GLUT4, c-jun, c-fos, and the insulin receptor did not change over the period of insulin infusion. However, overall RNA levels of the insulin receptor and c-jun were lower in insulin-resistant individuals. PMID: 8636261 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 520: Toxicol Appl Pharmacol. 1996 Feb;136(2):229-35. Metallothionein-I and -II knock-out mice are sensitive to cadmium-induced liver mRNA expression of c-jun and p53. Zheng H, Liu J, Choo KH, Michalska AE, Klaassen CD. Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City 66160-7417, USA. Zinc pretreatment has been shown in vitro (rat myoblasts) to induce metallothionein (MT) and inhibit cadmium (Cd)-induced protooncogenes c-myc and c-jun mRNA levels. therefore, the purpose of this study was to determine whether the mRNA expression of the protooncogene c-jun as well as the tumor suppressor gene p53 is increased by Cd in the intact animal and, more specifically, in the target organ for Cd toxicity, the liver. Additionally, modulation of the expression of these genes was investigated in the absence of MT. The effect of CdCl2 on the mRNA levels of c-jun and p53 was studied in livers of C57BL/6J (control) and MT-null mice by Northern- and slot-blot analyses. The mRNA for c-jun and p53 were increased by Cd in a dose-dependent fashion. In the control mice, Cd induced c-jun mRNA (5-fold) at 3 and 12 hr and p53 mRNA (1.8- to 2-fold) at 6 and 12 hr. Compared to controls, the MT-null mice were more sensitive to the Cd-induced gene expression. The magnitude of the inductions was more pronounced and the elevated mRNA levels of c-jun and p53 were seen at lower doses of Cd (10mumol/kg in MT-null mice vs 40 mmol/kg in control mice). In conclusion, these data demonstrate that Cd induces mRNA expression of the protooncogene c-jun and tumor suppressor gene p53 in liver, and that MT modulates this effect. PMID: 8619230 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 521: J Biol Chem. 1996 Jan 19;271(3):1695-701. ATF3 gene. Genomic organization, promoter, and regulation. Liang G, Wolfgang CD, Chen BP, Chen TH, Hai T. Ohio State Biochemistry Program, Ohio State University, Columbus 43210, USA. ATF3 gene, which encodes a member of the activating transcription factor/cAMP responsive element binding protein (ATF/CREB) family of transcription factors, is induced by many physiological stresses. As a step toward understanding the induction mechanisms, we isolated the human ATF3 gene and analyzed its genome organization and 5'-flanking region. We found that the human ATF3 mRNA is derived from four exons distributed over 15 kilobases. Sequence analysis of the 5'-flanking region revealed a consensus TATA box and a number of transcription factor binding sites including the AP-1, ATF/CRE, NF-kappa B, E2F, and Myc/Max binding sites. As another approach to understanding the mechanisms by which the ATF3 gene is induced by stress signals, we studied the regulation of the ATF3 gene in tissue culture cells by anisomycin, an approach that has been used to study the stress responses in tissue culture cells. We showed that anisomycin at a low concentration activates the ATF3 promoter and stabilizes the ATF3 mRNA. Significantly, co-transfection of DNAs expressing ATF2 and c-Jun activates the ATF3 promoter. A possible mechanism implicating the C-Jun NH2-terminal kinase/stress-activated protein kinase (JNK/SAPK) stress-inducible signaling pathway in the induction of the ATF3 gene is discussed. PMID: 8576171 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 522: EMBO J. 1996 Jan 15;15(2):319-33. GATA transcription factors associate with a novel class of nuclear bodies in erythroblasts and megakaryocytes. Elefanty AG, Antoniou M, Custodio N, Carmo-Fonseca M, Grosveld FG. National Institute for Medical Research, UK. The nuclear distribution of GATA transcription factors in murine haemopoietic cells was examined by indirect immunofluorescence. Specific bright foci of GATA-1 fluorescence were observed in erythroleukaemia cells and primary murine erythroblasts and megakaryocytes, in addition to diffuse nucleoplasmic localization. These foci, which were preferentially found adjacent to nucleoli or at the nuclear periphery, did not represent sites of active transcription or binding of GATA-1 to consensus sites in the beta-globin loci. Immunoelectron microscopy demonstrated the presence of intensely labelled structures likely to represent the GATA-1 foci seen by immunofluorescence. The GATA-1 nuclear bodies differed from previously described nuclear structures and there was no co-localization with nuclear antigens involved in RNA processing or other ubiquitous (Spl, c-Jun and TBP) or haemopoietic (NF-E2) transcription factors. Interestingly, GATA-2 and GATA-3 proteins also localized to the same nuclear bodies in cell lines co-expressing GATA-1 and -2 or GATA-1 and -3 gene products. This pattern of distribution is, thus far, unique to the GATA transcription factors and suggests a protein-protein interaction with other components of the nuclear bodies via the GATA zinc finger domain. PMID: 8617207 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 523: J Neurosci. 1996 Jan 15;16(2):836-42. Neonatal deprivation of maternal touch may suppress ornithine decarboxylase via downregulation of the proto-oncogenes c-myc and max. Wang S, Bartolome JV, Schanberg SM. Department of Pharmacology, Duke University, Durham, North Carolina 27710, USA. Previously, we have shown that short-term (1 hr) separation of neonatal rats from their mother (MS) suppresses basal ornithine decarboxylase (ODC) synthesis and tissue ODC response to trophic factors. This effect in the pup is caused by absence of maternal tactile stimulation (touch) but not from lack of maternal nutrients (food). This study was performed to examine in 10-d-old rats whether maternal touch deprivation affects expression of certain hepatic proto-oncogenes, the protein products of which are known to interact with the regulatory region of the ODC gene. Prolactin (PRL) injected subcutaneously increased hepatic ODC activity as well as mRNA levels of ODC and the proto-oncogenes c-fos, c-jun, junB, junD, c-myc, and max. MS significantly suppressed PRL-induced increases in ODC enzyme activity and c-myc, max, and ODC mRNAs but had little effect on expression of the other proto-oncogenes. PRL-induced stimulation of ODC, c-myc, and max mRNAs also was depressed in neonates placed with an anesthetized lactating dam (touch-deprived) but not in pups placed with nipple-ligated dams (food-deprived). Furthermore, unlike its effect on preweanling-age pups (< 20 d old), MS did not alter expression of either ODC or c-myc mRNAs in 25-d-old pups acutely separated from their mother. These findings indicate that suppression of ODC gene transcription in the neonatal pup during MS may be mediated by downregulation of the ODC gene transactivator proto-oncogenes c-myc and max. They are also consistent with our previous observation that lack of maternal touch, but not maternal milk, initiates the physiological alterations induced by MS. PMID: 8551363 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 524: Oncogene. 1996 Jan 4;12(1):135-42. Isolation and cloning of JTAP-1: a cathepsin like gene upregulated in response to V-Jun induced cell transformation. Hadman M, Gabos L, Loo M, Sehgal A, Bos TJ. Department of Microbiology and Immunology, Eastern Virginia Medical School, Norfolk, VA 23501, USA. The oncogenic potential of Jun in chicken embryo fibroblasts (CEF) varies depending on its structure. V-Jun, which has a number of structural differences from c-Jun is highly transforming and tumorigenic. C-Jun however, is only weakly transforming and is not tumorigenic. We have used this difference in oncogenic potential between v-Jun and c-Jun to screen for downstream target genes associated with the v-Jun induced transformed phenotype. We describe here the identification, cloning and characterization of one of these genes, JTAP-1. JTAP-1 is consistently overexpressed 7 to 10-fold in CEF transformed by v-Jun compared with c-Jun overexpressing or normal CEF. This pattern of expression suggests that JTAP-1 is associated with the transformed phenotype. DNA and amino acid homology search analysis revealed that JTAP-1 shares a high degree of similarity with over 100 cysteine proteases from a variety of species and is likely the chicken homolog of cathepsin O. Analysis of expression of JTAP-1 in CEF overexpressing other oncogenes including v-Ha-ras, v-Src, c-Fos, and Myc revealed that it's overexpression is unique to v-Jun transformed cells. Thus, JTAP-1 is likely a specific target of v-Jun overexpression and not simply a consequence of cell transformation. PMID: 8552384 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 525: Crit Rev Oncog. 1996;7(3-4):261-91. Oncogene-initiated aberrant signaling engenders the metastatic phenotype: synergistic transcription factor interactions are targets for cancer therapy. Denhardt DT. Department of Cell, Developmental and Neurobiology, Nelson Biological Laboratories, Rutgers University, Piscataway, NJ 08854, USA. Certain p21GTPases (notably Ras) and some of their guanine nucleotide exchange factors (e.g., Ost, Dbl, Tiam) and downstream mediators (e.g., Raf, Myc) have the potential to promote the development of malignancies because they can enhance the transcription of genes that foster the tumorigenic and metastatic phenotype. Among these are genes that stimulate cell proliferation, confer immortality, and facilitate the invasion of normal tissues. Oncogenes upstream of Ras-cell surface receptors such as ErbB2/Neu, Met, or Trk (and their ligands), and nonreceptor cytoplasmic protein tyrosine kinases such as Src and Abl-not only can act through Ras but also contribute additional signals. This review presents a synopsis of our understanding of signaling pathways controlled by the p21GTPases, with a focus on transcription factors regulated by the pathways. Mutations in one or more of the elements in these signaling pathways are invariably found in cancer cells. Crosstalk among the pathways may explain how some forms of stress can contribute to the development of a malignancy. Abnormal signaling leads to modified cytoskeletal structures and permanently altered (i.e., self-sustaining or epigenetic) transcription of target genes. A common therne is that genes whose transcription is elevated to the greatest extent by Ras often have in their promoters juxtaposed binding sites for two different transcription factors (particularly those in the Fos/Jun, CREB/ATF, NFkB, and Ets families) each of which is activated and such that together they synergize to augment transcription substantially. Some of these transcription factors can also act as oncogenes in certain cell types when appropriately modified and expressed. This unifying theme among many different cancers suggests that strategies to restore the balance among the signaling pathways or to suppress synergistic interactions between transcription factors may prove broadly useful in reversing the malignant phenotype. Publication Types: Review Review, Tutorial PMID: 9258606 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 526: Crit Rev Oncog. 1996;7(1-2):49-64. Oncogene- and tumor-suppressor gene-related proteins in plants and fungi. Loidl A, Loidl P. Department of Microbiology, University of Innsbruck-Medical School, Austria. Protooncogene- and tumor-suppressor gene proteins serve essential functions in the regulation of proliferation and differentiation of cells. Abnormal regulation or mutation of these genes, or transformation with retroviral homologs, may lead to tumor development in animals. In contrast to vertebrates, only few data on these genes exist in plants and fungi. Plant nuclear protooncogene homologs, such as myb and myc have multiple regulatory functions in metabolic pathways not existing in mammalian cells; they are involved in the complex regulation of anthocyanin (purple pigment) and phlobaphene (red pigment) biosynthesis, lignin production, trichome differentiation, dehydration stress gene expression and seed development. Apart from these well-characterized roles in plant-specific pathways, few experimental data have been reported on a functional significance in growth and development. A screening for nuclear protooncogene- and tumor-suppressor gene-related proteins in the myxomycete Physarum polycephalum revealed the existence of homologs of vertebrate c-myc, c-fos, c-jun, p53, and retinoblastoma proteins during the synchronous cell cycle or sclerotization. The p53 homologs of Physarum and Zea mays were shown to be specific for quiescent stages of their life cycles. Plants and lower eukaryotes, such as fungi, may be useful experimental systems to elucidate novel functions of protooncogene- and tumor-suppressor proteins in cell cycle regulation and development, or to reveal target genes that might be difficult to identify in complex mammalian systems. Recent data indicate that oncogenes and tumor suppressors in animals have more cellular targets than originally proposed; some of these might be as unexpected as in plant secondary metabolism. Publication Types: Review Review, Tutorial PMID: 9109497 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 527: Cancer Surv. 1996;27:25-40. Coupling of the IL2 receptor complex with non-receptor protein tyrosine kinases. Miyazaki T, Taniguchi T. Department of Immunology, Faculty of Medicine, University of Tokyo. IL2 induces the proliferation of T lymphocytes through the IL2 receptor (IL2R) following T lymphocyte activation. The IL2R consists of at least three subunits, IL2R alpha, beta and gamma chains. The cytoplasmic regions of the IL2R beta and gamma chains are critical for transduction of the IL2 signal to the cell interior. Although IL2R beta and gamma chains lack an intrinsic protein tyrosine kinase (PTK) domain, these chains recruit various non-receptor type PTKs, such as p56lck (and other Src family PTKs), Jak PTKs and Syk PTKs. The recruited PTKs are then activated following ligand stimulation to invoke intracellular signalling for the cell proliferation. Furthermore, it has been demonstrated that the IL2R is linked to at least three distinct signalling pathways leading to the induction of the c-fos/c-jun genes, c-myc gene induction and bcl-2 gene induction. All these pathways are essential for IL2 mediated proliferative signalling and co-operate with each other to ensure a full scale signal transduction. These signalling pathways, except that for bcl-2 pathway, appear to be mediated by multiple PTKs: p56lck is critical for the induction of the c-fos/c-jun genes, the activation of Syk PTKs results in the induction of the c-myc gene and Jak3 PTK is required for the induction of both c-fos and c-myc genes. Finally, the IL2 system may serve as a prototype in understanding the pleiotropic function of cytokine receptors that lack intrinsic PTK domains; the cytoplasmic structures of these cytokine receptors have evolved to allow the combined action of different PTK family members (and other signalling molecules) expressed in different cell types, which may determine the activity of cytokines. Publication Types: Review Review, Tutorial PMID: 8909793 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 528: Nutr Cancer. 1996;25(1):9-26. RRR-alpha-tocopheryl succinate induces apoptosis in avian retrovirus-transformed lymphoid cells. Qian M, Sanders BG, Kline K. Genetics Institute, University of Texas at Austin 78712-1097, USA. The RRR-alpha-tocopheryl succinate form of vitamin E [vitamin E succinate (VES)] inhibits the proliferation of avian reticuloendotheliosis virus-transformed RECC-UTC4-1 (C4-1) lymphoblastoid cells in a dose-dependent manner, blocks the cells in the G2/M cell cycle phase, and induces the cells to undergo apoptosis. Apoptosis was documented by demonstrating changes that are characteristic of this type of cell death, including morphological analyses of chromatin condensation by 4',6-diamidine-2'-phenylindole dihydrochloride (DAPI) staining using scanning confocal and traditional fluorescent microscopy; flow cytometry analyses of propidium iodide-labeled DNA showing fragmented DNA as a pre-G1 peak; two-color flow cytometry analyses of intact cells labeled first by the TUNEL procedure (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end-labeled DNA stained with fluorescein isothiocyanate-labeled avidin) and then by propidium iodide demonstrating fragmented DNA; and electrophoresis of DNA showing a DNA ladder created by internucleosomal DNA fragmentation. The percentage of apoptotic cells was determined by DAPI staining and showed 11%, 27%, and 49% of cells to be apoptotic after treatment with 10 micrograms/ml VES for one, two, and three days, respectively. Analyses of mRNA levels of genes that have been implicated in the apoptotic process, namely, bcl-2, c-myc, and c-jun, revealed no change in bcl-2, decreases in c-myc mRNA levels after 36 hours of treatment, and increases in c-jun mRNA levels within four hours after treatment. Western immunoblotting analyses of protein levels for the transcription factors c-Myc and c-Jun showed normal levels of c-Myc at early time points and decreased levels at 24 and 48 hours after treatment. c-Jun increased as early as 6 hours after treatment and returned to lower (yet still elevated over control) levels by 48 hours. To determine possible functional consequences of increased c-Jun expression, gel electrophoretic mobility assays were conducted that showed increased AP-1 binding at 24 and 48 hours after treatment. These data show that VES induces apoptosis in reticuloendotheliosis virus-transformed lymphoid cells and suggest that decreases of c-Myc protein and increases of c-Jun protein and DNA binding capacity may be playing a role in VES-mediated events leading to apoptosis in this cell type. PMID: 8837858 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 529: J Cell Biochem Suppl. 1996;24:218-27. Regulation of nuclear oncogenes expressed in lung cancer cell lines. Sabichi AL, Birrer MJ. Biomarkers and Prevention Research Branch, National Cancer Institute, Rockville, Maryland 20850, USA. Lung cancer is a major cause of mortality in the United States and accounts for the majority of all cancer deaths in both men and women. It is hoped that through broadening our understanding of the mechanisms involved in transformation of bronchial epithelial cells we will be able to improve methods of diagnosis and treatment of this disease, with the ultimate goal of reducing on lung cancer mortality. A knowledge of the molecular mechanisms involved in processes such as cell division and differentiation is paramount to this task, because it is known that aberrant responses to growth factors or cytokines found in the normal cellular milieu can lead to abnormal cell growth and/or transformation. Signals initiated at the cell membrane by tumor promoters, growth factors, or cytokines are transduced from the cell membrane to the nucleus and are, in part, mediated centrally by transcription factors encoded by nuclear protooncogenes. The transcription factors myc, jun, and fos have been characterized in both normal and transformed lung epithelial cells through detailed studies using cell lines. In this manuscript, we review what is known about the expression and regulation of these nuclear protooncogenes in normal and malignant epithelial cells of the lung, and their role in the development of lung cancer. Publication Types: Review Review, Tutorial PMID: 8806104 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 530: Cell Growth Differ. 1996 Jan;7(1):75-81. Antagonistic actions of phorbol ester in mammalian G0-->G1-->S cell cycle transition. Faria M, Armelin HA. Departamento de Bioquimica, Universidade de Sao Paulo, SP, Brazil. We have developed a protocol that reveals two antagonistic effects of phorbol-12-myristate-12-acetate (PMA) on the G0-->G1-->S transition of mammalian cell cycle. Balb-3T3 (Clone A31) cells arrested in G0 by serum starvation can be stimulated to traverse the G1 phase and initiate DNA synthesis 12 h later by a 2-h pulse with PMA. In contrast with this early stimulatory effect, PMA has an inhibitory effect when presented to the cells during the last 6 h of G1. PMA is able to inhibit DNA synthesis initiation irrespective of the triggering agent, i.e., serum, fibroblast growth factor, epidermal growth factor, platelet-derived growth factor, or PMA itself (presented as an early pulse). We have established that the critical period for the PMA inhibitory effect is between 6 and 8 h after cell stimulation. This dual effect of PMA is not a peculiarity of Balb-3T3 (clone A31) cells because it is also observed with other fibroblastic cell lines, namely, SWISS 3T3, NIL 8, and RAT 1, and also with the epithelial Y-1 adrenocortical cell line. Treatment with PMA for 0.5 or 2 h activates protein kinase C (PKC) in Balb-3T3-A31 cells, but is not sufficient to down-regulate the enzyme because a second 30-min PMA pulse applied between 6 and 6.5 h activates PKC again. On the other hand, a continuous 6.5-h PMA treatment causes PKC down-regulation; therefore, the inhibitory effect of PMA could be mediated by PKC. Growth factor early response proto-oncogenes c-myc, c-fos, and c-jun are induced transiently by both early and late PMA pulses, suggesting that these genes are not involved in the PMA inhibitory effect. PMID: 8788035 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 531: Arch Dermatol Res. 1996;288(1):2-6. Bradykinin upregulates immediate-early gene mRNA in human keratinocytes. Coutant KD, Ryder NS. Sandoz Research Institute, Dermatology Department, Vienna, Austria. We investigated the effect of bradykinin on the expression of the proto-oncogenes c-fos, c-jun and c-myc and on cell proliferation in the human keratinocyte cell line, HaCaT. Analysis of mRNAs was done by Northern blotting with single-stranded DNA hybridization probes. Bradykinin caused a rapid and transient accumulation in c-fos and c-jun mRNAs. In contrast, c-myc mRNA increased more slowly. Moreover, we report that bradykinin was a weak stimulator of HaCaT cell proliferation whereas epidermal growth factor, which induced the same degree of mRNA elevation, was shown as a powerful mitogen. Thus, while in HaCaT cells bradykinin promotes the expression of the proto-oncogenes c-fos, c-jun and c-myc, other biochemical events appear to be necessary for cell division. PMID: 8750926 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 532: Exp Clin Endocrinol Diabetes. 1996;104(2):111-22. Expression of estrogen receptor and proto-oncogene messenger ribonucleic acids in reproductive tracts of neonatally diethylstilbestrol-exposed female mice with or without post-puberal estrogen administration. Kamiya K, Sato T, Nishimura N, Goto Y, Kano K, Iguchi T. Graduate School of Integrated Science, Yokohama City University, Japan. Perinatal treatment of female mice with natural and synthetic estrogens including diethylstilbestrol (DES) results in estrogen-independent persistent proliferation and cornification of the vaginal epithelium. The dynamics of the induction of estrogen receptor (ER), c-jun, c-fos and c-myc mRNAs by 17 beta-estradiol (E2) was examined in the uterus and vagina of neonatally DES-exposed and -unexposed ovariectomized adult mice. In the uterus of neonatally DES-unexposed ovariectomized mice, the expression of ER mRNA increased within 1 h after E2 administration and declined by 12 h thereafter. ER mRNA in the vagina decreased within 1 h after the stimulation and recovered by 12 h thereafter. In the uterus, c-jun and c-fos mRNAs increased in concentration within 1 h after E2 administration, showing a peak 3 h after the stimulation; they decreased with time thereafter. In the vagina, the concentration of c-jun and c-fos mRNAs increased rapidly, reaching a peak within 1 h after the stimulation. However, the expression of c-myc in uterus and vagina was not changed by postpuberal E2. These results suggest that estrogen regulation of ER and proto-oncogene mRNAs in the vagina differs from those in the uterus. In the neonatally DES-exposed ovariectomized adult mice, uterine ER mRNA expression levels were significantly higher than in the unexposed ovariectomized controls; however, vaginal levels were drastically lower than in the controls. Expression of c-jun and c-fos mRNAs was greater in both the uterus (3- and 6-fold, respectively) and the vagina (18- and 4-fold) of neonatally DES-exposed mice than in controls. The ER mRNA and the increased levels of c-fos and c-jun mRNAs in both uterus and vagina of neonatally DES-exposed ovariectomized mice were not further altered by post-puberal E2 and may be related to ovary-independent persistent changes in the genital tract. PMID: 8740934 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 533: Brain Res Mol Brain Res. 1996 Jan;35(1-2):19-30. Gamma-radiation-induced cell death in the fetal rat brain possesses molecular characteristics of apoptosis and is associated with specific messenger RNA elevations. Borovitskaya AE, Evtushenko VI, Sabol SL. Laboratory of Biochemical Genetics, National Heart, Lung, and Blood Institute, Bethesda, MD 20892, USA. Low-dose ionizing irradiation of 16-18-day pregnant rats rapidly kills stem cells in the fetal forebrain. We have examined gamma-irradiated 17-day fetal rat brain tissue for molecular characteristics of apoptosis and changes in levels of mRNAs relevant to apoptosis. In many forebrain cells radiation elicits within 5 h nuclear condensation and fragmentation consistent with apoptosis. An electrophoretic DNA ladder indicative of internucleosomal chromatin cleavage was prominent within 3 h after irradiation. Pretreatment of pregnant rats with cycloheximide, or pretreatment of dissociated fetal brain cells in culture with actinomycin D, abolished the radiation-induced internucleosomal DNA fragmentation, demonstrating requirements for protein and RNA synthesis. Irradiation dramatically increased the level of the p53 transcription factor and the abundances of mRNAs coding for the cell-cycle inhibitor p21/Waf-1/Cip-1 and the AP-1-associated transcription factors Fos and JunB. Irradiation moderately increased the level of mRNA for the positive apoptosis regulator Bax. In contrast, irradiation reduced by 50-70% the abundances of most other mRNAs tested, including those for housekeeping proteins, p53, Jun, Myc, interleukin-1-beta-converting enzyme, and the negative apoptosis regulators Bcl-2 and Bcl-xL. These results indicate that radiation-elicited apoptosis of fetal brain cells is associated with activation of the p53 system, probable increases in AP-1 Fos/JunB heterodimers, and an increased ratio of Bax to Bcl-2 + Bcl-xL. PMID: 8717336 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 534: Recent Prog Horm Res. 1996;51:63-96. A nuclear matrix acceptor site for the progesterone receptor in the avian c-myc gene promoter. Spelsberg TC, Lauber AH, Sandhu NP, Subramaniam M. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905, USA. It has been the goal of this project to determine the location, composition, and biological function of the nuclear acceptor sites (i.e., the nuclear binding sites) for the avian oviduct progesterone (Pg) receptor (PR). Many laboratories have demonstrated a native-(in vivo) like cell-free binding of steroid receptor complexes to specific acceptor sites in nuclei/chromatin in a variety of target tissue systems. These sites appear to involve protein-DNA complexes and some of these have been shown to reside in the nuclear matrix, including the chromatin acceptor sites for the avian oviduct PR. We have purified a nuclear matrix "acceptor protein" for the avian PR. termed receptor binding factor-1 (RBF-1), based on its ability to generate specific, high-affinity PR binding on avian genomic DNA. This 10 kD nuclear matrix protein was found to be unique with minimal homology to a couple of other proteins. Using immunohistochemical techniques and antibodies against the purified RBF-1, the RBF-1 was localized to the nuclei of many avian and rat tissues. Co-localizations of RBF-1 and PR in select cell types in the avian oviduct and rat reproductive organs were also reported. A tissue specificity was found with regard to RBF-1 concentrations. The full length cDNA to RBF-1 has been isolated and used to identify a 0.7 kb mRNA whose levels in various avian tissues reflect the protein levels. Genomic sequences of RBF-1 have been isolated and characterized. Preliminary studies indicate that the over-expression of the RBF in human MCF-7 cells results in an inhibition of the c-myc gene promoter activity which is further inhibited by steroid hormone treatments of the cells. Past studies in our laboratory demonstrated that the c-myc mRNA steady state levels are rapidly (approximately 15 min) reduced by Pg and glucocorticoids in the avian oviduct. Further, partially purified fractions of RBF-1 were shown to generate specific PR binding sites only on the genomic DNAs of certain animal species and on the c-myc gene, but not ovalbumin gene. Using Southwestern blot analyses, the purified RBF-1 was shown to bind specifically to sequences in the 5' end of c-myc, c-jun proto-oncogenes but not to genomic sequences of the ovalbumin gene. A specific DNA binding element in the promoter region of the c-myc proto-oncogene has been identified as AT-rich domain of homopurine/pyrimidine stretches flanked by GC-rich sequences. Southern blot analyses using 200 bp matrix DNA fragments protected by the nuclear matrix structure indicate that the matrix is attached on either side of the RBF-1 binding element. A model is described for a nuclear matrix acceptor site attached to the c-myc promoter which may mediate the Pg-induced down-regulation of the c-myc gene expression. Publication Types: Review PMID: 8701093 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 535: Clin Exp Rheumatol. 1996 Jan-Feb;14(1):87-94. Sex hormones, proto-oncogene expression and apoptosis: their effects on rheumatoid synovial tissue. Cutolo M, Sulli A, Barone A, Seriolo B, Accardo S. Department of Internal Medicine, University of Genova, Italy. Programmed cell death (apoptosis), is a non-random physiological process characterized by cell fragmentation without leakage of the cellular contents into the extracellular space. Apoptosis is especially important in the immune system. On the other hand the capacity of cells to proliferate and to show local invasiveness, as in cancer cells or the "tumor-like" synoviocytes in rheumatoid arthritis (RA), seems to be controlled by a group of genes called "proto-oncogenes". The early metabolic events in cell apoptosis and proliferation are remarkably similar. The primary location of apoptotic cells in RA synovial tissue is at the level of the synovial lining, varying from rare positive cells to > 50% positive cells. C-jun, c-fos and c-myc oncoproteins seem to be largely restricted to the synovial cells attached to the sites of cartilage and bone destruction. Ovarian follicle atresia could serve as a useful model to study the hormonal regulation of apoptosis in different endocrine tissues. Based on ovarian studies it seems that estrogens generally prevent apoptosis whereas androgens induce apoptosis. The binding of steroids to their receptors forms a complex wherein the receptors are transformed, so that they can then pass through the nuclear membrane and associate with specific recognition sites on DNA. In the majority of cases, the steroid receptors mediate the rapid regulation of the nuclear proto-oncogene transcription. Therefore, they may serve as important "early" regulatory genes and as excellent universal markers in all tissues in steroid hormone action. Since the macrophages are considered to be target cells for sex hormones, we recently evaluated c-myc expression in cytocentrifuge preparations obtained from primary cultures of RA synovial macrophages treated with estrogens, and observed a marked upregulation. Further studies of the influence of sex hormones on synoviocyte apoptosis and proto-oncogene expression should offer new perspectives on the pathogenesis and therapy of synovitis in RA and other rheumatic diseases. Publication Types: Review Review, Tutorial PMID: 8697665 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 536: Anticancer Res. 1996 Jan-Feb;16(1):401-6. Differential EGF action on nuclear protooncogenes in human endometrial carcinoma RL95-2 cells. Sallot M, Ordener C, Lascombe I, Propper A, Adessi GL, Jouvenot M. Service de Biochimie-Biologie, Moleculaire, Institut d'Etude et de Transfert de Genes, Besancon, France. Our aim was to analyze EGF action on nuclear protooncogenes in RL95-2 since it has not been documented so far. Synchronization and partial' growth arrest were obtained by maintaining cells for 15 hours in L-methionine-free medium. After this depletion, EGF transiently increased fos and jun mRNAs: the expression peaked at 45 minutes for c-fos (5.5 fold induction) and at 60 minutes for c-jun and jun-B (3 fold induction) and the mRNA levels returned to the basal value within 3 hours. Upon EGF addition, c-myc mRNAs peaked at 12 hours (7.6 fold induction) and surprisingly remained higher than the control up to 48 hours. Unlike fetal calf serum, EGF did not increase the cell number and this could be linked to steadily induced c-myc expression. These data provide evidence for a differential EGF action on fos/jun and c-myc in RL95-2 cells. PMID: 8615644 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 537: Mol Cell Biochem. 1995 Dec 6-20;153(1-2):59-67. Unique and selective mitogenic effects of vanadate on SV40-transformed cells. Wang H, Scott RE. Department of Pathology, The University of Tennessee College of Medicine, Memphis, Tennessee 38163, USA. Vanadate and insulin both function as unique complete mitogens for SV40-transformed 3T3T cells, designated CSV3-1, but not for nontransformed 3T3T cells. The mitogenic effects induced by vanadate and insulin in CSV3-1 cells are mediated by different signaling mechanisms. For example, vanadate does not stimulate the tyrosine phosphorylation of the insulin receptor beta-subunit nor the 170 kDa insulin receptor substrate-1. Instead, vanadate induces a marked increase in tyrosine phosphorylation of 55 and 64 kDa proteins that is not observed in insulin-stimulated CSV3-1 cells. Perhaps most interestingly, vandate-induced mitogenesis is associated with the selective induction of c-jun and junB expression without significantly inducing c-fos or c-myc. Furthermore, treatment of CSV3-1 cells with genistein abolishes the effects of vanadate on protein tyrosine phosphorylation and c-jun induction. These and related data suggest that modulation of protein tyrosine phosphorylation and c-jun and junB expression may serve the critical roles in mediating vandate-induced mitogenesis in SV40-transformed cells. Publication Types: Review Review, Tutorial PMID: 8927049 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 538: Gen Pharmacol. 1995 Dec;26(8):1643-9. Mitogenic signal transduction in human breast cancer cells. Strobl JS, Wonderlin WF, Flynn DC. Department of Pharmacology & Toxicology, West Virginia University, Morgantown 26506, USA. 1. Signal transduction pathways activated during growth of human breast cancer cells in tissue culture are reviewed. 2. Steroid hormones and growth factors stimulate similar mitogenic pathways and frequently modulate each other's activity. 3. A response common to estrogen, progestins and most polypeptide mitogens is induction of the nuclear transcription factors myc, fos and jun in early G1 phase of the cell cycle. 4. Some growth factors also stimulate cyclin D1, a regulatory protein responsible for the activation of cell cycle-dependent kinases in G1. 5. In addition, insulin, IGF-I and EGF activate tyrosine kinase receptors. 6. Several tyrosine phosphorylated proteins occur in human breast cancer cells, and include the EGF and estrogen receptors. 7. Cyclic AMP plays a critical role in breast cancer cell proliferation through the activation of protein kinase A, and it also modulates the activity of estrogen and progesterone receptors. 8. EGF is the only breast cell mitogen known to raise intracellular free calcium levels. 9. Calcium may play a dual role in breast cancer cell proliferation, activating both calmodulin-dependent processes and regulating cell membrane potential through the activation of potassium channels. 10. Potassium channel activity and cell proliferation are linked in breast cancer cells, the cell membrane potential shifting between a depolarized state in G1/G0 cells and a hyperpolarized state during S phase. 11. Activation of an ATP-sensitive potassium channel is required for breast cancer cells to undergo the G1/G0-S transition. Publication Types: Review Review, Tutorial PMID: 8745151 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 539: Eur J Cell Biol. 1995 Dec;68(4):457-62. Hypertonic stress induces c-fos but not c-jun expression in the human embryonal EUE epithelial cell line. Rossi D, Fuhrman Conti AM, De Grada L, Larizza L. Department of Biology and Genetics, University of Milan, Italy. Recent evidence has indicated a role for the two early response genes c-fos and c-jun in transcriptional regulation of genes acting in osmoregulation. On this basis we investigated their expression in response to hypertonic stress in the human embryonal EUE epithelial cell line. EUE cells have proven to be a useful tool for studying long-term in vitro adaptation to hypertonic stress. After culturing EUE cells in hypertonic medium a marked c-fos induction was observed, both at the mRNA and the protein level. Northern analysis of fos-mRNA showed a peak expression at 4 h, followed by a progressive decline till complete extinction at 8 h. Immunofluorescence analysis of FOS protein evidenced a similar, although slightly delayed kinetics of expression. Conversely, neither c-jun nor c-myc up-regulation could be detected. The treatment of EUE cells with cycloheximide led to superinduction of c-fos expression, (with high levels up to 12 h), and to a c-jun expression that was just detectable. Hypertonic stimulation of the transformed cell lines A549, MCF7 and JR induced both c-fos and c-jun only in JR cells. Hypertonic shock was also effective in inducing c-fos expression in fetal human diploid fibroblasts, although the response was earlier and more transient than in EUE cells. These findings indicate that c-fos is a primary response gene in hypertonic stress-activated cells, although the pattern and kinetics of its induction may differ according to the type of cell. PMID: 8690026 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 540: J Clin Invest. 1995 Dec;96(6):2768-74. Alteration of growth responses in established cardiac pressure overload hypertrophy in rats with aortic banding. Schunkert H, Weinberg EO, Bruckschlegel G, Riegger AJ, Lorell BH. The Charles A. Dana Research Institute and Harvard-Thorndike Laboratory, Beth Israel Hospital, Boston, Massachusetts 02215, USA. We examined the acute effects of elevated wall stress, norepinephrine, and angiotensin II on cardiac protein synthesis as well as protooncogene expression in hearts with established pressure overload left ventricular hypertrophy. Isolated rat hearts with chronic hypertrophy (LVH) were studied 12 wk after ascending aortic banding when systolic function was fully maintained. New protein synthesis (incorporation of [3H]phenylalanine [Phe]) was analyzed in isolated perfused rat hearts after a 3-h protocol; c-fos, c-jun, c-myc, and early growth response gene-1 (EGR-1) mRNA levels (Northern blot) were studied over a time course from 15 to 240 min of perfusion. Under baseline conditions (i.e., before mechanical or neurohormonal stimulation), [3H]-Phe-incorporation (280 nmoles/gram protein/h) and protooncogene mRNA levels were similar in age-matched control and LVH hearts. However, hearts with chronic LVH were characterized by a markedly blunted or absent [3H]-Phe-incorporation after acute imposition of isovolumic systolic load (90 mmHg/gram left ventricle), as well as norepinephrine (10(-6)M), or angiotensin II infusion (10(-8)M plus prazosin 10(-7)M) compared with nonhypertrophied control hearts. Similarly, stimulation of LVH hearts with acute systolic load or norepinephrine was associated with a significantly blunted increase of protooncogene mRNA levels relative to control hearts. The blunted induction of c-fos mRNA in LVH hearts was not due to feedback inhibition, since cycloheximide perfusion of hearts exposed to elevated wall stress further increased the differences between age-matched control and LVH hearts. The data suggest that acute molecular growth responses to mechanical or neurohormonal stimulation are altered in rat hearts with established LVH relative to nonhypertrophied control hearts. This alteration of molecular adaptations in hearts with compensatory hypertrophy may prevent inappropriate excess cardiac growth in response to mechanical and neurohormonal stimuli. PMID: 8675646 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 541: Cell Prolif. 1995 Dec;28(12):631-44. Transient inhibition of protein synthesis induces expression of proto-oncogenes and stimulates resting cells to enter the cell cycle. Rosenwald IB, Setkov NA, Kazakov VN, Chen JJ, Ryazanov AG, London IM, Epifanova OI. Harvard-MIT Division of Health Sciences and Technology, Cambridge, USA. There is evidence that resting cells are able to produce molecules with antiproliferative activity, some of which behave as short-lived repressor proteins. We suggest that transient inhibition of protein synthesis in resting cells would lead to a decrease in the levels of these negative growth regulators and might, therefore, promote mitogenic responses. We report that treatment of resting (serum-deprived) NIH 3T3 cells with cyclocheximide (CH) or puromycin induces expression of c-fos, c-jun and c-myc proto-oncogenes in a manner similar to that of platelet-derived growth factor (PDGF). Actinomycin D (Act D) abrogates the induction of proto-oncogene expression. Transient inhibition of protein synthesis by CH or puromycin also induces the resting NIH 3T3 and C3H 1OT1/2 cells to enter the cell cycle. Inhibition of new RNA or protein synthesis abolishes the proliferative response. These findings show that control mechanisms at both transcriptional and translational levels are operative in the resting cells treated with protein synthesis inhibitors. Cell fusion experiments with resting and serum-stimulated NIH 3T3 cells revealed that brief pre-incubation of resting cells with either PDGF, CH or puromycin abrogates their ability to suppress the onset of DNA synthesis in the nuclei of stimulated cells in heterodikaryons. However, the abrogative effect of PDGF disappeared in the presence of Act D, whereas the effects of protein synthesis inhibitors did not, indicating their independence of the induction of transcription. The data suggest that the observed effects of protein synthesis inhibitors are connected with elimination of some short-lived negative growth regulators, since a brief translational arrest is sufficient for the resumption of DNA synthesis in the nuclei of stimulated cells blocked by resting cells in heterodikaryons. PMID: 8634371 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 542: Mol Reprod Dev. 1995 Dec;42(4):459-67. Transcriptional regulation by MAP kinases. Davis RJ. Department of Biochemistry and Molecular Biology, Howard Hughes Medical Institute, University of Massachusetts Medical Center, Worcester, MA 01605, USA. Tyrosine kinase growth factor receptors activate MAP kinase by a complex mechanism involving the SH2/3 protein Grb2, the exchange protein Sos, and Ras. The GTP-bound Ras protein binds to the Raf kinase and initiates a protein kinase cascade that leads to MAP kinase activation. Three MAP kinase kinase kinases have been described--c-Raf, c-Mos, and Mekk--that phosphorylate and activate Mek, the MAP kinase kinase. Activated Mek phosphorylates and activates MAP kinase. Subsequently, the activated MAP kinase translocates into the nucleus where many of the physiological targets of the MAP kinase signal transduction pathway are located. These substrates include transcription factors that are regulated by MAP kinase phosphorylation (e.g., Elk-1, c-Myc, c-Jun, c-Fos, and C/EBP beta). Thus the MAP kinase pathway represents a significant mechanism of signal transduction by growth factor receptors from the cell surface to the nucleus that results in the regulation of gene expression. Three MAP kinase homologs have been identified in the rat: Erk1, Erk2, and Erk3. Human MAP kinases that are similar to the rat Erk kinases have also been identified by molecular cloning. The human Erk1 protein kinase has been shown to be widely expressed as a 44-kDa protein in many tissues. The human Erk2 protein kinase is a 41-kDa protein that is expressed ubiquitously. In contrast, a human Erk3-related protein kinase has been found to be expressed at a high level only in heart muscle and brain. The loci of these MAP kinase genes are widely distributed within the human genome: erk2 at 22q11.2; erk1 at 16p11.2; and ek3-related at 18q12-21. In the yeast Saccharomyces cerevisiae, five MAP kinase gene homologs have been described: smkl, mpk1, hog1, fus3, and kss1. Together, these kinases are a more diverse group than the human erks that have been identified. Thus the erks are likely to represent only one subgroup of a larger human MAP kinase gene family. A candidate for this extended family of MAP kinases is the c-Jun NH2-terminal kinase (Jnk), which binds to and phosphorylates the transcription factor c-Jun at the activating sites Ser-63 and Ser-73. Evidence is presented here to demonstrate that Jnk is a distant relative of the MAP kinase group that is activated by dual phosphorylation at Tyr and Thr. Publication Types: Review Review, Tutorial PMID: 8607977 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 543: Development. 1995 Dec;121(12):3969-77. Differential regulation of AP-1 and novel TRE-specific DNA-binding complexes during differentiation of oligodendrocyte-type-2-astrocyte (O-2A) progenitor cells. Barnett SC, Rosario M, Doyle A, Kilbey A, Lovatt A, Gillespie DA. University Department of Neurology, CRC Beatson Laboratories, Glasgow, UK. AP-1 is an ubiquitous transcription factor which is composed of the Jun and Fos proto-oncogene proteins and is thought to play a role in both cell proliferation and differentiation. We have used an immortal, bipotential oligodendrocyte-type-2 astrocyte progenitor cell line (O-2A/c-myc) which can differentiate into oligodendrocytes or type-2 astrocytes in vitro, to investigate whether AP-1 DNA-binding activity fluctuates during glial cell differentiation. Unexpectedly, DNA-mobility shift assays using a TRE-containing oligonucleotide derived from the promoter of the glial-specific gene, glial fibrillary acidic protein (GFAP/AP-1), revealed that O-2A/c-myc progenitor cells were devoid of conventional AP-1 DNA-binding complexes. O-2A/c-myc cells did however contain several novel GFAP/AP-1-specific DNA-binding complexes, which we have termed APprog. APprog complexes recognise the TRE consensus motif present in the GFAP/AP-1 oligonucleotide together with adjacent 3' sequences but do not contain c-Jun or any other known Jun-related proteins. When O-2A/c-myc cells underwent terminal differentiation APprog complexes were lost and conventional AP-1 DNA-binding activity became evident, particularly in astrocytes. These changes appear to be closely linked to the differentiation process since they did not occur in a derivative of the O-2A/c-myc cell line that contains an activated v-ras oncogene and which fails to differentiate under appropriate culture conditions. The inverse regulation of conventional AP-1 and APprog complexes within the O-2A lineage suggests that these factors may play a role in the regulation of glial cell differentiation or glial cell-specific gene expression. PMID: 8575297 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 544: J Steroid Biochem Mol Biol. 1995 Dec;55(3-4):279-89. SKOV3 ovarian carcinoma cells have functional estrogen receptor but are growth-resistant to estrogen and antiestrogens. Hua W, Christianson T, Rougeot C, Rochefort H, Clinton GM. Department of Molecular and Medical Genetics, Oregon Health Sciences University, Portland 97201, USA. Estrogen receptor positive ovarian cancer is often refractile to antiestrogen therapy. Here we describe the SKOV3 human ovarian carcinoma cell line as an in vitro model for estrogen and antiestrogen resistant ovarian cancer. While SKOV3 cells expressed estrogen receptor (ER) mRNA and protein at a similar level as the estrogen responsive T47D breast carcinoma cell line, their growth was not responsive to estradiol (E2) and was not inhibited by the antiestrogens OH-tamoxifen and ICI 164,384. The ER in SKOV3 cells was normal with respect to apparent Kd for binding with E2, E2 regulation of a transiently transfected ERE driven reporter gene, and E2 stimulation of expression of the early growth response genes c-myc and c-fos. However, the SKOV3 cells exhibited no expression of the progesterone receptor gene (PR) even after addition of E2, and the protein products of the estrogen responsive genes HER-2/neu and cathepsin D were expressed at constitutive levels that were not regulated by E2. Therefore, estrogen resistance in these cells may be a result of constitutive expression and loss of E2 regulation of selected growth regulatory gene products rather than a defect in estrogen activation of ER as a transcriptional regulator. PMID: 8541224 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 545: J Cell Physiol. 1995 Dec;165(3):459-67. Growth control and gene expression in a new hepatocellular carcinoma cell line, Hep40: inhibitory actions of vitamin K. Bouzahzah B, Nishikawa Y, Simon D, Carr BI. Pittsburgh Transplantation Institute, Department of Surgery, University of Pittsburgh, Pennsylvania 15213, USA. The growth characteristics of a newly established cell line, Hep40, derived from a human hepatoma are described. An absolute requirement was found for serum to mediate cell growth. Neither EGF, TGF-alpha, nor HGF altered cell growth in the presence or absence of serum. A partial suppression of cell growth was achieved by several TGF-beta family proteins. Affinity crosslinking gels using 125I-labeled TGF-beta showed a significant decrease in the TGF-beta cell-surface type II receptor in Hep40 cells, compared to the TGF-beta-sensitive Hep3B cell line. However, growth could be completely suppressed by addition of vitamins K to the culture medium in both Hep40 and several other hepatoma cell lines. Growth suppression by vitamins K was accompanied by an increased level of transcripts for c-myc, c-jun, and prothrombin genes, in contrast to the actions of TGF-beta 1 protein, which caused a decrease in the level of c-myc transcripts. These data show that this new human hepatoma cell line has partial resistance to growth inhibition by TGF-beta with a unique TGF-beta receptor defect. However, growth was completely suppressed by vitamins K. The differing gene expression patterns in response to TGF-beta as compared to vitamin K suggest that these two growth inhibitors act through differing pathways. PMID: 7593224 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 546: Arterioscler Thromb Vasc Biol. 1995 Dec;15(12):2273-83. Sodium butyrate inhibits platelet-derived growth factor-induced proliferation of vascular smooth muscle cells. Ranganna K, Joshi T, Yatsu FM. Department of Neurology, University of Texas Health Science Center at Houston 77030, USA. Sodium butyrate (SB), a naturally occurring short-chain fatty acid, was investigated for its therapeutic value as an antiproliferative agent for vascular smooth muscle cells (SMCs). At 5-mmol/L concentration, SB had no significant effect on rat SMC proliferation. However, at the same concentration, SB inhibited platelet-derived growth factor (PDGF)-AA-, -AB-, and -BB-induced proliferation of SMCs. Exposure of SMCs to PDGF-BB resulted in activation of receptor intrinsic tyrosine kinase activity and autophosphorylation of beta-PDGF-receptor (beta-PDGFR). The activated beta-PDGFR physically associated and phosphorylated signaling molecules such as ras-GTPase activating protein (GAP) and phospholipase C gamma (PLC gamma). SB, in the absence of PDGF-BB, caused neither beta-PDGFR tyrosine phosphorylation nor phosphorylation and association of GAP and PLC gamma with beta-PDGFR. PDGF-BB-enhanced activation of receptor intrinsic tyrosine kinase activity and autophosphorylation of tyrosine residues of beta-PDGFR were unaffected by SB irrespective of whether SMCs were preincubated with SB before exposure to PDGF-BB plus SB or incubated concomitantly with PDGF-BB plus SB. Likewise, phosphorylation and association of GAP and PLC gamma with PDGF-BB-activated beta-PDGFR were unaffected. In addition, SB did not block PDGF-BB-stimulated, PLC gamma-mediated production of inositol triphosphate. Similarly, PDGF-BB-induced beta-PDGFR degradation was unaffected when SMCs were exposed to PDGF-BB plus SB, and SB by itself had no influence on beta-PDGFR degradation. Unlike beta-PDGFR kinase activity, mitogen-activated protein kinase (MAP-kinase) activity was stimulated by SB by about 2.7-fold. Exposure of SMCs to PDGF-BB caused an approximately 11.4-fold increase in MAP-kinase activity and this increase in activity was not significantly affected when cells were coincubated with PDGF-BB and SB (10.3-fold). However, pretreatment of SMCs with SB for 30 minutes and subsequent incubation in PDGF-BB plus SB abolished most of the PDGF-BB-induced MAP-kinase activity (4.6-fold). Transcription of growth response genes such as c-fos, c-jun, and c-myc were induced by PDGF-BB, and their induction was suppressed, particularly c-myc, by incubating SMCs with PDGF-BB plus SB. Similarly, preincubation of cells with SB for 30 minutes and subsequent incubation in PDGF-BB plus SB diminished PDGF-BB-induced transcription of c-fos, c-jun, and c-myc. However, SB by itself had no significant effect on c-fos, c-jun, and c-myc transcription.(ABSTRACT TRUNCATED AT 400 WORDS) PMID: 7489253 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 547: Nature. 1995 Nov 30;378(6556):509-12. Comment in: Nature. 1995 Nov 30;378(6556):438-9. Myc but not Fos rescue of PDGF signalling block caused by kinase-inactive Src. Barone MV, Courtneidge SA. Differentiation Programme, European Molecular Biology Laboratory, Heidelberg, Germany. Growth factors such as platelet-derived growth factor (PDGF) elicit the transcriptional activation of a large number of immediate early genes (many of which encode transcription factors), and ultimately DNA synthesis. Both AP1 and Myc are activated in fibroblasts in response to growth factor stimulation, and various experiments suggest their importance in proliferation. Src family kinases are required for PDGF (and other growth factors) to induce DNA synthesis. We have examined which transcription factors, when constitutively expressed, 'rescue' the block elicited by dominant negative Src. We report here that Myc, but not Fos and/or Jun, was able to rescue the block. In contrast, Fos and Jun, but not Myc, rescued the block induced by dominant negative Ras. Our data suggest that Src kinases control the transcriptional activation of Myc. PMID: 7477410 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 548: J Biol Chem. 1995 Nov 24;270(47):28337-41. Gastrin and glycine-extended progastrin processing intermediates induce different programs of early gene activation. Todisco A, Takeuchi Y, Seva C, Dickinson CJ, Yamada T. Department of Internal Medicine, University of Michigan Medical Center, Ann Arbor 48109-0368, USA. We recently reported that gastrin and glycine-extended progastrin processing intermediates (G-Gly) exert growth-promoting effects on AR4-2J cells (derived from rat pancreas) via interaction with distinct receptors. In this study we sought to investigate the mechanisms by which gastrin and G-Gly stimulate cell proliferation. While gastrin increased [Ca2+]i in AR4-2J cells, G-Gly had no effect. Similarly, G-Gly had no effect either on basal and 10(-7) M vasoactive intestinal polypeptide-stimulated cAMP generation, although gastrin is known to inhibit cAMP generation. Gastrin dose dependently stimulated AR4-2J cell mRNA content of both c-fos and c-jun, two genes known to function in regulating cell proliferation, but G-Gly had no effect. Gastrin also induced the expression of luciferase in AR4-2J cells transfected with a construct consisting of a luciferase reporter gene coupled to the serum response element of the c-fos gene promoter. In similar fashion, gastrin stimulated the activity of mitogen-activated protein kinase, an enzyme known to mediate the induction of the c-fos serum response element in response to growth factor stimulation. Although G-Gly had none of these effects of gastrin in AR4-2J cells, it stimulated activity of c-Jun amino-terminal kinase, an enzyme known to phosphorylate and transcriptionally activate c-Jun. These data support the notion that gastrin stimulates cell proliferation by inducing c-fos and c-jun gene expression, while G-Gly acts by post-translationally regulating early gene transcriptional activation. Our studies represent a novel model in which both the precursor and the product of a key processing reaction, peptide alpha-amidation, act cooperatively to stimulate cell proliferation via distinct receptors linked to different signal transduction pathways. PMID: 7499334 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 549: Mol Cell Biochem. 1995 Nov 22;152(2):131-41. Molecular remodelling in hypertrophied hearts from polyomavirus large T-antigen transgenic mice. Holder EL, Al Moustafa AE, Chalifour LE. Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, Montreal, Quebe. Polyomavirus large T-antigen transgenic mice develop cardiac hypertrophy characterized by an increase in atrial natriuretic factor and beta-myosin heavy chain isoform expression. The aim of this study was to examine changes in proto-oncogene expression in hypertrophied hearts from the transgenic mice. Expression of early growth response-1 (Egr-1) mRNA was detected in hearts from all 15 transgenic mice, but was not detectable in 13 control mice. Reverse transcriptase-polymerase chain reaction experiments using Egr-1-specific primers confirmed the increase in Egr-1 mRNA in enlarged hearts from the transgenic mice. Expression of c-jun, junD and Ha-ras mRNAs was increased in the transgenic hearts 3, 17 and 2.8-fold respectively. Western blots showed an increase in c-myc, c-jun and ras protein in hypertrophied transgenic hearts. Immunofluorescence analyses confirmed an increase in Egr-1 and c-jun protein in transgenic cardiomyocytes. Proliferating cell nuclear antigen, Ki-ras and HSP 90 mRNAs were decreased 22, 2.7 and 3-fold, respectively in the transgenic hearts. Not altered in most hypertrophied hearts was expression of c-fos, junB, p53, c-neu, c-myc, HSP70, HSP27, TGF-beta or IGF 1 mRNAs. Proto-oncogene and growth factor gene expression in hypertrophy induced by PVLT expression is modulated with some proto-oncogenes increased and others decreased in expression. PMID: 8751159 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 550: Cancer Lett. 1995 Nov 6;97(2):169-75. Modulation of expression of genes encoding nuclear proteins following exposure to JANUS neutrons or gamma-rays. Woloschak GE, Chang-Liu CM. Center for Mechanistic Biology and Biotechnology, Argonne National Laboratory, IL 60439-4833, USA. Previous work has shown that exposure of cells to ionizing radiations causes modulation of a variety of genes, including those encoding c-fos, interleukin-1, tumor necrosis factor, cytoskeletal elements, and many more. The experiments reported herein were designed to examine the effects of either JANUS neutron or gamma-ray exposure on expression of genes encoding nucleus-associated proteins (H4-histone, c-jun, c-myc, Rb, and p53). Cycling Syrian hamster embryo cells were irradiated with varying doses and dose rates of either JANUS fission-spectrum neutrons or gamma-rays; after incubation of the cell cultures for 1 h following radiation exposure, mRNA was harvested and analyzed by Northern blot. Results revealed induction of transcripts for c-jun, H4-histone, and (to a lesser extent) Rb following gamma-ray but not following neutron exposure. Interestingly, expression of c-myc was repressed following gamma-ray but not following neutron exposure. Radiations at different doses and dose rates were compared for each of the genes studied. PMID: 7497459 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 551: J Neuroendocrinol. 1995 Nov;7(11):875-9. Expression of early genes in estrogen induced phenotypic conversion of neuroblastoma cells. Santagati S, Ma ZQ, Ferrarini C, Pollio G, Maggi A. Milano Molecular Pharmacology Laboratory, University of Milan, Italy. Estrogens are known to modulate the growth rate and differentiation state of a number of cells. In uterine, as well as in mammary tumor cells, estrogen-dependent proliferation and differentiation are correlated to a series of biochemical responses, including increased expression of proto-oncogenes such as: c-fos, c-jun and c-myc. Since estrogens were shown to regulate the proliferation and the differentiation state of cells of nervous origin, the aim of the present study was to investigate whether these effects were associated to changes in the expression of early genes. In the model system utilized, the human cell line SK-ER3, an increase in c-fos mRNA and Fos protein without change of c-jun and related genes mRNA concentration was observed after short term treatment with 17 beta-estradiol (E2). A significant decrease of c-fos, c-jun and jun-D proto-oncogene mRNA levels were found after prolonged hormonal treatment. The exposure to the hormone did not determine any change in N-myc expression. Since the three protooncogene mRNAs are rapidly induced following estrogen treatment in other cell systems and target tissues, it is concluded that the estrogen-induced differentiation of neuroblastoma cells is correlated to a pattern of expression of early genes that might be peculiar for the activity of this hormone in neural cells. PMID: 8748125 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 552: Eur J Cancer B Oral Oncol. 1995 Nov;31B(6):384-91. Differential c-myc, c-jun, c-raf and p53 expression in squamous cell carcinoma of the head and neck: implication in drug and radioresistance. Riva C, Lavieille JP, Reyt E, Brambilla E, Lunardi J, Brambilla C. Lung and Airways Cancer Research Group, Albert Bonniot Institute, La Tronche, France. The expression of oncogenes c-myc, c-jun and c-raf and tumour suppressor gene p53 was assessed by northern blot analysis of 42 tumours and p53 protein expression by immunohistochemistry on paraffin-embedded sections from 36 specimens of squamous cell carcinoma of the head and neck (SCCHN) obtained before therapy. Of the 42 tumours, 89, 100 and 100% expressed c-myc, c-jun and c-raf oncogenes, respectively. These oncogene expressions did not correlate with sex, age or clinical stage of the disease. However, an association was found between low c-myc expression (P = 0.0001) and high c-jun expression (P = 0.0001) and absence of tumoral response to neoadjuvant chemotherapy. On the other hand, c-raf overexpression was observed in patients resistant to radiation therapy (P = 0.0494). Forty-two per cent of the tumours showed p53 protein overexpression, which did not correlate with any clinical parameter. This p53 protein overexpression was associated with high p53 mRNA levels (REL) (P = 0.0223). A correlation was found between increased c-myc RNA expression and lack of p53 protein expression (P = 0.0407). In addition, a lack of p53 protein expression was indicative of tumour relapse (P = 0.05). None of these biological parameters were associated with disease-free survival (Cox-Mantel test). In conclusion, the overexpression of c-myc, c-jun and c-raf may be independently associated to tumoral response to chemotherapy or radiotherapy, or to tumour relapse, but fail to predict long-term survival. PMID: 8746269 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 553: Biotechniques. 1995 Nov;19(5):706-8, 710. Validation of northern blot analysis for quantitating the expression of several genes in rat liver. Triest S, Maiter D, Ketelslegers JM. University of Louvain School of Medicine, Brussels, Belgium. PMID: 8588900 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 554: Bone. 1995 Nov;17(5):479-84. In vivo, human parathyroid hormone fragment (hPTH 1-34) transiently stimulates immediate early response gene expression, but not proliferation, in trabecular bone cells of young rats. Onyia JE, Bidwell J, Herring J, Hulman J, Hock JM. Endocrine Division, Lilly Research Labs, Indianapolis, IN 46285, USA. Intermittent PTH increases trabecular bone mass in vivo by stimulating osteoblast differentiation to increase bone formation. The molecular events that mediate the anabolic effect of PTH on osteoblasts have not been characterized. We investigated if PTH regulated mRNA expression of proto-oncogenes, c-fos, c-jun, and c-myc, early response genes that have been shown to be involved in the regulation of both cell proliferation and differentiation. As PTH also regulates the early expression of the cytokine, interleukin-6 (IL-6), in bone cells in vitro, we also investigated if this occurred in vivo, in concert with the other early response genes. Northern blot hybridization was used to analyze mRNA expression in the metaphysis of the distal femur of young rats. To determine the proliferative state in these femurs, mRNA expression of the cell proliferation marker histone, H4, was assessed. Subcutaneous administration of a single injection of human PTH (1-34) at 8 micrograms/100 g, a dose known to increase bone forming surfaces, induced rapid and transient expression of c-fos, c-jun, c-myc, and IL-6 mRNA. A second novel transcript for IL-6 was detected, but its significance remains unknown. Induction of all these messages was evident by 1 h; the levels of mRNA returned to baseline after 3-6 h. Concurrently, PTH had a small inhibitory effect on the expression of histone H4 mRNA. We conclude that, in vivo, PTH upregulates cell differentiation in trabecular bone by transient stimulation of the early response genes and IL-6, while downregulating cell proliferation. PMID: 8579960 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 555: Differentiation. 1995 Nov;59(4):243-52. Inhibitors of ADP-ribosylation suppress phenotypic modulation and proliferation of smooth muscle cells cultured from rat aorta. Thyberg J, Hultgardh-Nilsson A, Kallin B. Department of Cell and Molecular Biology, Karolinska Institute, Stockholm, Sweden. The effects of hexamethylenebisacetamide (HMBA), an inhibitor of poly-ADP-ribosylation, and meta-iodobenzylguanidine (MIBG), an inhibitor of mono-ADP-ribosylation, on the phenotypic properties and proliferation of cultured rat aortic smooth muscle cells were studied using a combination of structural and chemical methods. The results show that HMBA and MIBG both slowed down the transition of the cells from a contractile to a synthetic phenotype in primary culture. While the control cells rapidly lost most of their myofilaments and built up an extensive endoplasmic reticulum and Golgi complex, a conspicuous fraction of the drug-treated cells retained a characteristic smooth muscle morphology for at least 6 days. Moreover, most of the treated cells remained positive for smooth muscle alpha-actin and desmin throughout this period. In contrast, the drugs lacked distinct effects on cell morphology and cytoskeletal organization in secondary cultures. Nevertheless, they strongly inhibited serum-stimulated cell growth both in primary and secondary cultures. The ability of serum-starved cells to synthesize DNA after exposure to platelet-derived growth factor or serum was also restrained. Notably, the drugs could be added several hours after the mitogens without loss of effect, suggesting that they did not prevent the entrance into but rather the progression through the cell cycle. Accordingly, the expression of early response genes like c-fos, c-jun and c-myc was not blocked by the drugs. On the other hand, HMBA reduced the expression of transcripts for smooth muscle alpha-actin, type IV collagenase, collagen type I, and osteopontin both in primary and secondary cultures. Weaker and more variable effects were obtained with MIBG. Taken together, the findings support the notion that poly- and mono-ADP-ribosylation of proteins are involved in the control of smooth muscle cell differentiation and growth. PMID: 8575646 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 556: Cancer Immunol Immunother. 1995 Nov;41(5):280-6. A flow cytometric study of c-erbB-3 expression in breast cancer. Brotherick I, Shenton BK, Angus B, Waite IS, Horne CH, Lennard TW. Department of Surgery, Medical School, University of Newcastle upon Tyne, England. In order to assess the specificity of biotinylated anti-c-erbB-3 antibody, screening was performed on a series of tumour cell lines and lymphocytes. Staining was found to be consistent, with good reproducibility. Twenty-nine consecutive breast cancer samples were obtained from women treated with tamoxifen and undergoing elective mastectomy. Twenty-eight invasive ductal carcinomas and 1 DCIS were stained for c-erbB-3 expression: 2 were grade I (Bloom and Richardson), 15 grade II, and 11 grade III tumours, 1 being unclassified; 16 were axillary node positive and 10 node negative; in 2 cases no nodes were sampled. Tumours examined by flow cytometry were stained with cytokeratin FITC antibody and the cytokeratin-positive population gated. Using Mann-Whitney analysis no association was seen between c-erbB-3 expression and Bloom and Richardson grade or axillary node status. In the tumour samples c-erbB-3 expression was found to show as association with EGF-R (P = 0.021 r2 = 0.16), PgR (P = 0.02, r2 = 0.16), c-myc (P < 0.0001, r2 = 0.5), c-jun (P = 0.001, r2 = 0.4) and c-fos (P = 0.001, r2 = 0.5) but not with c-erbB-2 (P = 0.2, r2 = 0.06), ER (P = 0.4, r2 = 0.02) or p53 1801 (P = 0.05, r2 = 0.2). Expression of c-erbB-3 may not be an independent marker of prognosis, but it is associated with other markers of poor prognosis and early cellular events linked with aberrant growth and differentiation. PMID: 8536273 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 557: Proc Soc Exp Biol Med. 1995 Nov;210(2):162-70. Transforming growth factor-alpha and epidermal growth factor activate mitogen-activated protein kinase and its substrates in intestinal epithelial cells. Oliver BL, Sha'afi RI, Hajjar JJ. Department of Physiology, University of Connecticut Health Center, Farmington 06030, USA. The signal transduction pathways of mitogenic stimuli in intestinal epithelial cells are not clearly understood. We report here a possible signaling pathway of two closely related agonists, transforming growth factor-alpha (TGF alpha) and epidermal growth factor (EGF). Both increase thymidine incorporation in the intestinal epithelial cell (IEC) line IEC-6. This increase is dose dependent and inhibited by the tyrosine kinase inhibitors genistein and tyrphostin. The addition of either TGF alpha or EGF to IEC-6 cells also stimulates the activities of the two forms of mitogen-activated protein kinase, p42erk2 MAPK and p44erk1 MAPK, as evidenced by increased incorporation of radiolabeled phosphate in myelin basic protein. The main difference between the MAPK activity levels induced by the two agonists is in the intensity of the response. Maximum TGF alpha-induced stimulation of p42erk2 MAPK activity is 9-fold at 2 ng/ml, while maximum EGF stimulation is only 4.5-fold at 25 ng/ml. These doses correlated closely with the dose required for maximum thymidine incorporation. The activity of the 90-kDa ribosomal S6 kinase, a downstream substrate for activated MAPK, is also enhanced as evidenced by increased incorporation of radiolabeled phosphate in the rsk kinase substrate peptide in IEC-6 cells following stimulation with either TGF alpha or EGF. This increase correlates closely with the stimulus-induced increase in MAPK activity with respect to dose, but the time of increased activity is more prolonged, especially after EGF stimulation. TGF alpha induced the synthesis of both c-Fos and c-Myc, two nuclear substrates for MAPK, and increased c-fos and c-myc message levels as well. However, c-Jun protein and c-jun mRNA were not induced. The increase in IEC-6 cell proliferation in response to TGF alpha and EGF stimulation may then be due, in part, to an increase in immediate early gene expression as a direct result of MAPK and RSK activation. PMID: 7568287 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 558: Biochemistry. 1995 Oct 17;34(41):13554-64. The leucine zippers of the HLH-LZ proteins Max and c-Myc preferentially form heterodimers. Muhle-Goll C, Nilges M, Pastore A. EMBL, Heidelberg, Germany. c-Myc and Max are members of a subfamily of the helix-loop-helix transcription-regulating proteins. Their function is mediated by switches in the dimerization partners; c-Myc does not homodimerize in vivo but competes with Mad, another member of the subfamily, to form heterodimers with Max, leading to either activation or repression of transcription. Max is also able to form homodimers. In an attempt to identify which regions of the proteins carry the information to determine specific recognition of the dimerization partner, we have investigated the dimerization properties of synthetic peptides corresponding to the leucine zipper sequence of Max and c-Myc using circular dichroism and nuclear magnetic resonance techniques. We show that the heterodimer is obtained readily by simply mixing the peptides and that at neutral pH it is more stable than the homodimer of the Max leucine zipper. We have shown in a previous paper [Muhle-Goll, C. et al. (1994) Biochemistry 33, 11296-11306] that the leucine zipper of c-Myc does not form stable homodimers under these conditions. Thus, the leucine zipper regions of these two proteins by themselves display the same behavior as the entire proteins. However, even the heterodimer is less stable than dimers of leucine zippers of the basic leucine zipper family such as GCN4 and Fos-Jun. The specificity of the interaction between different monomers can be explained by polar interactions. We investigate the structural role of the polar and charged residues in the hydrophobic interface by molecular-modeling studies. PMID: 7577944 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 559: J Physiol. 1995 Oct 15;488 ( Pt 2):493-9. Expression of immediate early genes in rat gastric myenteric neurones: a physiological response to feeding. Dimaline R, Miller SM, Evans D, Noble PJ, Brown P, Poat JA. Physiological Laboratory, University of Liverpool, UK. 1. Expression of the immediate early genes c-fos, c-jun and c-myc in rat stomach in response to feeding and gastric distension was examined by Northern blot analysis and in situ hybridization. 2. Refeeding of fasted rats induced a transient increase in c-fos mRNA abundance in gastric corpus and antrum that was sixfold within 15 min and declined within 4 h. The response was not mediated by gastrinergic or muscarinic cholinergic mechanisms; it was reduced but not abolished by hexamethonium. No changes in expression of c-jun, c-myc or the constitutively expressed protein glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were observed. 3. In conscious rats prepared with a gastric fistula, gastric distension with nutritive and non-nutritive solutions at a physiological pressure for 30 min induced expression of c-fos, c-jun and c-myc, but not GAPDH. 4. Messenger RNA encoding c-fos was localized by in situ hybridization to gastric myenteric neurones of animals that underwent gastric distension, but not of undistended controls. 5. The results suggest that expression of c-fos in gastric myenteric neurones is an early response to the physiological stretching of the stomach wall that accompanies feeding. With supraphysiological distension, other immediate early genes may be recruited. PMID: 8568687 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 560: Oncogene. 1995 Oct 5;11(7):1403-7. Changes in expression of members of the fos and jun families and myc network during terminal differentiation of human keratinocytes. Gandarillas A, Watt FM. Keratinocyte Laboratory, Imperial Cancer Research Fund, Lincoln's Inn Fields, London, UK. Normal human epidermal keratinocytes are induced to undergo terminal differentiation when disaggregated and placed in suspension: commitment occurs at about 5 h and overt differentiation by 24 h. In contrast, the squamous cell carcinoma line SCC12B2 does not initiate terminal differentiation until 75 h in suspension and the ndk strain of keratinocytes undergoes growth arrest but does not differentiate at all. In order to identify genes that may regulate terminal differentiation we have examined mRNA levels of members of the fos and jun gene families and myc gene network in suspension cultures of normal keratinocytes, SCC12B2 and ndk. The major changes observed were an up-regulation of c-Fos and Fra-1 and a decrease in c-Myc at the time of commitment, followed by an increase in Mad, Mxi1, Fra-2 and JunB expression at the onset of differentiation. The sequence of events shows a striking similarity to that which occurs during myeloid differentiation and suggests a role for AP-1 complexes and Myc and Mad complexes in regulating keratinocyte differentiation. PMID: 7478564 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 561: Tanpakushitsu Kakusan Koso. 1995 Oct;40(14):2126-33. [Targeted ablation of transcription factor genes] [Article in Japanese] Kondoh H. Institute for Molecular and Cellular Biology, Osaka University, Japan. Publication Types: Review Review, Tutorial PMID: 8532868 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 562: Carcinogenesis. 1995 Oct;16(10):2423-8. Altered expression of cyclins and c-fos in R6 cells that overproduce PKC epsilon. Han EK, Cacace AM, Sgambato A, Weinstein IB. Columbia-Presbyterian Cancer Center, NY 10032, USA. Since PKC epsilon functions as an oncogene when stably overexpressed in R6 rat fibroblasts (Cacace et al. 1993) in the present study we examined whether transformed R6-PKC epsilon cells display abnormalities in the expression of specific early response and cyclin genes. When vector control and R6-PKC epsilon cells were starved of serum for 72 h they arrested in G0/G1 and showed passage through the cell cycle at similar rates after subsequent stimulation with 10% fetal calf serum plus TPA. In PKC epsilon cells, induction of cyclin D1 protein was markedly reduced, and that of cyclin A was slightly reduced when compared to control cells. Northern blot analyses indicated that decreased expression of cyclin D1 and A protein in PKC epsilon cells is due to translational or post-translational effects. A study of early response gene expression in PKC epsilon cells indicated that there was a marked reduction in the expression of c-fos mRNA but not in c-jun or c-myc mRNAs. The marked decreases in cyclin D1 and c-fos expression seen in PKC epsilon cells were not seen in R6 cells that overexpress PKCs alpha or beta. These findings suggest that PKC epsilon cells bypass certain normal signal transduction and cyclin-controlled pathways involved in cell proliferation. PMID: 7586146 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 563: Clin Exp Immunol. 1995 Oct;102(1):6-10. Defective expression of early activation genes in cartilage-hair hypoplasia (CHH) with severe combined immunodeficiency (SCID). Castigli E, Irani AM, Geha RS, Chatila T. Children's Hospital, Department of Pediatrics, Harvard Medical School, Boston, MA, USA. Cartilage-hair hypoplasia (CHH) is an autosomal recessive disease of unknown etiology characterized by metaphyseal dysostosis, unpigmented hair, and defective cellular immunity. We studied peripheral blood mononuclear cells (PBMC) of a boy with CHH and combined immunodeficiency in an attempt to characterize further the immune defect in this disease. Stimulation of his PBMC with mitogens was associated with severely depressed IL-2 and interferon-gamma (IFN-gamma) synthesis and IL-2 receptor alpha-chain (IL-2R alpha) expression and resulted in poor lymphocyte proliferation that was only modestly upregulated by the addition of recombinant IL-2 (rIL-2). The defective proliferation and lymphokine synthesis were not corrected by the addition of phorbol myristate acetate (PMA) and ionomycin, agents that bypass receptor-mediated signalling, indicative of a distal abnormality. Importantly, the levels of mRNA encoding c-myc, IL-2R alpha, IL-2 and IFN-gamma were markedly decreased in patient lymphocytes stimulated with PMA+ionomycin as compared to control lymphocytes. The defect in the expression of these early activation genes was selective in that induction by mitogens of mRNA encoding other early activation gene products such as c-fos and c-jun was not impaired. These results suggest that the underlying defect in this patient and perhaps others with CHH may be an abnormality in a component of intracellular signalling pathways or in a trans-acting factor which regulates the expression of a selected number of early activation genes. PMID: 7554401 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 564: Br J Cancer. 1995 Oct;72(4):922-7. Effects of transforming growth factor beta-1 on growth-regulatory genes in tumour-derived human oral keratinocytes. Paterson IC, Patel V, Sandy JR, Prime SS, Yeudall WA. Department of Oral and Dental Science, University of Bristol Dental Hospital and School, UK. This study examined the effect of transforming growth factor beta-1 (TGF-beta 1) on c-myc, RB1, junB and p53 expression together with pRb phosphorylation, in carcinoma-derived and normal human oral keratinocytes with a range of inhibitory responses to this ligand. Amplification of c-myc was observed in eight of eight tumour-derived cell lines and resulted in corresponding mRNA expression. The down-regulation of c-myc expression by TGF-beta 1 predominantly reflected growth inhibition by TGF-beta 1, but in two of eight tumour-derived cell lines which were partially responsive to TGF-beta 1 c-myc expression was unaltered by this ligand. While RB1 mRNA levels were unaltered by TGF-beta 1, the ligand caused the accumulation of the underphosphorylated form of the Rb protein in all cells irrespective of TGF-beta 1-induced growth arrest. junB expression was up-regulated by TGF-beta 1 in cells with a range of growth inhibitory responses. All cells contained mutant p53. TGF-beta 1 did not affect p53 mRNA expression in both tumour-derived and normal keratinocytes and there was no alteration in p53 protein levels in keratinocytes expressing stable p53 protein following TGF-beta 1 treatment. The data indicate that TGF-beta-induced growth control can exist independently of the presence of mutant p53 and the control of Rb phosphorylation and c-myc down-regulation. It may be that TGF-beta growth inhibition occurs via multiple mechanisms and that the loss of one pathway during tumour progression does not necessarily result in the abrogation of TGF-beta-induced growth control. PMID: 7547241 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 565: Biochem Biophys Res Commun. 1995 Sep 25;214(3):941-8. Involvement of CD45 in dexamethasone- and heat shock-induced apoptosis of rat thymocytes. Troiano L, Monti D, Cossarizza A, Lovato E, Tropea F, Barbieri D, Morale MC, Gallo F, Marchetti B, Franceschi C. Department of Biomedical Sciences, University of Modena, Italy. CD45 is a transmembrane tyrosine-specific phosphatase which participates in lymphoid cell signal transduction during T cell activation, as well as in intrathymic negative and positive selection. In mammals, this molecule exhibits a variety of isoforms of different molecular weight, whose roles have still to be fully elucidated. We report here that apoptosis of rat thymocytes after in vitro dexamethasone and heat shock treatment was accompanied by an early significative increase of cells expressing CD45RC, the high molecular weight isoform of CD45 molecule. The same phenomenon was observed in thymocytes derived from in vivo dexamethasone-treated rats. However, the increase of CD45RC+ cells was not apparently characteristic of cells undergoing apoptosis, as the same phenomenon was also observed in rat thymocytes induced to proliferate by Concanavalin A. On the whole, these results suggest that CD45 modulation can be added to the list of early molecular events, such as the increased expression of genes (ornithine decarboxylase), proto-oncogenes (c-fos, c-jun, c-myc) and activation of transcription factors (AP-1, NFkB), we previously demonstrated in the same experimental model to occur and to be shared by these two apparently opposite biological processes, i.e., cell proliferation and apoptosis, both likely depending on a complex balance of protein phosphorylation and dephosphorylation. PMID: 7575567 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 566: Gene. 1995 Sep 11;162(2):285-9. Interleukin-6 and ciliary neurotrophic factor trigger janus kinase activation and early gene response in rat hepatocytes. Wang Y, Fuller GM. Department of Cell Biology, University of Alabama at Birmingham 35294, USA. Interleukin-6 (IL-6) and ciliary neurotrophic factor (CNTF) share a signal-transducing molecule called gp130. Previous studies showed that CNTF regulates fibrinogen gene expression in rat hepatocytes by competitive binding to the IL-6 receptor. This report explores the post ligand-binding events in the control of fibrinogen and early response gene production stimulated by IL-6 and CNTF in primary rat hepatocytes. Metabolic labeling, using [32P]orthophosphate or anti-phosphotyrosine antibody (Ab) blot experiments revealed that both IL-6 and CNTF induced tyrosine phosphorylation of gp130, and the Jak1 and Jak2 kinases in a dose- and time-dependent manner. Additional experiments revealed that only one of the early response genes, junb, but not c-myc or c-fos, was stimulated by the addition of either IL-6 or CNTF. These data suggest that activation of Jak kinases and stimulation of junb reflect a divergence of the IL-6/CNTF signaling pathway and further suggest that junb may participate in cytokine control of acute-phase protein production in the inflammatory response. PMID: 7557445 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 567: Mol Biol (Mosk). 1995 Sep-Oct;29(5):1137-44. [Concentration of proto-oncogenes in hepatocyte nuclei] [Article in Russian] Boikov PIa, Kostiuk GV, Terent'ev AA, Shevchenko NA. PMID: 8538608 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 568: Am J Pathol. 1995 Sep;147(3):699-706. Effect of vitamin A deficiency on the integrity of hepatocytes after partial hepatectomy. Evarts RP, Hu Z, Omori N, Omori M, Marsden ER, Thorgeirsson SS. Laboratory of Experimental Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4255, USA. The effect of vitamin A deficiency on hepatic regeneration in male and female rats was studied after partial hepatectomy. A fourfold increase in the number of positive dUTP end-labeled nuclei was observed in the deficient animals as early as 30 minutes after partial hepatectomy and their number reached a peak by 8 hours after the operation. The bile duct cells were both morphologically and biochemically intact at all time points. Administration of retinyl palmitate 1 hour before partial hepatectomy significantly reduced the number of positive nuclei, and treatment with retinyl palmitate 24 or 48 hours before the operation reduced the number of positive cells to the level observed in control vitamin A-supplemented rats. The level of transcripts for c-jun, c-fos, c-myc, and transforming growth factor-beta 1 were increased for an extended period of time in livers of deficient animals, whereas the expression of both p53 and max were unchanged. Immunocytochemistry demonstrated the presence of latent transforming growth factor-beta 1 in cells showing evident apoptotic or necrotic changes in their nuclei. This study demonstrates the importance of vitamin A for the survival of hepatocytes both in intact vitamin A-deficient liver and after partial hepatectomy, whereas the ductal cells appear to be less sensitive to vitamin A deficiency. PMID: 7677181 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 569: Radiat Res. 1995 Sep;143(3):263-72. Induction of transcription of "immediate early genes" by low-dose ionizing radiation. Prasad AV, Mohan N, Chandrasekar B, Meltz ML. Department of Radiology, University of Texas Health Science Center, San Antonio 78284-7800, USA. The induction of transcription of specific genes after exposure to ionizing radiation has previously been reported after lethal doses of radiation (2-50 Gy). Little attention has been focused on expression of "immediate early genes" after low doses of ionizing radiation, where cell viability remains high. This dose range (0.25-2.0 Gy) is above the diagnostic dose level but at or below the doses typical for a single exposure in fractionated radiotherapy treatment of cancer. In this study, it was observed that doses in the range of 0.25-2.0 Gy induced different amounts of the mRNAs of the proto-oncogenes c-fos, c-jun, c-myc and c-Ha-ras at a given dose and time in Epstein-Barr virus-transformed human lymphoblastoid 244B cells. A maximum response was seen after a dose of 0.5 Gy for all but c-fos, which showed a maximum response after exposure to 0.25 Gy. Time-course studies demonstrated that, for all four proto-oncogenes, the induction was transient, reaching a maximum at 1 h and declining to the constitutive level at 4 h after irradiation. Using second-messenger specific inhibitors, the signaling pathways involved in the induction of these proto-oncogenes was also investigated. The results showed that all four of the proto-oncogenes induced after 0.5 Gy shared a common pathway of tyrosine kinase activation. Other signaling pathways included protein kinase C, reactive oxygen intermediates and calcium-dependent kinases; these were found to be differentially involved in the induction of transcription of the individual proto-oncogenes. In summary, this study suggests that low-dose ionizing radiation (0.25-2.0 Gy) can modulate expression of immediate early genes. Secondly, the activation of immediate early genes after low-dose exposure involves multiple second-messenger signaling pathways. Third, the magnitude of involvement of the different signaling pathways after low-dose radiation is different for each proto-oncogene expressed. PMID: 7652163 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 570: Br J Rheumatol. 1995 Sep;34(9):805-8. Oncogene expression in synovial fluid cells in reactive and early rheumatoid arthritis: a brief report. Roivainen A, Isomaki P, Nikkari S, Saario R, Vuori K, Toivanen P. Department of Medical Microbiology, Turku University, Finland. Since it has been implied that cellular oncogenes might have a role in the pathogenesis of rheumatoid arthritis (RA), we have examined the expression of c-myc, c-myb, c-fos, c-jun and c-Ha-ras oncogenes in the cells from synovial fluid (SF) and peripheral blood (PB) of patients with reactive arthritis (ReA) and early RA. Oncogene expression was studied using RNA hybridizations with 32P-labelled probes. From the SF, mononuclear and granulocyte cell fractions were used separately. Significant differences between ReA and RA were observed only for c-myb in PB mononuclear cells and c-jun in SF granulocytes. Regarding the expression of c-myc, c-fos and c-Ha-ras oncogenes, no difference between ReA and RA was observed. Comparison to normal controls was made using PB mononuclear cells; only the expression of c-fos tended to be slightly increased in RA, without statistical significance, however. We conclude that oncogene activation in the synovial inflammation is not a phenomenon specific for RA. PMID: 7582717 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 571: Leuk Res. 1995 Sep;19(9):645-50. Augmentation by aphidicolin of 1-beta-D-arabinofuranosylcytosine-induced c-jun and NF-kappa B activation in a human myeloid leukemia cell line: correlation with apoptosis. Kuwakado K, Kubota M, Bessho R, Kataoka A, Usami I, Lin YW, Okuda A, Wakazono Y. Department of Pediatrics, Faculty of Medicine, Kyoto University, Japan. 1-beta-D-arabinofuranosylcytosine (ara-C) (2 microM) can induce apoptosis in a human myeloid leukemia cell line, U937, after 4 h of incubation. Pretreatment of cells with aphidicolin (2 microM) augments ara-C-induced apoptosis, since it was first observed at 0.4 microM ara-C and became more intense at 2 and 10 microM. Although aphidicolin itself had a marginal effect on c-jun expression, it significantly augmented ara-C induced c-jun upregulation by shortening the lag time and lowering ara-C concentrations necessary for the induction of detectable c-jun transcripts. Aphidicolin and ara-C acted synergistically to increase NF-kappa B DNA binding activity as determined by an electrophoretic mobility shift assay. Expression of c-myc was slightly increased through the DNA degradative phase, and was then downregulated. Thus, the activation of NF-kappa B and c-jun expression seems to be well correlated with the potentiation by aphidicolin of ara-C-induced apoptosis. PMID: 7564475 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 572: Proc Natl Acad Sci U S A. 1995 Aug 29;92(18):8458-62. Asbestos induces nuclear factor kappa B (NF-kappa B) DNA-binding activity and NF-kappa B-dependent gene expression in tracheal epithelial cells. Janssen YM, Barchowsky A, Treadwell M, Driscoll KE, Mossman BT. Department of Pathology, University of Vermont College of Medicine. Nuclear factor kappa B (NF-kappa B) is a transcription factor regulating expression of genes intrinsic to inflammation and cell proliferation--features of asbestos-associated diseases. In studies here, crocidolite asbestos caused protracted and dose-responsive increases in proteins binding to nuclear NF-kappa B-binding DNA elements in hamster tracheal epithelial (HTE) cells. This binding was modulated by cellular glutathione levels. Antibodies recognizing p65 and p50 protein members of the NF-kappa B family revealed these proteins in two of the DNA complexes. Transient transfection assays with a construct containing six NF-kappa B-binding DNA consensus sites linked to a luciferase reporter gene indicated that asbestos induced transcriptional activation of NF-kappa B-dependent genes, an observation that was confirmed by northern blot analyses for c-myc mRNA levels in HTE cells. Studies suggest that NF-kappa B induction by asbestos is a key event in regulation of multiple genes involved in the pathogenesis of asbestos-related lung cancers. PMID: 7667311 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 573: Biochem Biophys Res Commun. 1995 Aug 24;213(3):789-95. Coexpression pattern of c-myc associated genes in a small cell lung cancer cell line with high steady state c-myc transcription. Dooley S, Wundrack I, Blin N, Welter C. Department of Human Genetics, Univ of Saarland, Homburg, Germany. The c-Myc protein is involved in cellular transformation and mitogenesis, but also works as a potent inducer of differentiation and programmed cell death. Max as an obligate heterodimeric partner for Myc mediates its functions as a specific transcriptional activator and a transforming protein. Mad and Mxi1 proteins both heterodimerize with Max and compete with each other for limiting amounts of Max. Transcriptional activation by Myc can be suppressed by increasing the amount of Mad or Mxi1. This report shows the expression pattern of these Myc related factors at the mRNA level in a small cell lung cancer (SCLC) cell line (GLC4) which is characterized by c-myc amplification and strong constitutive c-myc overexpression. We found these genes transcriptionally active but uninfluenced from high c-myc transcription. Max was constantly transcribed at a relatively low level during cell cycle progression. Mad and mxi1 mRNA was at a surprisingly high level in proliferating cells. Mad was further upregulated and mxi1 was downregulated to basal levels during serum starvation of the cells. We further analyzed the activity of c-fos, c-jun, c-myb and nm23 which are described to be involved in c-myc transcriptional activation, c-jun and c-fos were not constitutively activated and can be excluded as regulators. High steady state c-myc in contrast influences the serum stimulated transient activation mechanism of these two genes. We identified high copy number nm23 mRNA whose role as a putative c-myc transcriptional activator is under investigation. Our results indicate that constitutive overexpression of c-myc does not require the activity of the nuclear oncogenes tested and that the m-RNA expression pattern of functionally related proteins is not influenced. PMID: 7654239 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 574: Cell Growth Differ. 1995 Aug;6(8):945-53. Insulin regulates expression of c-fos and c-jun and suppresses apoptosis of lens epithelial cells. Rampalli AM, Zelenka PS. Laboratory of Molecular and Developmental Biology, National Eye Institute, NIH, Bethesda, Maryland 20892, USA. This study investigates whether insulin (a differentiation factor for lens epithelial cells) acts as a survival factor. In the absence of insulin, 6-day embryonic chicken lens epithelial explants undergo apoptosis as shown by changes in cell morphology, DNA fragmentation, and loss of trypan blue exclusion. Insulin inhibits these changes and promotes survival of the cells. Aurintricarboxylic acid suppresses the apoptosis of lens explants. In contrast to 6-day embryonic explants, 19-day embryonic explants survive in the absence of insulin, presumably due to an endogenous survival factor. To explore the mechanism of the action of insulin as a survival factor for 6-day embryonic lens explants, we compared the pattern of cell cycle markers (c-fos, c-jun, c-myc, p53, histone H3, thymidine kinase, and cyclin B) in both apoptotic and differentiating lens explants. In the presence of insulin, the expression of c-fos and c-jun was down-regulated after an initial induction. Expression of these genes was also induced in the absence of insulin, but mRNA levels remained elevated as the cells underwent apoptosis. In contrast, expression of c-myc, p53, histone H3, thymidine kinase, and cyclin B showed only minor differences in differentiating and apoptotic cells. Since c-fos and c-jun have been shown to play a role in apoptosis in other cell types, the ability of insulin to regulate expression of these genes may be central to its ability to act as a survival factor for lens epithelial cells. PMID: 8547223 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 575: Gen Comp Endocrinol. 1995 Aug;99(2):127-36. Changes in proto-oncogene activity in the testis of the frog, Rana esculenta, during the annual reproductive cycle. Chieffi P, Minucci S, Cobellis G, Fasano S, Pierantoni R. Dipartimento di Fisiologia Umana e Funzioni Biologiche Integrate F. Bottazzi, II Universita di Napoli, Italy. Proto-oncogenes are said to influence the regulation of cellular growth and differentiation. Myc, Fos, Jun, and Mos protein localization has been studied by immunocytochemistry in the testis of the frog, Rana esculenta, during the annual reproductive cycle. Oncoproteins have been localized in the primary and secondary (I and II) spermatogonia (SPG). Myc and Mos also appear in I and II spermatocytes (SPC) while Jun appears in II SPC. Myc, Fos, and Jun in SPG translocate in the nucleus during the periods of active spermatogenesis. Myc, Fos, and Jun are also localized in Sertoli cells. Fos is present in interstitial cells during the period characterized by the androgen peak which precedes the sharp increase of estradiol. It is suggested that proto-oncogene activity exerts a regulatory role in steroidogenesis and spermatogenesis. PMID: 8536921 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 576: J Clin Invest. 1995 Aug;96(2):842-7. Linoleic acid and its metabolites, hydroperoxyoctadecadienoic acids, stimulate c-Fos, c-Jun, and c-Myc mRNA expression, mitogen-activated protein kinase activation, and growth in rat aortic smooth muscle cells. Rao GN, Alexander RW, Runge MS. Division of Cardiology, University of Texas Medical Branch, Galveston 77555, USA. Previous studies from other laboratories suggest that linoleic acid and its metabolites, hydroperoxyoctadecadienoic acids, play an important role in modulating the growth of some cells. A correlation has been demonstrated between hydroperoxyoctadecadienoic acids and conditions characterized by abnormal cell growth such as atherosclerosis and psoriasis. To determine if linoleic acid and its metabolites modulate cell growth in atherosclerosis, we measured DNA synthesis, protooncogene mRNA expression, and mitogen-activated protein kinase (MAPK) activation in vascular smooth muscle cells (VSMC). Linoleic acid induces DNA synthesis, c-fos, c-jun, and c-myc mRNA expression and MAPK activation in VSMC. Furthermore, nordihydroguaiaretic acid, a potent inhibitor of the lipoxygenase system, significantly reduced the growth-response effects of linoleic acid in VSMC, suggesting that conversion of linoleic acid to hydroperoxyoctadecadienoic acids (HPODEs) is required for these effects. HPODEs also caused significant induction of DNA synthesis, protooncogene mRNA expression, and MAPK activation in growth-arrested VSMC, suggesting that linoleic acid and its metabolic products, HPODEs, are potential mitogens in VSMC, and that conditions such as oxidative stress and lipid peroxidation which provoke the production of these substances may alter VSMC growth. PMID: 7635978 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 577: Kidney Int. 1995 Aug;48(2):383-9. Cadmium-induced expression of immediate early genes in LLC-PK1 cells. Matsuoka M, Call KM. Department of Molecular and Cellular Toxicology, Harvard School of Public Health, Boston, Massachusetts, USA. To identify molecular mechanisms underlying renal cell damage by cadmium, the effect of this heavy metal on the level of immediate early genes (IEGs) transcripts in LLC-PK1 cells was studied. Cadmium chloride (CdCl2) induced the expression of four IEGs examined, but with differing time courses. The level of c-fos mRNA peaked at 30 minutes, and then decreased. The levels of c-jun and c-myc transcripts reached a maximum at one hour, and remained elevated up to four hours. Egr-1 mRNA level peaked at one hour, and returned to the control level by three hours. Experiments with cycloheximide and actinomycin D showed, respectively, that induction of IEGs by cadmium occurred in a protein synthesis-independent and transcriptional activation-dependent manner. Cadmium induction of c-fos mRNA was reduced markedly by the intracellular calcium chelator, bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)-ester (BAPTA/AM), and was decreased partially by a protein kinase C (PKC) inhibitor, 1-(5-isoquinolinylsulfonyl)-2- methylpiperazine (H-7). These data indicate that IEG induction by cadmium requires intracellular calcium mobilization and occurs in part by a PKC-dependent pathway. Exposure of LLC-PK1 cells to CdCl2 (20 microM for 1 to 24 hr) resulted loss of cell viability and DNA fragmentation, which was indicative of apoptosis. PMID: 7564105 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 578: Yakugaku Zasshi. 1995 Aug;115(8):570-83. [Structure and function of IL-5 receptor] [Article in Japanese] Takatsu K. Department of Immunology, University of Tokyo, Japan. Interleukin-5 (IL-5) regulates the production and function of B cells, eosinophils and basophils. In particular, IL-5 plays a critical role in the development of CD5-positive B (B-1) cells. The pleiotropic activity of IL-5 on target cells is directly dependent on the initial binding to IL-5 specific cell-surface receptor (IL-5R). The IL-5 signals are mediated through the high affinity IL-5R which is composed of two different polypeptide chains, alpha and beta. The alpha chain is a membrane-penetrated glycoprotein that specifically binds IL-5 and retains features common to the cytokine receptor superfamily. The beta chain by itself does not bind IL-5, but it can convert the low affinity IL-5R into the high affinity IL-5R and in indispensable for IL-5 signal transduction. The beta chain is shared among receptors for IL-5, IL-3 and GM-CSF and is called beta c. The cytoplasmic comains of both IL-5R alpha and beta c are essential for signal transduction. The membrane proximal proline-rich sequence of the cytoplasmic domain of IL-5R alpha was found to be essential for the IL-5-induced proliferative response, expression of nuclear proto-oncogenes such as c-jun, c-fos and c-myc, and activation of Bruton's tyrosine and JAK2 kinases. Furthermore, JAK2 activation correlates with proline residues in Pro-Pro-X-Pro motif in the cytoplasmic domain of IL-5R alpha. These results indicate that activation of JAK2 and its substrate is critical to coupling IL-5-induced tyrosine phosphorylation and ultimately mitogenesis. I will discuss about molecular mechanisms of IL-5 signaling and B cell defect in X-linked immunodeficient mice in relation to IL-5 signaling. Publication Types: Review Review, Tutorial PMID: 7473055 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 579: Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):7041-5. A splice variant of alpha 6 integrin is associated with malignant conversion in mouse skin tumorigenesis. Tennenbaum T, Belanger AJ, Glick AB, Tamura R, Quaranta V, Yuspa SH. Laboratory of Cellular Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. The epithelial-specific integrin alpha 6 beta 4 is suprabasally expressed in benign skin tumors (papillomas) and is diffusely expressed in carcinomas associated with an increase in the proliferating compartment. Analysis of RNA samples by reverse transcriptase-PCR and DNA sequencing revealed that chemically or oncogenically induced papillomas (n = 8) expressed a single transcript of the alpha 6 subunit, identified as the alpha 6 A splice variant. In contrast, carcinomas (n = 13) expressed both alpha 6A and an alternatively spliced form, alpha 6B. Primary keratinocytes and a number of keratinocyte cell lines that vary in biological potential from normal skin, to benign papillomas, to well-differentiated slowly growing carcinomas exclusively expressed alpha 6A. However, I7, an oncogene-induced cell line that produces highly invasive carcinomas, expressed both alpha 6A and alpha 6B transcript and protein. The expression of alpha 6B in I7 cells was associated with increased attachment to a laminin matrix compared to cell lines exclusively expressing alpha 6A. Furthermore, introduction of an alpha 6B expression vector into a papilloma cell line expressing alpha 6A increased laminin attachment. When a papilloma cell line was converted to an invasive carcinoma by introduction of the v-fos oncogene, the malignant cells expressed both alpha 6A and alpha 6B, while the parent cell line and cells transduced with v-jun or c-myc, which retained the papilloma phenotype, expressed only alpha 6A. Comparative analysis of alpha 6B expression in cell lines and their derived tumors indicate that alpha 6B transcripts are more abundant in tumors than cell lines, and alpha 6B is expressed to a greater extent in poorly differentiated tumors. These results establish a link between malignant conversion and invasion of squamous tumor cells and the regulation of transcript processing of the alpha 6 beta 4 integrin. PMID: 7624366 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 580: Int J Cancer. 1995 Jul 4;62(1):107-14. The effect of active oxygen generated by xanthine/xanthine oxidase on genes and signal transduction in mouse epidermal JB6 cells. Singh N, Aggarwal S. Department of Biochemistry, All India Institute of Medical Sciences, New Delhi. To gain insight into the gene regulation and signal transduction effects of active oxygen in tumour promotion and progression, we studied the effect of active oxygen generated extracellularly by xanthine/xanthine oxidase (X/XO) in promotion-insensitive (P-), promotion-sensitive (P+) and transformed (Tx) mouse epidermal JB6 cells. Active oxygen inhibited growth, particularly of P- cells and increased poly ADPR transferase activity and PKC activity more significantly in P- cells. No phenotypic differences in the distribution pattern of PKC isotypes alpha, beta and gamma were seen in JB6 cells. PKC alpha was expressed abundantly, whereas beta and gamma were not detected. Basal levels of the antioxidant enzymes catalase and CuZn. Superoxide dismutase were higher in P+ and Tx cells. X/XO resulted in an initial decrease in the activity of these enzymes, followed by recovery or transient induction in Tx and P+ cells. X/XO induced c-myc and c-fos expression in JB6 cells, with c-fos induction being more pronounced in P- cells, whereas a biphasic increase in c-jun was seen in P+ cells. These early genes may play a role in proliferation whereas post-translational poly ADP-ribosylation and, perhaps, phosphorylation suggest a genetic-epigenetic mechanism in oxidant tumour promotion and progression. PMID: 7601557 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 581: Endocrinology. 1995 Jul;136(7):2975-83. Retinoic acid inhibits estrogen-induced uterine stromal and myometrial cell proliferation. Boettger-Tong HL, Stancel GM. Department of Pharmacology, University of Texas Medical School at Houston 77225, USA. Retinoic acid, a potent natural derivative of vitamin A, influences proliferation in many cell types. However, little is known about the role of retinoic acid in estrogen-induced proliferation in normal physiological systems. In this study we sought to determine if in vivo administration of retinoic acid influences the proliferation of a normal estrogen target tissue, the immature rat uterus. The results indicate that treatment of animals with 30 mg/kg all-trans-retinoic acid for 3 days before 17 beta-estradiol (E2) administration diminishes DNA synthesis and cell division by approximately 50% in uterine stromal and myometrial cells. Luminal epithelial cell proliferation is not inhibited, indicating that the antiproliferative effects of all-trans-retinoic acid treatment are cell type-specific. The inhibition is retinoid-specific and fully reversible 1 week after discontinuing all-trans-retinoic acid treatment. The inhibitory effect of all-trans-retinoic acid is not due to a change in E2 receptor levels assessed by ligand binding. E2 induction of c-jun, a gene expressed primarily in myometrial cells, is unaffected in retinoid-treated animals. This is the first demonstration that retinoic acid inhibits estrogen-induced proliferation of uterine stromal and myometrial cells in a physiological setting. PMID: 7789323 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 582: Mol Carcinog. 1995 Jul;13(3):146-56. Effects of 12-O-tetradecanoylphorbol-13-acetate on human papillomavirus type 16-positive keratinocytes at different stages of transformation. Sears WL, Goto-Mandeville R, Mirapuri M, Braun L. Department of Pathology and Laboratory Medicine, Brown University, Providence, RI 02912, USA. Normal human keratinocytes grown under serum-free conditions can be triggered to differentiate by exposure to serum or to the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA). We found that TPA treatment of human papillomavirus (HPV) type 16-immortalized cells in culture induced formation of cornified envelopes indicative of squamous differentiation. Concurrent with differentiation, TPA inhibited the expression of HPV 16 E6 and E7 mRNA transcripts. Adaptation of the immortalized cells to growth in serum-containing medium led to the selection of a subpopulation of HPV-transformed cells that was resistant to TPA-induced differentiation. In this cell line, a transient suppression of HPV transcripts was observed at 5 h, whereas in differentiation-resistant, carcinoma-derived lines, TPA had little effect on HPV oncogene expression. c-myc transcripts were suppressed for the duration of exposure to TPA in only the differentiation-competent cells; c-fos and c-jun were transiently induced in all cell lines. Transforming growth factor-alpha mRNAs were also increased approximately eightfold as HPV 16-immortalized cells were induced to differentiate. These results demonstrate that, in HPV 16-immortalized keratinocytes, acquisition of resistance to inducers of squamous differentiation is accompanied by altered regulation of cell growth and gene expression. PMID: 7619217 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 583: J Clin Invest. 1995 Jul;96(1):69-77. Immediate-early gene induction and MAP kinase activation during recovery from metabolic inhibition in cultured cardiac myocytes. Yao A, Takahashi T, Aoyagi T, Kinugawa K, Kohmoto O, Sugiura S, Serizawa T. Second Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan. To investigate how cardiac myocytes recover from a brief period of ischemia, we used a metabolic inhibition (MI) model, one of the in vitro ischemic models, of chick embryo ventricular myocytes, and examined the induction of immediate-early (IE) genes mRNAs and the activity of mitogen-activated protein (MAP) kinase. We performed Northern blot analysis to study the expression of c-jun, c-fos, and c-myc mRNAs during MI using 1 mM NaCN and 20 mM 2-deoxy-d-glucose, and also during the recovery from MI of 30 min. The c-fos mRNA was induced transiently at 30 and 60 min during the recovery. The expression of c-jun mRNA was significantly augmented at 30, 60, 90, and 120 min during the recovery (3.0-, 4.7-, 2.4-, and 1.9-fold induction, respectively) and so did the expression of c-myc mRNA (1.4-, 1.7-, 1.8-, and 2.0-fold induction, respectively). In contrast, the levels of these mRNAs remained unchanged during MI. The electrophoretic mobility shift assay revealed that AP-1 DNA binding activity markedly increased at 120 min during the recovery. When the cells were pretreated with protein kinase C (PKC) inhibitors, 100 microM H-7 or 1 microM staurosporine, the induction of c-jun mRNA at 60 min during the recovery was markedly suppressed (95 or 82% reduction, respectively). The c-jun induction was partially inhibited when the cells were treated with 2 mM EGTA during MI and the recovery (42% reduction). MAP kinase activity quantified with in-gel kinase assay was unchanged during MI, but significantly increased at 5, 10, and 15 min during the recovery (3.0-, 4.1-, and 3.4-fold increase, respectively). S6 kinase activity was also augmented significantly at 15 min during the recovery. Thus, these data suggest that IE genes as well as MAP kinase may play roles in the recovery process of cardiac myocytes from MI, and that the augmentation of c-jun expression needs the activation of PKC and to some extent, [Ca2+]i. PMID: 7615838 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 584: Mol Cell Endocrinol. 1995 Jul;112(1):35-43. Estrogen action in target cells: selective requirements for activation of different hormone response elements. Hyder SM, Shipley GL, Stancel GM. Department of Pharmacology, University of Texas Medical School, Houston 77225, USA. RENE1 cells, an estrogen receptor positive rat uterine endometrial cell line immortalized with the E1A oncogene, were analyzed for the presence of estrogen-dependent signal transduction pathways using the induction of transfected as well as endogenous genes. RENE1 cells express the estrogen receptor as analyzed by Northern blots and ligand binding assays (40 fmoles/mg protein). The receptor system appears functional, based on the induction of reporter constructs containing the consensus estrogen response element (ERE) in transient transfection assays and alterations in endogenous transcripts visualized by utilizing differential display methodology. However, neither transfected repoter constructs containing the c-fos ERE, nor the endogenous c-fos, c-jun, or c-myc genes are induced by estrogens in these cells despite being induced by estrogens in the uterus in vivo. In addition, estradiol did not induce endogenous c-fos expression or the activity of CAT reporters containing the c-fos ERE in a stable transfectant of RENE1 cells with a 3-fold elevation in estrogen receptor content. Under identical conditions, TPA and serum rapidly induce c-fos transcription in RENE1 cells, indicating that the lack of inducibility by estradiol is not due to a general inhibitor of transcription of these genes. These results suggest that RENE1 cells lack factors present in normal uterine cells which are required for the estrogenic induction of a specific subset(s) of EREs. These observations support the generally evolving hypothesis that steroid hormones may act through composite response elements via interactions with other transcription factors, in addition to functioning as homodimers at classical palindromic response elements. PMID: 7589783 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 585: Biochem Mol Biol Int. 1995 Jul;36(3):465-74. Hepatocyte growth factor-induced signal transduction in two normal mouse epithelial cell lines. Johnson M, Kochhar K, Nakamura T, Iyer A. Rush Medical School, Department of Biochemistry, Chicago, Illinois 60610, USA. We have investigated HGF-induced signal transduction in two normal mouse epithelial cell lines (M23 and MM55). Both cell lines display HGF-induced mitogenesis and high level HGF-induced autophosphorylation of MET/HGFR. In both M23 and MM55 cells, HGF induces association with MET/HGFR and increased tyrosine phosphorylation of the SH2-domain containing proteins PI3K, GAP and NCK. PLC-gamma exhibited neither HGF-induced increases in tyrosine phosphorylation nor an association with MET/HGFR in these cell lines. Additionally, HGF induced increased transcription of c-fos, c-jun, junB, junD, and c-myc early response genes in both cell lines. We therefore suggest that the second messenger proteins PI3K, GAP and NCK, and possibly the protein products of the c-fos, c-jun, junB, junD and c-myc genes, are important elements in the HGF-induced mitogenic pathway in the normal mouse epithelial cell lines M23 and MM55. PMID: 7549943 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 586: J Cell Physiol. 1995 Jun;163(3):623-30. Activation of junB and c-myc primary response genes by interleukin 9 in a human factor-dependent cell line. Kang LY, Yang YC. Walther Oncology Center, Department of Biochemistry/Molecular Biology, Indiana University School of Medicine, Indianapolis 46202, USA. Interleukin 9 (IL-9) stimulates the proliferation of various hematopoietic cell types. To elucidate the molecular mechanisms underlying the cell proliferation action, immediate-early gene expression elicited by IL-9 in a human factor-dependent cell line, MO7e, was studied. IL-9 stimulation resulted in a rapid and transient elevation of primary response genes including junB and c-myc. The differential effects of protein kinase inhibitors, herbimycin A, genistein, and H-7 on the steady-state mRNA level and the transcription rate of junB and c-myc genes triggered by IL-9 were also investigated. Herbimycin A, but not genistein, specifically inhibited the expression of junB steady-state mRNA and the in vitro transcription of the junB gene. IL-9-enhanced c-myc gene expression was completely inhibited by both herbimycin A and genistein at the level of transcriptional initiation. H-7 failed to inhibit c-myc, but partially abolished junB mRNA induction. The role of protein kinase C in IL-9-mediated junB induction was also examined. The different responses of junB and c-myc messages to protein kinase inhibitors suggested that more than one pathway may be involved in IL-9-mediated signal transduction which leads to the expression of junB and c-myc genes. PMID: 7775604 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 587: J Cell Biochem. 1995 Jun;58(2):135-50. Cells en route to apoptosis are characterized by the upregulation of c-fos, c-myc, c-jun, cdc2, and RB phosphorylation, resembling events of early cell-cycle traverse. Pandey S, Wang E. Bloomfield Centre for Research in Aging, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, Department of Medicine, McGill University, Montreal, Quebec, Canada. Density-arrested quiescent murine Balb/c-3T3 cells are dependent upon growth factors for their survival. Withdrawal of serum from their medium induces rapid cell death, the mechanism of which is not yet fully understood. We have studied the effect of serum deprivation on density-inhibited quiescent Swiss 3T3 cells and found that they undergo rapid cell death upon total withdrawal of serum. The nature of this cell death is similar to apoptosis, as shown by cellular and nuclear morphology and DNA fragmentation into oligonucleosomal fragments. Investigating the regulation of early cell-cycle genes during this process, we found that c-myc, c-jun, c-fos, and cdc2 protein presence is induced after serum deprivation, when the phosphorylated form of the RB protein also appears. The upregulation of these genes' protein products is coupled with the appearance of PCNA, a proliferation-specific nuclear antigen, as well as significant incorporation of BrdU, which may reflect DNA repair activity; in situ analysis shows that BrdU-positive cells are also positive for DNA fragmentation. These results suggest that en route to apoptosis, cells undergo events typical of early cell-cycle traverse by expressing early G1 genes and may even experience the late G1/S phase boundary, as shown by the presence of PCNA. However, the demonstrated ability of these cells to traverse the G1 phase of the cell cycle seems to be an abortive event, since they die shortly afterwards. Publication Types: Review Review, Tutorial PMID: 7673322 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 588: Mol Biol Cell. 1995 Jun;6(6):627-36. Characterization of cis-regulatory elements of the c-myc promoter responding to human GM-CSF or mouse interleukin 3 in mouse proB cell line BA/F3 cells expressing the human GM-CSF receptor. Watanabe S, Ishida S, Koike K, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Interleukin 3 (IL-3) or granulocyte macrophage colony-stimulating factor (GM-CSF) activates c-fos, c-jun, and c-myc genes and proliferation in both hematopoietic and nonhematopoietic cells. Using a series of deletion mutants of the beta subunit of human GM-CSF receptor (hGMR) and inhibitors of tyrosine kinase, two distinct signaling pathways, one for activation of c-fos and c-jun genes, and the other for cell proliferation and activation of c-myc gene have been elucidated. In contrast to wealth of information on the pathway leading to activation of c-fos/c-jun genes, knowledge of the latter is scanty. To clarify the mechanisms of activation of c-myc gene by cytokines, we established a transient transfection assay in mouse proB cell line BA/F3 cells expressing hGMR. Analyses of hGMR beta subunit mutants revealed two cytoplasmic regions involved in activation of the c-myc promoter, one is essential and the other is dispensable but enhances the activity. These regions are located at the membrane proximal and the distal regions covering amino acid positions 455-544 and 544-589, respectively. Characterization of cis-acting regulatory elements of the c-myc gene showed that the region containing the P2 promoter initiation site is sufficient to mediate the response to mIL-3 or hGM-CSF. Electrophoretic mobility shift assay using an oligonucleotide corresponding to the distal putative E2F binding site revealed that p107/E2F complex, the negative regulator of E2F, decreased, and free E2F increased after mIL-3 stimulation. These results support the thesis that mIL-3 or hGM-CSF regulates the c-myc promoter by altering composition of the E2F complexes at E2F binding site. PMID: 7579683 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 589: Oncogene. 1995 May 18;10(10):2037-49. c-fos is a positive regulator of carcinogen enhancement of adenovirus transformation. Su ZZ, Yemul S, Stein CA, Fisher PB. Department of Urology, Institute of Cancer Research, Columbia University, College of Physicians and Surgeons, New York, New York 10032, USA. The early gene expression changes mediating carcinogen enhancement of viral transformation (CET) remain to be elucidated. A model cell culture system has been developed that is now permitting a molecular analysis of CET. Pretreatment of cloned rat embryo fibroblast (CREF) cells with methyl methanesulfonate (MMS) prior to infection with the cold-sensitive host-range type 5 adenovirus mutant, H5hr1, results in a dose-dependent increase in viral transformation. The present study investigates the role of immediate-early response genes, specifically c-fos, in the CET process. MMS pretreatment, alone or in combination with infection with H5hr1 temporally and differentially increases c-fos, c-jun, jun-B, jun-D and c-myc steady-state mRNA levels. Maximum induction occurs with c-fos and c-jun 8 to 12 h posttreatment and the magnitude of response is generally greatest in CREF cells pretreated with MMS and then infected with H5hr1. Enhancement in RNA levels is observed in the presence of cycloheximide indicating that ongoing protein synthesis is not required for induction of c-fos, c-jun, jun-B or c-myc expression. Nuclear run-on analysis indicates an enhancement in transcriptional rates for c-fos, c-jun, jun-B and c-myc in CREF cells treated with MMS or MMS plus infection with H5hr1. A requirement for elevated c-fos in the early stages of CET is indicated by the ability of c-fos antisense oligonucleotides to prevent the CET process. Direct evidence implicating early increases in c-fos as a mediator of the CET process is demonstrated by stably expressing mouse mammary tumor virus promoter-regulated human sense and antisense c-fos genes in CREF cells. Induction of c-fos sense expression by dexamethasone (DEX) in the absence of MMS treatment results in enhanced c-fos mRNA, Fos protein, AP-1 DNA-binding activity and H5hr1-induced transformation and CET. Induction of c-fos expression by DEX in stable c-fos-sense CREF constructs also results in elevated levels of c-jun, jun-B and c-myc mRNA and protein. Conversely, induction of c-fos antisense expression prevents the increase in c-fos mRNA, Fos protein and AP-1 DNA-binding activity and eliminates CET. In the antisense-c-fos constructs, increases in c-jun, jun-B and c-myc mRNA and protein normally induced by MMS also are not apparent. Thus, induction or inhibition in c-fos expression affects the level of expression of additional immediate-early response genes, including c-jun, jun-B and c-myc.(ABSTRACT TRUNCATED AT 400 WORDS) PMID: 7761104 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 590: Br J Cancer. 1995 May;71(5):1018-24. Nuclear proto-oncogene products transactivate the human papillomavirus type 16 promoter. Nurnberg W, Artuc M, Vorbrueggen G, Kalkbrenner F, Moelling K, Czarnetzki BM, Schadendorf D. Universitatsklinikum Rudolf Virchow, Hautklinik, Freie Universitat, Berlin, Germany. Human papillomavirus (HPV) type 16 and 18 viral genomes are frequently detected in cervical and penile cancer biopsies. Although this strongly suggests a prominent role for HPV infection in the development of genital cancer, other genetic or environmental factors are also involved. Genital cancer is postulated to result from loss of cellular control functions, which leads to an unregulated expression of HPV oncogenic proteins. In our study, we determined the trans-activating properties of nuclear proto-oncogene proteins c-Fos, c-Jun and c-Myc on P97 enhancer/promoter activity of HPV16. Using a CAT-reporter construct containing the HPV16 enhancer/promoter element, we investigated the trans-activating effects of c-Fos, c-Jun, c-Myc, and E2 in cervical HT-3 cells. c-Fos and c-Jun overexpression resulted in a 3.3- and 3.1-fold up-regulation of CAT activity. Only 2-fold induction was determined by co-transfection with c-myc and the viral transcription factor E2. Based on these findings, we investigated the expression of HPV DNA (16 and 18) as well as nuclear proto-oncogenes (c-fos, c-jun and c-myc) in nine cervical cancers by in situ hybridisation. In six out of nine carcinomas, HPV16 and/or HPV18 DNA was detectable. All tumours showed an intense and homogeneous expression of c-fos and c-jun mRNA, while the signal for c-myc was detectable only in four specimens. These data suggest that deregulation of nuclear proto-oncogene expression may contribute to an overexpression of HPV-derived oncogenic proteins (E6 and E7), which is generally hypothesised to be an important step in the malignant transformation of HPV-associated tumours. PMID: 7734293 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 591: Biochem Soc Trans. 1995 May;23(2):329S. Immunolocalisation of proto-oncogene expression in mechanically stimulated skeletal muscle. Lee DM, Dawes NJ, Cox VM, Hesketh JE, Goldspink DF. Dept. of Clinical Medicine, University of Leeds, U.K. PMID: 7672360 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 592: Biochem Soc Trans. 1995 May;23(2):328S. Growth hormone induction and somatostatin suppression of hepatic proto-oncogenes and IGF-I in the rat. Penman J, Liu S, Belchetz P, Goldspink DF. Dept. of Clinical Medicine, University of Leeds, U.K. PMID: 7672359 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 593: Cell Growth Differ. 1995 May;6(5):505-13. Involvement of ornithine decarboxylase and polyamines in glucocorticoid-induced apoptosis of rat thymocytes. Desiderio MA, Grassilli E, Bellesia E, Salomoni P, Franceschi C. Institute of General Pathology and Consiglio Nazionale delle Richerche Center for Research on Cell Pathology, University of Milano, Italy. Ornithine decarboxylase (ODC), the first and rate-limiting enzyme of polyamine metabolism, has been shown to be required for entry into and progression through the cell cycle. However, the role of ODC and polyamines in apoptosis remains to be determined. We have examined ODC expression and polyamine levels in thymocytes activated to undergo apoptosis by dexamethasone treatment. We have demonstrated a rapid and reversible induction of ODC (mRNA and activity), as previously reported for the mRNA expression of other "early" genes, c-fos, c-jun, and c-myc, in the same experimental model. Surprisingly, polyamine levels diminished progressively starting at 2-4 h after dexamethasone treatment, and spermine was depleted at 8-12 h. This seemed to be relevant since increasing the intracellular polyamine levels by exogenous spermine administration prevented the DNA "laddering" (2-4 h) and the DNA loss from the nucleus (8-18 h) due to dexamethasone treatment. Moreover, the activities of spermidine/spermine N1-acetyltransferase, which controls the cytosolic polyamine interconversion pathway, and of spermidine N8-acetyltransferase, which regulates the nuclear pool and functions of polyamines, were measured in apoptotic cells. Spermidine/spermine N1-acetyltransferase activity progressively increased and might be responsible for spermidine and spermine excretion as acetyl derivatives. In contrast, spermidine N8-acetyltransferase activity remained unchanged. A completely different scenario was observed in proliferating concanavalin A-treated thymocytes, studied for comparison. In this case, polyamine levels increased, remaining at high values until 12 h. This is likely a consequence of the rapid and prolonged induction of ODC (mRNA and activity), accompanied by that of spermidine/spermine N1-acetyltransferase (mRNA and activity).(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 7647033 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 594: Anticancer Res. 1995 May-Jun;15(3):729-33. Differential oncogene expression and susceptibility to apoptosis in the human leukemia HL60 cell lines: implications for etoposide resistance. Eliot HE, Borner MM, Sinha BK. Biochemical and Molecular Pharmacology Section, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA. Mechanisms of etoposide (VP-16) resistance have been evaluated in a human promyelocytic leukemia HL60 cell line. HL60 resistant (HL60/AR) cells were selected for resistance with adriamycin and were 250-fold resistant to VP-16. We have found that while a significantly higher (10 to 15-fold more) dose of VP-16 was required to induce similar amounts of SDS-KCI-precipitable DNA-protein complex formation in the resistant cell line, there was no difference in the repair of VP-16-induced DNA damage, indicating that differential DNA repair was not involved in VP-16 resistance in HL60 cells. VP-16 treatment significantly inhibited c-myc expression and induced c-jun and c-fos expressions in sensitive cells. In contrast, VP-16 had no effect on c-myc, c-jun or c-fos expressions in resistant cells. The level of bcl2 oncogene was similar in both cell lines; however, treatment with VP-16 resulted in a time- and dose-dependent degradation of the genomic DNA into oligo-sized DNA only in the sensitive cells, indicating that differential expressions of oncogenes (c-myc, c-jun, and c-fos) and susceptibility to apoptosis may play important roles in the sensitivity and resistance to VP-16 in HL60 cells. PMID: 7645949 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 595: Am J Physiol. 1995 May;268(5 Pt 2):F793-801. Subtotal but not unilateral nephrectomy induces hyperplasia and protooncogene expression. Terzi F, Ticozzi C, Burtin M, Motel V, Beaufils H, Laouari D, Assael BM, Kleinknecht C. Institut National de la Sante et de la Recherche Medicale Unite 192, Hopital Necker, Enfants-Malades, Paris, France. It is generally accepted that renal compensatory growth after unilateral nephrectomy (Uni) is due to prominent hypertrophy with no involvement of protooncogenes. Neither the balance between hypertrophy and hyperplasia nor the expression of the early-growth-related genes has been studied after subtotal nephrectomy (Nx). The occurrence of cystic tubular dilatations after Nx may suggest an excessive cell proliferation in this model. We measured DNA, RNA, and protein content, number of nuclei per tubular section, as well as c-fos, c-jun, c-myc, c-H-ras, c-sis, and c-erb-B2 protooncogene expression in kidneys taken at time of surgery and 2, 7, and 14 days after sham operation (control rats), Uni, or Nx. After Uni, hyperplasia was greater than expected (+79% for DNA at day 14) and was associated with moderate hypertrophy (+11% for protein/DNA ratio). After Nx, compensatory growth was due only to hyperplasia (+117% for DNA at day 14), with unchanged protein/DNA ratio (vs. Uni, P < 0.02). The greater hyperplasia after Nx was confirmed by nuclei counting. The protooncogene mRNA expression was constantly absent in control and Uni rats, whereas that of c-fos and c-jun genes was detected in Nx rats at day 14 with a 2- to 12-fold increment. The c-fos and c-jun protein levels were also increased at that time in Nx rats. This suggests the following: 1) the cellular events following Uni and Nx are not the same, and 2) the late protooncogene expression in Nx exclusively could favor a particular type of cell proliferation possibly more related with cystic formation than with actual compensatory growth. PMID: 7539585 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 596: Mol Carcinog. 1995 May;13(1):44-9. Combined effects of ionizing radiation and cycloheximide on gene expression. Woloschak GE, Felcher P, Chang-Liu CM. Center for Mechanistic Biology and Biotechnology, Argonne National Laboratory, IL 60439-4833, USA. We performed experiments to determine the effects of ionizing radiation exposure on expression of genes such as beta-actin, c-fos, histone H4, c-myc, c-jun, Rb, and p53 after exposure of Syrian hamster embryo (SHE) cells to the protein synthesis inhibitor cycloheximide. The purpose of these experiments was to determine the role of a labile protein in the radiation-induced response. The results revealed that when ionizing radiation (either fission-spectrum neutrons or gamma rays) was administered 15 min after cycloheximide treatment of SHE cells, the radiation exposure reduced cycloheximide-mediated gene induction of c-fos, histone H4, and c-jun. In addition, dose-rate differences were found when radiation exposure most significantly inhibited the cycloheximide response. Our results suggest that ionizing radiation does not act as a general protein-synthesis inhibitor and that the presence of a labile protein is required for the maintenance of specific gene transcription and mRNA accumulation after radiation exposure, especially at high dose-rates. PMID: 7539271 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 597: Cell. 1995 Apr 21;81(2):223-31. Three distinct IL-2 signaling pathways mediated by bcl-2, c-myc, and lck cooperate in hematopoietic cell proliferation. Miyazaki T, Liu ZJ, Kawahara A, Minami Y, Yamada K, Tsujimoto Y, Barsoumian EL, Permutter RM, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. Two interleukin-2 receptor-dependent signaling pathways have thus far been identified: the c-fos/c-jun induction pathway mediated by src family protein-tyrosine kinases and the c-myc induction pathway. Here, we provide evidence for the existence of a third, rapamycin-sensitive pathway, which results in the induction of another proto-oncogene, bcl-2. In the hematopoietic cell line BAF-B03, the expression of any two of lckF505 (an active form of p56lck), Bcl-2, or c-Myc is sufficient to promote transit of the cell cycle, regardless of the activation state of the third pathway. We also provide evidence that epidermal growth factor receptor signaling may act through the same pathway that involves p56lck. These studies demonstrate a novel approach to dissecting signaling pathways regulating cellular proliferation. PMID: 7736574 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 598: J Biol Chem. 1995 Apr 21;270(16):9615-21. Granulocyte macrophage-colony stimulating factor-dependent replication of polyoma virus replicon in hematopoietic cells. Analyses of receptor signals for replication and transcription. Watanabe S, Ito Y, Miyajima A, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Granulocyte macrophage-colony stimulating factor (GM-CSF) stimulates proliferation of various hematopoietic cells. Using cytoplasmic deletion mutants of the human GM-CSF receptor (hGMR) beta subunit and tyrosine kinase inhibitors, we previously showed that distinct signaling pathways of hGMR are involved in the induction of c-fos/c-jun mRNAs and of c-myc mRNA/cell proliferation. We used polyoma virus (Py) replicon to analyze the initiation of DNA replication induced by hGM-CSF in mouse BA/F3 pro-B cells expressing hGMR. hGM-CSF efficiently stimulated Py replication in the presence of Py enhancer and Py large T antigen supplied in trans. Analyses of Py enhancer mutants revealed that hGM-CSF promoted Py replication and activated transcription of the Py early promoter through the PEA3/PEBP5 region of Py enhancer. The membrane proximal region of hGMR beta subunit is required for activation of PEA3/PEBP5-dependent replication which is also required for activation of DNA synthesis in the host cells. In contrast, a more distal region which is essential for activation of c-fos and c-jun genes is required for the PEA3/PEBP5-dependent transcription of Py early promoter. These results indicate that distinct signaling pathways of hGMR are required to activate PEA3/PEBP5-dependent replication and transcription although the same enhancer is required for both activities. PMID: 7721893 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 599: Oncogene. 1995 Apr 6;10(7):1325-33. Cell cycle arrest by tyrosine kinase Abl involves altered early mitogenic response. Mattioni T, Jackson PK, Bchini-Hooft van Huijsduijnen O, Picard D. Departement de Biologie Cellulaire, Universite de Geneve, Switzerland. Activated forms of the nuclear and cytoplasmic tyrosine kinase c-Abl are completely cytoplasmic and oncogenic. The overexpression of c-Abl, and in certain fibroblast cell lines even of v-Abl, leads to a cell cycle arrest revealing an alternative Abl function. To facilitate the analysis of this growth inhibitory function we have taken advantage of regulable Abl-estrogen receptor (ABL:ER) fusion proteins. Oncogenic in the presence of estrogen, they are reversibly switched to inhibit cell proliferation upon removal of hormone. Using this system, we demonstrate that inhibition is effected by Abl derivatives which we have previously shown to be hypo-phosphorylated and to have low kinase activity. Since an almost exclusively cytoplasmic ABL:ER protein is fully growth inhibitory, relevant interactions may occur in the cytoplasm. We identify the cell cycle arrest as an early G1 or G0-like block. Interestingly, growth inhibition correlates with an altered expression pattern of early serum response genes; c-Jun mRNA and c-Fos protein levels are elevated in Abl-blocked cells. In view of the two functional modes of overexpressed Abl proteins, one can speculate that normal c-Abl may be involved in relaying growth regulatory signals from the membrane to the nucleus. PMID: 7731683 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 600: Exp Cell Res. 1995 Apr;217(2):477-83. The growth-inhibitory block of TGF-beta is located close to the G1/S border in the cell cycle. Kletsas D, Stathakos D, Sorrentino V, Philipson L. Institute of Biology, N.C.S.R. Demokritos, Athens, Greece. Transforming growth factor-beta (TGF-beta) inhibits DNA synthesis in dense cultures of young human embryonic fibroblasts and antagonizes the mitogenic action of platelet-derived growth factor (PDGF). The inhibition of the PDGF-BB action by TGF-beta was independent of the induction of mRNAs for the PDGF-A chain and PDGF-beta receptor, the predominant types of PDGF receptor in human fibroblasts. The TGF-beta-mediated inhibition did not influence the expression of various genes that are involved in the transition from the arrested (GO) state to the S phase of the cell cycle. Indeed, TGF-beta upregulated the "early" genes c-myc, c-fos, and junB and downregulated the growth arrest-specific (gas) genes. These results suggest that the inhibition of DNA synthesis by TGF-beta in human fibroblasts is independent of modulation of expression of early and gas genes, placing the TGF-beta block comparatively late in the GO to S transition. In cultures of senescent human fibroblasts TGF-beta stimulated DNA synthesis but, nevertheless, had the same effect as in young cells on the expression of PDGF chains and receptor genes, as well as on early and gas genes, with the exception of a significantly lower induction of c-fos. PMID: 7698248 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 601: Nippon Ronen Igakkai Zasshi. 1995 Apr;32(4):259-65. [Aging and cellular senescence] [Article in Japanese] Fujiwara Y. Department of Radiation Biophysics and Genetics, Kobe University School of Medicine. Mechanisms of aging involve genetic programs and error accumulation. Cellular aging is an aspect of organismal aging from a point of view of age-dependent declines of tissue cells during the postreproductive aging process and a parallelism between enhanced individual and cellular aging in some genetic progeroid syndromes. Cellular senescence involves the gene-directed inhibition of replicative potential of cells. Cell fusion analysis has indicated that senescent normal and presenescent Werner syndrome cells cause the dominant suppression of DNA synthesis in the partner of either actively growing cells or any cells of the four complementation groups of immortalized human cells. Membrane proteins produced in senescent cells showed the biphasic DNA synthesis-inhibiting activity when assayed for young cells. Senescent cells showed the strong transcriptional repressions of early serum responsive genes (c-fos, c-jun, c-myc), late responsive genes of transcription factor E2F1 and cyclin E. In addition, the protein levels of CDK2 and cyclin E are also extremely low, with an increased level of the p53-dependent p21 Cip 1 protein which inhibits the kinase activity of cyclins/CDKs by forming complexes. Such characteristic molecular factors and mechanisms feature irreversible G1-arrest in cellular senescence. PMID: 7616677 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 602: Ann N Y Acad Sci. 1995 Mar 27;752:394-405. Reactive oxygen intermediates (ROIs) are involved in the intracellular transduction of angiotensin II signal in C2C12 cells. Puri PL, Avantaggiati ML, Burgio VL, Chirillo P, Collepardo D, Natoli G, Balsano C, Levrero M. Fondazione Andrea Cesalpino, University of Rome La Sapienza, Italy. Increasing evidence suggests that angiotensin II may act as a growth factor for several muscle cell types. Angiotensin II stimulation activates many immediate early response genes like c-Fos, c-Jun, c-Myc and Egr-1 in both vascular smooth muscle cells and cardiomyocytes, independently of whether a hyperplastic or hypertrophic response is taking place. In this study we report that angiotensin II significantly stimulates AP1-driven transcription in mouse skeletal muscle cells C2C12 stably transfected with a TRE-tk-CAT plasmid in a dose-dependent manner (peak stimulation at 10(-5) M of angiotensin II). Moreover, angiotensin II increases the binding of the AP1 complex to its DNA target in both quiescent C2C12 myoblasts and in differentiated C2C12 myotubes. Most of the TRE-bound complexes in both unstimulated and angiotensin II-treated cells consist of c-jun/c-fos heterodimers. Using a set of different protein kinase inhibitors, including HA1004, H7, tyrphostin, genistein and staurosporine, we could demonstrate that the angiotensin II-induced AP1 binding increase is not mediated by the cAMP-dependent pathway and that protein kinase C and tyrosine kinases are involved. Treatment of C2C12 cells with H2O2 induces a dose-dependent increase in c-jun/c-fos heterodimer binding, specifically reverted by the cysteine derivative and glutathione precursor N-acetyl-L-cysteine (NAC). The observation that the induction by angiotensin II of both the AP1 DNA binding activity and DNA synthesis in quiescent C2C12 myoblasts is abolished by NAC strongly suggests a role for reactive oxygen intermediates (ROIs) in the intracellular transduction of angiotensin II signals for immediate early gene induction and for cell proliferation. Publication Types: Review Review, Tutorial PMID: 7755283 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 603: Oncogene. 1995 Mar 16;10(6):1119-23. Trichostatin A inhibits both ras-induced neurite outgrowth of PC12 cells and morphological transformation of NIH3T3 cells. Futamura M, Monden Y, Okabe T, Fujita-Yoshigaki J, Yokoyama S, Nishimura S. Oncogene Research Laboratory, Banyu Tsukuba Research Institute (Merck), Japan. During screening for inhibitors of ras-mediated differentiation of PC12 cells, trichostatin A (TSA) was isolated from the metabolites of Streptomyces as a potent inhibitor. TSA blocked both oncogenic ras- and NGF-induced neurite outgrowth from PC12 cells. However, addition of TSA 1 h after NGF-stimulation did not inhibit neuronal differentiation, suggesting that TSA affects an early step in the NGF-signaling pathway mediated by ras. Northern blotting analysis showed that TSA prolonged the maximum expression period of c=fos mRNA triggered by NGF and delayed its return to the basal level. TSA reduced c-jun mRNA induction by NGF but greatly enhanced c-myc mRNA induced by NGF. Yoshida et al. (J. Biol. Chem, 265, 17174-17179, 1990) showed that TSA inhibits histone deacetylation, which might influence the gene expression involved in cellular differentiation. In this study, we also found that TSA prevents histone deacetylation in PC12 cells as well as other cell lines, suggesting that inhibition of histone deacetylation by TSA might affect the expression of early-response genes. We also demonstrated that TSA induced reversion of oncogenic ras-transformed NIH3T3 cells to a normal morphology, suggesting that inhibitors of ras-mediated differentiation of PC12 cells may be effective as anticancer agents. PMID: 7700637 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 604: Cell Growth Differ. 1995 Mar;6(3):219-27. Nuclear factor I interferes with transformation induced by nuclear oncogenes. Schuur ER, Kruse U, Iacovoni JS, Vogt PK. Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037, USA. The four nuclear factor I genes (NFI-A, NFI-B, NFI-C, and NFI-X) give rise to multiple isoforms by alternative splicing in many tissues. These NFI proteins cooperate with AP-1, Myc, and other transcription factors in regulating transcription of numerous cellular and viral genes. We have investigated the growth-regulatory potential of NFI by overexpressing cDNAs from chicken NFI genes -A, -B, -C, and -X in chicken embryo fibroblasts (CEF). None of the NFI cDNAs induced oncogenic transformation of CEF. However, overexpression of each of the NFI proteins caused similar morphological alteration of the cells, inducing them to become flattened and polygonal and to show increased adherence. The growth properties of these cells were similar to normal CEF. When these morphologically altered CEF were challenged by superinfection with oncogenic retroviruses, they were resistant to transformation by the nuclear oncogenes jun, fos, junD, myc, and qin but were readily transformed by cytoplasmic oncogenes src, mil/raf, ras, and fps. The NFI-A1 protein was able to alter transactivation by the cellular and viral Jun proteins in a promoter-dependent manner. The changes in cell morphology and reduced susceptibility to nuclear oncogenes were not seen with a carboxy-terminal truncation in the transactivation domain of NFI, suggesting that this region of the protein is essential for the observed effects. The dichotomy between the activities of nuclear and of cytoplasmic oncogenes in this system is discussed. PMID: 7794790 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 605: Kidney Int. 1995 Mar;47(3):782-8. Expression of growth-related protooncogenes during diabetic renal hypertrophy. Shankland SJ, Scholey JW. Department of Medicine, University of Toronto, Ontario, Canada. Experimental type 1 diabetes mellitus is characterized by an early increase in kidney weight and glomerular volume, but changes in gene expression accompanying diabetic renal growth have not been elucidated. The early response genes, c-fos, c-jun, and c-myc encode proteins that regulate gene transcription, thus influencing the cellular responses to a stimulus. Accordingly, we studied c-fos, c-jun, and c-myc expression in glomeruli during the rapid phase of glomerular hypertrophy that follows the onset of hyperglycemia in diabetic rats. Total RNA was extracted by the method of Chomczynski from isolated glomeruli of streptozotocin (60 mg/kg i.v.) induced diabetic rats 24, 48, and 96 hours, and 1 week after the onset of hyperglycemia (blood glucose > 15 mmol/liter). A second group of rats, studied after streptozotocin administration, received twice daily insulin to maintain normoglycemia. A group of age-matched normal rats served as the control group. Northern blot analysis was performed with cDNA probes for c-fos, c-jun, and c-myc, and GAPDH. mRNA levels for c-fos increased fourfold 24 hours after the onset of hyperglycemia, but returned to baseline by 48 hours. mRNA levels for c-jun increased threefold 24 hours after the onset of hyperglycemia, in diabetic glomeruli, and the increase was sustained for one week. Intensive insulin treatment normalized blood glucose levels and abrogated the increases in c-fos and c-jun expression. There was no discernable increase in c-myc mRNA levels in the diabetic glomeruli.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 7752577 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 606: DNA Cell Biol. 1995 Feb;14(2):95-101. Activation of the nuclear oncogenes N-myc and c-jun in carcinoid [correction of cartinoid] tumors of transgenic mice carrying the human adenovirus type 12 E1 region gene. Sagara M, Sugiyama F, Horiguchi H, Kamma H, Ogata T, Yagami K, Murakami K, Fukamizu A. Institute of Applied Biochemistry, University of Tsukuba, Ibaraki, Japan. The adenovirus (Ad) E1 region genes, E1A and E1B, are well known cooperatively to transform primary rodent cells and activate a number of cellular promoters, including nuclear oncogenes such as N-myc and c-jun, in transfected cell lines. However, there is still less information available on the in vivo mechanism(s) by which the E1 region gene, when chromosomally integrated in the living animals, exerts its effect on nuclear oncogene activation coupled with transformation. To investigate such in vivo activity of E1A we have used a series of microinjection experiments into fertilized eggs to generate three transgenic mice carrying the Ad12-type E1A/E1B genes under the control of the human renin gene. This transgene caused an early onset of bowel cartinoid tumors that express neural cell adhesion molecules, but do not metastasize to any region. Northern blot analysis revealed that the transgenes were considerably expressed in the tumors, but not in other tissues at detectable levels. Interestingly, the levels of N-myc and c-jun mRNAs in the cartinoid tumors were elevated 19- and 8-fold, respectively, as compared with those found in the control intestine. In contrast, the major histocompatibility complex (MHC) class I mRNA level was not altered between the tumor and control intestines, suggesting that this unchanged expression may reflect the loss of tumor metastasis. These findings provide the first in vivo evidence that the expression of the Ad12 E1 region gene induces cartinoid tumors associated with the activation of the nuclear oncogenes N-myc and c-jun. PMID: 7865136 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 607: EMBO J. 1995 Feb 1;14(3):452-60. Glucocorticoid-induced apoptosis of human leukemic cells is caused by the repressive function of the glucocorticoid receptor. Helmberg A, Auphan N, Caelles C, Karin M. Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla 92093-0636. Induction of apoptosis in lymphocytes, which may account for the therapeutic effects of glucocorticoids in various diseases including leukemia, depends on the glucocorticoid receptor. However, the events leading from the activated receptor to cell lysis are not understood. A prevailing hypothesis postulates induction of so-called 'lysis genes' by the activated receptor. In this study, we show that an activation-deficient glucocorticoid receptor mutant is as effective as the wild-type receptor in repression of AP-1 activity, inhibition of interleukin-2 production, inhibition of c-myc expression and induction of apoptosis. Furthermore, we show that retinoic acid can also induce apoptosis in these cells through the retinoic acid receptor, whose repressive functions but not target site specificity, are similar to those of the glucocorticoid receptor. Therefore, the primary effect of the receptor in glucocorticoid-mediated apoptosis correlates with transcriptional repression rather than activation and could be mediated by interference with other transcription factors required for cell survival. PMID: 7859735 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 608: J Cell Physiol. 1995 Feb;162(2):234-45. Regulation of human ornithine decarboxylase expression following prolonged quiescence: role for the c-Myc/Max protein complex. Pena A, Wu S, Hickok NJ, Soprano DR, Soprano KJ. Department of Microbiology and Immunology, Temple University School of Medicine, Philadelphia, Pennsylvania 19140. WI-38 cells can remain quiescent for long periods of time and still be induced to reenter the cell cycle by the addition of fresh serum. However, the longer these cells remain growth arrested, the more time they require to enter S phase. This prolongation of the prereplicative phase has been localized to a point early in G1, after the induction of "immediate early" G1 genes such as c-fos and c-jun but before maximal expression of "early" G1 genes such as ornithine decarboxylase (ODC). Understanding the molecular basis for ODC mRNA induction can therefore provide information about the molecular events which regulate the progression of cells out of long-term quiescence into G1 and subsequently into DNA synthesis. Studies utilizing electrophoretic mobility shift assays (EMSA) of nuclear extracts from short- and long-term quiescent WI-38 cells identified a region of the human ODC promoter at -491 bp to -474 bp which exhibited a protein binding pattern that correlated with the temporal pattern of ODC mRNA expression. The presence of a CACGTG element within this fragment, studies with antibodies against c-Myc and Max, the use of purified recombinant c-Myc protein in the mobility shift assay, and antisense studies suggest that these proteins can specifically bind this portion of the human ODC promoter in a manner consistent with growth-associated modulation of the expression of ODC and other early G1 genes following prolonged quiescence. These studies suggest a role for the c-Myc/Max protein complex in regulating events involved in the progression of cells out of long-term quiescence into G1 and subsequently into S. PMID: 7822433 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 609: EMBO J. 1995 Feb 1;14(3):473-83. Differential roles of PI3-kinase and Kit tyrosine 821 in Kit receptor-mediated proliferation, survival and cell adhesion in mast cells. Serve H, Yee NS, Stella G, Sepp-Lorenzino L, Tan JC, Besmer P. Molecular Biology Program, Sloan Kettering Institute, New York, NY 10021. The pleiotropic effects of the Kit receptor system are mediated by Kit-Ligand (KL) induced receptor autophosphorylation and its association with and activation of distinct second messengers, including phosphatidylinositol 3'-kinase (PI3-kinase), p21ras and mitogen-activated protein kinase (MAPK). To define the role of PI3-kinase, p21ras and MAPK in Kit-mediated cell proliferation, survival and adhesion in bone marrow-derived mast cells (BMMC), mutant Kit receptors were expressed in Wsh/Wsh BMMC lacking endogenous c-kit expression. The introduction of both murine Kit(S) and KitL (isoform containing a four amino acid insert) into Wsh/Wsh BMMC restored KL-induced proliferation, survival and adhesion to fibronectin, as well as activation of PI3-kinase, p21ras and MAPK, and induced expression of c-fos, junB, c-myc and c-myb mRNA. Substitution of tyrosine 719 in the kinase insert with phenylalanine (Y719F) abolished PI3-kinase activation, diminished c-fos and junB induction, and impaired KL-induced adhesion of BMMC to fibronectin. In addition, the Y719F mutation had partial effects on p21ras activation, cell proliferation and survival, while MAP kinase activation was not affected. On the other hand, Y821F substitution impaired proliferation and survival without affecting PI3-kinase, p21ras and MAPK activation, and induction of c-myc, c-myb, c-fos and c-jun mRNA, while KL-induced cell adhesion to fibronectin remained intact. In agreement with a role for PI3-kinase in Kit-mediated cell adhesion, wortmannin blocked Kit-mediated cell adhesion at concentrations known to specifically inhibit PI3-kinase. We conclude, that association of Kit with p85PI3-K, and thus with PI3-kinase activity, is necessary for a full mitogenic as well as adhesive response in mast cells. In contrast, tyrosine 821 is essential for Kit-mediated mitogenesis and survival, but not cell adhesion. PMID: 7532131 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 610: Biochim Biophys Acta. 1995 Jan 25;1270(1):12-8. Relationships between proto-oncogene expression and apoptosis induced by anticancer drugs in human prostate tumor cells. Sinha BK, Yamazaki H, Eliot HM, Schneider E, Borner MM, O'Connor PM. Biochemical and Molecular Pharmacology Section, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892. A variant of human prostate PC3 cells, isolated from PC3 cells, was shown to be significantly resistant (> 10-fold) to several clinically active anticancer drugs, including VP-16 and cisplatin. Previous studies showed that resistance to these drugs was not due to expression of the mdr1 gene, or modifications in topoisomerases but may have resulted from high expressions of certain proto-oncogenes (Yamazaki et al. (1994) Biochim. Biophys. Acta 1226, 89-96). Flow cytometry, DNA gel electrophoresis and northern blot analysis were used to further characterize drug responses in sensitive and resistant cells. Treatment of the sensitive PC3 cells with VP-16 and CDDP resulted in accumulation of cells in S and G2, and G1 and S phases, respectively, and caused significant degradation of the genomic DNA into internucleosomal sized DNA fragments, indicating apoptosis. In contrast, resistant PC3 cells showed little or no DNA fragmentation. Resistant PC3(R) cells expressed 2-3-fold more bcl2 protein than the parental PC3 cells, and overexpressed c-myc, c-jun and H-ras mRNA compared to sensitive cells. Treatment with VP-16 or CDDP significantly induced c-myc mRNA levels in sensitive PC3 cells. H-ras message was not affected by either VP-16 or CDDP treatment in PC3 cells. These studies, taken together, suggest that a differential susceptibility to apoptosis and chemosensitivity may be related to altered levels of bcl2 and/or oncogene overexpression in PC3(R) cells. PMID: 7827130 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 611: J Natl Cancer Inst. 1995 Jan 18;87(2):117-22. Apoptosis in human adenocarcinoma HT29 cells induced by exposure to hypoxia. Yao KS, Clayton M, O'Dwyer PJ. Fox Chase Cancer Center, Philadelphia, PA 19111, USA. BACKGROUND: Recent evidence indicates that programmed cell death or apoptosis may be an important mechanism for the lethality of cancer chemotherapy drugs in tumor cells. Apoptosis may be induced in tumor cells by a number of physical stresses, including radiation, UV light, heat shock, and cold. In preliminary studies, we have found that exposure of cells to hypoxia rapidly induces the expression of several immediate early genes, including c-jun, jun-D, and c-fos, and of a bifunctional redox protein/endonuclease, Ref-1. PURPOSE: Our purpose was to determine if hypoxia-induced overexpression of the endonuclease was associated with altered DNA integrity and manifestations of apoptosis. METHODS: We examined cultured cells for the induction of apoptosis by electrophoresis and immunofluorescence techniques and related these findings to the induction of gene expression by hypoxia. RESULTS: We found that an 8-hour exposure of human adenocarcinoma HT29 cells to hypoxia (which results in only 14% loss of viability) induced internucleosomal DNA fragmentation beginning after 8 hours of hypoxia and before reoxygenation. Immunofluorescence of hypoxia-treated cells showed that apoptotic cells were detectable initially after 4 hours of hypoxia and reached a peak (31.3% of cells) at 12 hours after reoxygenation and that by 36 hours after reoxygenation all apoptotic cells had been eliminated from the population. Hypoxia-induced apoptosis was associated with a marked induction of c-myc messenger RNA (mRNA). We have previously shown that hypoxia is associated with ref-1 mRNA induction. While expression of c-myc declined with a time course similar to that of the disappearance of apoptotic cells, that of ref-1 remained elevated. Coincident with the decline of c-myc, we observed a late increase in the expression of bcl-2 beginning 24 hours after reoxygenation. CONCLUSIONS AND IMPLICATIONS: These results suggest a possible relationship between Ref-1 induction and the occurrence of apoptosis following a hypoxic exposure. PMID: 7707382 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 612: J Biol Chem. 1995 Jan 6;270(1):78-86. Two closely related isoforms of protein kinase C produce reciprocal effects on the growth of rat fibroblasts. Possible molecular mechanisms. Borner C, Ueffing M, Jaken S, Parker PJ, Weinstein IB. Columbia-Presbyterian Center, Columbia University, New York, New York 10032. We have previously reported that two closely related protein kinase C (PKC) isoforms, PKC alpha and PKC beta I, had divergent effects on the growth and transformation of the same parental R6 rat embryo fibroblast cell line (Housey, G. M., Johnson, M. D., Hsiao, W.-L. W. O'Brian, C. A., Murphey, J. P., Kirschmeier, P., and Weinstein, I. B. (1988) Cell 52, 343-354; Borner, C., Filipuzzi, I., Weinstein, I. B., and Imber, R. (1991) Nature 353, 78-80). Whereas cells that overexpress PKC beta I lost anchorage dependence, grew to higher saturation densities, and generated small tumors when injected into nude mice, none of these properties were seen with cells that overexpress PKC alpha. In fact, the latter cells grew even slower and to lower saturation densities as compared to control cells. Here we investigate possible molecular mechanisms underlying the reciprocal effects of PKC alpha and PKC beta I. Overexpression of both isoforms enhanced 12-O-tetradecanoyl phorbol-13 acetate-induced expression of the growth regulatory genes c-jun, c-myc, and collagenase and enhanced feedback inhibition of epidermal growth factor receptor binding and cellular levels of diacylglycerol. However, the cells overexpressing PKC beta I differed from those overexpressing PKC alpha by displaying a decreased requirement for growth factors and by the production of a mitogenic factor. Thus, the basis for enhanced growth and transformation of cells overexpressing PKC beta I may be the establishment of an autocrine growth factor loop. These findings may be relevant to the roles of specific isoforms of PKC in carcinogenesis and tumor growth. PMID: 7814423 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 613: Ciba Found Symp. 1995;191:254-65; discussion 265-8. Oestrogen- and anti-oestrogen-regulated genes in human breast cancer. Rochefort H. INSERM U148, Unite Hormones et Cancer, Universite de Montpellier, Faculte de Medecine, France. The study of several human breast cancer cell lines containing oestrogen receptors has allowed characterization of a number of oestrogen-induced proteins (e.g. progesterone receptor, cathepsin D, pS2, Hsp27, c-Myc). In primary tumours these markers have different prognostic significance for predicting whether the tumour will be hormone responsive (e.g. pS2, progesterone receptor) and whether it will metastasize (e.g. cathepsin D). The mechanism of regulation of gene expression by oestrogens and anti-oestrogens in breast cancer is complex and varies according to the nature of both the gene and the cell in which it is transcribed. Our laboratory has identified the sequences mediating oestrogen activity in the proximal region of cathepsin D, including a non-consensus oestrogen-responsive element located at -260 which acts in synergy with other regulatory elements. In addition to the classical effect of oestrogen receptor in stimulating transcription of genes controlled by the oestrogen-responsive element, we found that estrogen receptor is able to modulate transcription of AP-1-responsive genes without interacting directly with DNA. Cross-talk between oestrogen receptor and members of the Fos/Jun family via protein-protein interactions may explain how anti-oestrogens inhibit the mitogenic effect of growth factors in the apparent absence of oestrogens and why tamoxifen is able to stimulate cathepsin D gene expression and induce apoptosis in certain oestrogen receptor-positive breast cancer cells. The nature and degree of this cross-talk appears to vary according to the gene, the cell type and the type of oestrogen receptor ligand involved. Studies of oestrogen-regulated genes are not only useful for classifying breast cancers according to their ability to metastasize and respond to therapies, but also should lead to new therapeutic approaches for hormone-dependent and hormone-resistant cancers. Publication Types: Review Review, Tutorial PMID: 8582202 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 614: Cancer Chemother Pharmacol. 1995;35(5):423-31. Molecular characterization of the in vivo alkylating agent resistant murine EMT-6 mammary carcinoma tumors. Chatterjee D, Liu CJ, Northey D, Teicher BA. Dana-Farber Cancer Institute, Joint Center for Radiation Therapy, Boston, MA 02115. The expression of several early-response genes and genes associated with malignant disease was assessed in the EMT-6/parent tumor and the EMT-6/CTX and EMT-6/CDDP in vivo resistant tumor lines growing as tumors or as monolayers in culture. In the absence of treatment the levels of mRNA for the genes c-jun, c-fos, c-myc, Ha-ras and p53 were increased in the EMT-6/CTX and EMT-6/CDDP as compared with the EMT-6/parent tumor, whereas the expression of erb-2 was similar in all three tumors. Although the cells from each of the three tumors show increased expression of early response genes after exposure to cisplatin (CDDP; 100 microM, 2 h) or 4-Hydroxyperoxycyclophosphamide (4-HC; 100 microM, 2 h) in culture, in mRNA extracted from tumor tissue these changes are absent or very small. Both C-jun and erb-2 were detectable in liver. There was increased expression of both of these genes in the livers of tumor-bearing animals as compared with non-tumor-bearing animals. The highest expression of both c-jun and erb-2 occurred in the livers of animals bearing the EMT-6/CDDP tumor. Treatment of the animals with CDDP or cyclophosphamide, in general, resulted in increased expression of both genes at 6 h post treatment. The increased expression of these genes may impart metabolic changes in the tumors and/or hosts that contribute to the resistance of these tumors to specific antitumor alkylating agents. PMID: 7850925 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 615: Mol Carcinog. 1995 Jan;12(1):14-22. Expression of epidermal ornithine decarboxylase and nuclear proto-oncogenes in phorbol ester tumor promotion-sensitive and -resistant mice. Kennard MD, Kang DC, Montgomery RL, Butler AP. University of Texas M. D. Anderson Cancer Center, Smithville 78957. This study was undertaken to assess the effects of a single or two sequential topical applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the expression of c-fos, c-jun, junB, c-myc, and ornithine decarboxylase (ODC) in promotion-sensitive SSIN mice and the relatively promotion-resistant C57BL/6 strain. Northern blot analysis demonstrated that a single promoting dose of TPA induced ODC mRNA expression 10- to 15-fold in both strains. Treatment of each strain with a second dose of TPA, 48 h (in C57BL/6 mice) or 72 h (in SSIN mice) after the first, led to hyperinduction of ODC activity. Although this involved transcription of new ODC mRNA, the hyperinduction of ODC enzyme activity was primarily posttranscriptional. Induction of c-fos mRNA or protein was maximal about 3 h after a single treatment in either strain but was sustained for at least 6 h in C57BL/6 mice. In contrast, two treatments of SSIN mice with TPA caused a rapid, strong c-fos induction 1-2 h after treatment, whereas C57BL/6 mice responded no more strongly than after a single treatment. c-jun mRNA and protein were induced only slightly in either strain, but junB was induced about fivefold in SSIN mice and tenfold in C57BL/6 mice. Although c-myc was induced to comparable levels in both strains, the response was more prolonged in C57BL/6 mice. Compared with SSIN mice, C57BL/6 mice responded to TPA treatment, in general, with changes in proto-oncogene mRNA to a higher level or for longer or both. Thus, although small differences in the expression of these genes were observed, they were not positively correlated with the differential sensitivity of SSIN and C57BL/6 mice toward tumor promotion by phorbol esters, with the possible exception of c-fos. PMID: 7818761 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 616: J Invest Dermatol. 1995 Jan;104(1):78-82. Changes in expression of apoptosis-associated genes in skin mark early catagen. Seiberg M, Marthinuss J, Stenn KS. Skin Biology Research Center, R.W. Johnson Pharmaceutical Research Institute, Raritan, NJ 08869-0602. Programmed cell death is central to hair biology, as the hair follicle undergoes cycles of growth (anagen), regression (catagen), and rest (telogen). During catagen, the hair follicle shortens via a pathway of programmed cell death and apoptosis. The molecular mechanisms involved in this process have not been elucidated yet. Using reverse transcriptase-polymerase chain reaction, we examined in this study the expression in total skin, throughout one hair cycle, of a series of regulatory genes associated with apoptosis. We show that gene expression within skin is hair-cycle-dependent. Transforming growth factor-beta was expressed immediately before catagen; therefore, it might be involved in the early signaling of this process. Tumor necrosis factor-beta was expressed during catagen and might be involved in follicular apoptosis. Several proto-oncogenes and transcription factors have been described in the regulation of apoptosis in other systems. Here we show that the transcript levels of c-myc, c-myb, and c-jun changed immediately before or during early catagen and thus could be involved in the signaling or regulation of catagen. Levels of p53 remained constant throughout anagen and catagen, suggesting that p53 is not involved in the developmentally induced apoptosis of the hair follicle. The variable expression throughout the hair cycle of the genes described demonstrates the dynamic changes of the skin and underscores the importance of studying the complete hair cycle when characterizing any molecule in skin. PMID: 7798646 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 617: J Mol Cell Cardiol. 1995 Jan;27(1):133-40. Molecular mechanism of cardiac cellular hypertrophy by mechanical stress. Yamazaki T, Komuro I, Yazaki Y. Department of Medicine III, University of Tokyo School of Medicine, Japan. Mechanical stress is a major cause of cardiac hypertrophy. Although the mechanisms by which mechanical load induces cardiac cellular hypertrophy have long been a subject of great interest for cardiologists, the lack of a good in vitro system has hampered the understanding of the biochemical mechanisms. For these past several years, however, an in vitro cardiocyte culture system has made it possible to examine the biochemical basis for the signal transduction of mechanical stress. Passive stretch of cardiomyocytes cultured on silicone membranes activates protein kinase cascades of phosphorylation and induces an increase in protein synthesis and the expression of both immediate early genes such as c-fos, c-myc, c-jun, Egr-1, and late response genes such as beta-myosin heavy chain and skeletal alpha-actin. Although an important question regarding how mechanical stimulus is converted into biochemical signals remains unknown, the cultured cardiomyocyte is a good model to examine the signal transduction pathways of mechanical stress. Publication Types: Review PMID: 7760338 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 618: J Hypertens. 1995 Jan;13(1):105-12. Angiotensin converting enzyme inhibition prevents proto-oncogene expression in the vascular wall after injury. Van Belle E, Bauters C, Wernert N, Delcayre C, McFadden EP, Dupuis B, Lablanche JM, Bertrand ME, Swynghedauw B. Department of Cardiology, University of Lille, France. OBJECTIVES: Angiotensin converting enzyme (ACE) inhibitors reduce neointimal hyperplasia after balloon denudation, but the mechanisms are not completely understood. It has been demonstrated that nuclear oncogenes are induced in the vascular wall in the hours immediately after injury, and that the same genes are induced by angiotensin II in vascular smooth muscle cells. It has therefore been suggested that the effects of ACE inhibitors on the response of the vessel wall could be mediated by an inhibition of proto-oncogene expression. METHODS AND RESULTS: Sixteen New Zealand White rabbits were randomly assigned for histologic analysis to receive placebo (n = 9) or 1 mg/kg per day perindopril (n = 7). After treatment for 7 days balloon aortic injury was performed. The treatment was continued and the rabbits were killed 28 days after injury. In the perindopril group the neointimal cross-sectional area was significantly smaller than in the control group. Six untreated rabbits were used to assess the time course of proto-oncogene expression in the aortic wall after injury in the present model. After extraction, total aortic RNA was hybridized with myc, fos and jun probes. Based on the results, the effects of ACE inhibition on proto-oncogene expression were tested 1 h after balloon denudation. Accordingly, 24 rabbits were randomly assigned to pretreatment for 7 days with placebo or with 1 or 10 mg/kg per day perindopril (n = 8, for each group) and were killed 1 h after injury. Expression of c-myc was not altered by pretreatment. However, 1 mg/kg per day perindopril induced significant reductions of 50% in c-jun and 45% in c-fos expression compared with control. No additional effect was obtained with the higher dose. CONCLUSION: The effect of ACE inhibition on intimal hyperplasia is associated with a reduction in early cellular events such as c-fos and c-jun expression. These results suggest that potent ACE inhibition at the time of vascular injury may be required to limit the hyperplastic response of the vessel wall. PMID: 7759840 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 619: Virus Genes. 1995 Jan;9(2):161-70. Augmentation of c-fos and c-jun expression in transgenic mice carrying the human T-cell leukemia virus type-I tax gene. Iwakura Y, Tosu M, Yoshida E, Saijo S, Nakayama-Yamada J, Itagaki K, Asano M, Siomi H, Hatanaka M, Takeda T, et al. Institute of Medical Science, University of Tokyo, Japan. To analyze the effect of human T-cell leukemia virus type I (HTLV-I) on cellular gene expression and its relation to tumorigenesis, two lines of transgenic mice carrying the long terminal repeat (LTR)-env-pX-LTR regions of the HTLV-I genome were produced. The transgene was expressed in many organs, including the brain, salivary gland, spleen, thymus, skin, muscle, and mammary gland. We found that the expression of the c-fos and c-jun genes, but not of the lyn and c-myc genes, was augmented 2- to 20-fold in histologically normal skin and muscle of these mice. The augmentation was tissue specific, suggesting the involvement of a cellular factor in the transgene action. In these mice, a three to seven times higher incidence of tumors was seen as compared with the control mice. These tumors included mesenchymal tumors, such as fibrosarcoma, neurofibroma, and lipoma, and adenocarcinomas of the mammary gland, salivary gland, and lung. The c-fos and c-jun genes were also activated in these tumors. The possible roles of elevated c-fos and c-jun gene expression in tumorigensis are discussed. PMID: 7732661 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 620: Vestn Ross Akad Med Nauk. 1995;(3):53-7. [Psoriasis and the problem of gene expression regulation] [Article in Russian] Vasileiskii SS. During the epidermis maturation different keratin genes are expressed in strictly definite sequences according to the level of epidermis. In psoriasis this sequence in deranged (permutation) Perhaps this is due to implication of homeobox system similar to what occurs in (Ftz) gene. This problem is reviewed and discussed in this paper. All keratin as well as involukrine genes have been sequenced. Regions of K5 gene have been found which play regulatory role in transcription. It has been achieved using transfection K5 into keratinocytes. Psoriatic injury reversed by yEPP peptide of chick Y. sack cells. It is well established that protooncogenes are involved in transcription regulation. Expression of c-myc is increased in psoriatic plagues whereas that of c-fos and c-jun is decreased. This fact may be regarded as an indirect evidence for abnormal functioning c-myc in regulation. It is not ruled out that the system implicated in shedding in reptiles recapitulate in psoriasis. PMID: 7540450 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 621: Ann N Y Acad Sci. 1994 Dec 15;747:172-82. Transcription factors as molecular mediators in cell death. Soares HD, Curran T, Morgan JI. Roche Institute of Molecular Biology, Roche Research Center, Nutley, New Jersey 07110. Publication Types: Review PMID: 7531406 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 622: Oncogene. 1994 Dec;9(12):3493-8. Transformation by Raf and other oncogenes renders cells differentially sensitive to growth inhibition by a dominant negative c-jun mutant. Rapp UR, Troppmair J, Beck T, Birrer MJ. Laboratory of Viral Carcinogenesis, National Cancer Institute, Frederick Cancer Research and Development Center, Maryland 21702-1201. In NIH3T3 cells expressing active Raf-1 protein serine/threonine kinase (PSK) c-jun expression is constitutive while c-fos expression is attenuated. This alteration prompted us to determine whether oncogene transformation would render cells differentially sensitive to growth inhibition by a dominant negative mutant of c-jun, TAM 67. Growth inhibition was observed in three types of assays: (1) transfection of TAM 67 into cells stably transformed by a variety of oncogenes, (2) cotransfection of TAM 67 with oncogene expression plasmids into NIH3T3 cells and (3) titration of oncogene-expressing retroviruses on cells stably expressing TAM 67. The results clearly demonstrate that Raf-1 dependent oncogenes, which include receptor protein tyrosine kinases (PTKs)-, intracellular PTKs- and Ras-derived genes share the Raf phenotype of constitutive c-jun expression, attenuated c-fos induction, and high sensitivity to growth suppression by TAM 67. Additionally, the intracellular PSK oncogene, mos and the nuclear oncogenes c-myc, c-fos, and SV40 T antigen were TAM 67-sensitive for transformation. This universal pattern of altered growth regulation in oncogene transformed fibroblast cell lines highlights the potential usefulness of c-jun based inhibitors for control of tumor cell growth. PMID: 7970709 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 623: Pathologe. 1994 Dec;15(6):321-30. [Oncogenes and tumor suppressor genes in the pathogenesis of lung tumors] [Article in German] Wiethege T, Voss B, Muller KM. Institut fur Pathologie, Berufsgenossenschaftliche Kliniken Bergmannsheil, Bochum. The recent results obtained from investigations based on molecular biological techniques have led to a better understanding of recurrent genetic causes important for the pathogenesis of tumors. Several genes have been identified as being involved in the development of cancer. In many cases, the activation of oncogenes or the inactivation of tumor-suppressor genes is the predominant reason for cancerogenic cell transformation. Functional dysregulation is frequently the consequence of mutations, resulting in an alteration of the primary structure of the DNA. As our understanding of the nature, function, and interaction of these genes evolves, new opportunities for early diagnosis, classification, prevention, and treatment of malignant tumors will arise. The present report summarizes the current molecular biological aspects of several oncogenes (erbB, ras, myc, raf, fos, jun, bcl, mdm2, myb, kit CSF1R, met) and tumor suppressor genes (p53, rb, mts) involved in lung-cancer development with respect to the pathology of lung tumors, including the importance of these genes as far as the clinical course of the disease is concerned. Publication Types: Review Review, Tutorial PMID: 7855100 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 624: Bull Cancer. 1994 Dec;81 Suppl 2:105s-111s. [Drug resistance, oncogenes, and anti-oncogenes in epithelial tumors] [Article in French] Riva-Lavieille C. Institut Albert-Bonniot, Universite Joseph-Fourier, Grenoble, France. Twenty four squamous cell carcinomas of the head and neck (HNSCC) of stage II to IV were evaluated for the expression of potential markers such as oncogenes and tumor suppressor genes in drug-resistance behavior. We have analysed the c-myc, c-jun, c-raf and N-ras and p53 expression in total RNA preparation from tumor biopsies obtained before treatment. The patients underwent chemotherapy including 5-fluorouracil and cisplatinum. No significant differences in c-raf and N-ras expression were found in responding or resistant patients. However, resistance to chemotherapy was associated with low expression of c-myc (P < 0.025) or high expression of c-jun (P < 0.001). In addition, p53 mRNA pre-therapeutic level was increased in unresponsive patients to chemotherapy (P < 0.05). Therefore, analysis of the expression of c-myc, c-jun oncogenes and p53 tumor suppressor gene in tumor cells before initiation of therapy may define a subset of patients with potentially better prognosis. PMID: 7727855 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 625: Cancer Lett. 1994 Nov 25;87(1):85-9. Curcumin inhibits TPA induced expression of c-fos, c-jun and c-myc proto-oncogenes messenger RNAs in mouse skin. Kakar SS, Roy D. Department of Physiology and Biophysics, University of Alabama at Birmingham 35294. Curcumin is a major chemical constituent of turmeric normally eaten by humans. 12-O-Tetradecanoyl phorbol-13-acetate (TPA) is a strong promoter of chemically induced skin cancer. The exact mechanism by which TPA promotes skin cancer is not clear. However, it is known that TPA elevates the expression of oncogenes involved in cell proliferation. Recently, it has been shown that turmeric or curcumin significantly inhibits TPA-induced tumor promotion on mouse skin. However, the mechanism by which curcumin inhibits TPA-induced tumor promotion is not known. In the present studies, we investigated the effect of curcumin on the expression of c-fos, c-jun and c-myc oncogenes in TPA-treated mouse skin in CD-1 mice. A 30-nmol dose of TPA increased the levels of mRNAs for c-fos, c-jun and c-myc oncogenes by 2-3-fold compared with control. Topical application on the dorsal side of the skin with 1 mumol, 10 mumol, 20 mumol or 30 mumol of curcumin 30 min before TPA treatment inhibited the TPA-induced expression of these proto-oncogenes. Inhibition of expression of c-fos and c-jun was more pronounced than that of c-myc. A dose of 10 mumol of curcumin was found to inhibit 90% TPA-induced expression of c-fos and c-jun, and 60% of c-myc. These data strongly suggest that curcumin may inhibit skin cancer through the modulation of expression of these proto-oncogenes. PMID: 7954373 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 626: J Biol Chem. 1994 Nov 4;269(44):27756-61. Characterization of conventional protein kinase C (PKC) isotype expression during F9 teratocarcinoma differentiation. Overexpression of PKC alpha alters the expression of some differentiation-dependent genes. Kindregan HC, Rosenbaum SE, Ohno S, Niles RM. Department of Biochemistry, Boston University School of Medicine, Massachusetts 02118. F9 teratocarcinoma is a useful model for studying early embryogenesis since these cells can differentiate into primitive or parietal endoderm under the influence of retinoic acid or retinoic acid and cyclic AMP, respectively. We have found that three isoforms of protein kinase C (PKC alpha, -beta, and -gamma) were expressed in undifferentiated stem cells. When the cells were treated with retinoic acid either alone or in the presence of cAMP for 120 h, PKC alpha mRNA and protein levels increased, whereas those of PKC beta and PKC gamma became undetectable. These changes began within 24 h of drug treatment and were complete by 48-72 h. In order to determine the functional significance of the induction of PKC alpha during F9 differentiation, we established two stable transfectants that overexpressed PKC alpha protein between 4- and 5-fold compared to wild type cells. Characterization of these cell lines revealed an altered pattern of expression of some of the markers of F9 differentiation. The clone that had the highest amount of PKC alpha protein constitutively expressed mRNA for type IV collagen and c-Jun, which are not normally expressed until 24-48 h of treatment with differentiation agents. In the other overexpressing clone, these markers were induced much faster than in wild type cells. The growth rate of both overexpressing clones was less than wild type cells, while the expression of the PKC beta protein in these clones was similar to the levels found in differentiated F9 cells. However, other markers of differentiation, including the cellular morphology and levels of pST6-135 and c-myc RNA, responded to agents identically in both wild type and PKC-alpha-overexpressing clones. Therefore, overexpression of PKC alpha is not sufficient to induce full differentiation of F9 cells. However, our data suggest that certain pathways that lead to the expression of differentiation-dependent genes are regulated by PKC alpha protein levels. PMID: 7961696 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 627: Am J Physiol. 1994 Nov;267(5 Pt 2):F805-15. Anti-AP-1 activity of all-trans retinoic acid in glomerular mesangial cells. Simonson MS. Department of Medicine, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106. Functional antagonism between retinoic acid (RA) receptors and activator protein-1 (AP-1) transcription factors might regulate expression of genes involved in the response to injury in the kidney. We designed experiments to analyze the mechanisms by which RA inhibits AP-1-directed transcriptional responses in glomerular mesangial cells. RA inhibited serum-stimulated mesangial cell proliferation as assessed by measurements of [3H]thymidine uptake and cell number. In transient transfection assays with a chloramphenicol acetyltransferase reporter, RA completely blocked transcription directed by an AP-1 cis-element in cells stimulated by serum. AP-1 DNA binding was analyzed in electrophoretic gel mobility shift assays using nuclear extracts from control or RA-pretreated cells stimulated with serum. RA did not abolish AP-1 DNA binding activity under the conditions of this assay. The apparent equilibrium dissociation constant, maximal density of binding, and association rate for the AP-1-DNA interaction were similar in serum-stimulated cells or RA-pretreated cells stimulated with serum. RA repressed serum-stimulated induction of the immediate early genes c-fos and c-jun, whose protein products dimerize to form AP-1. Repression was relatively selective for c-fos/c-jun; induction of other immediate early transcription factors (junB, c-myc, and egr-1) was not downregulated by RA. That repression of c-fos by RA might contribute to anti-AP-1 activity was suggested by experiments with an antisense c-fos expression vector, which demonstrated that c-fos induction was required for serum-stimulated AP-1 activity. Together, these data demonstrate that RA antagonizes AP-1-directed transcription without inhibiting AP-1 DNA-binding in mesangial cells. Selective repression of c-fos and c-jun might contribute to the anti-AP-1 activity of RA. PMID: 7977784 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 628: Mol Cell Biol. 1994 Nov;14(11):7404-13. Comment in: Mol Cell Biol. 1995 Aug;15(8):4657-8. A critical cytoplasmic domain of the interleukin-5 (IL-5) receptor alpha chain and its function in IL-5-mediated growth signal transduction. Takaki S, Kanazawa H, Shiiba M, Takatsu K. Department of Immunology, University of Tokyo, Japan. Interleukin-5 (IL-5) regulates the production and function of B cells, eosinophils, and basophils. The IL-5 receptor (IL-5R) consists of two distinct membrane proteins, alpha and beta. The alpha chain (IL-5R alpha) is specific to IL-5. The beta chain is the common beta chain (beta c) of receptors for IL-3 and granulocyte-macrophage colony-stimulating factor (GM-CSF). The cytoplasmic domains of both alpha and beta chains are essential for signal transduction. In this study, we generated cDNAs of IL-5R alpha having various mutations in their cytoplasmic domains and examined the function of these mutants by expressing them in IL-3-dependent FDC-P1 cells. The membrane-proximal proline-rich sequence of the cytoplasmic domain of IL-5R alpha, which is conserved among the alpha chains of IL-5R, IL-3R, and GM-CSF receptor (GM-CSFR), was found to be essential for the IL-5-induced proliferative response, expression of nuclear proto-oncogenes such as c-jun, c-fos, and c-myc, and tyrosine phosphorylation of cellular proteins including JAK2 protein-tyrosine kinase. In addition, analysis using chimeric receptors which consist of the extracellular domain of IL-5R alpha and the cytoplasmic domain of beta c suggested that dimerization of the cytoplasmic domain of beta c may be an important step in activating the IL-5R complex and transducing intracellular growth signals. PMID: 7935454 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 629: Leuk Lymphoma. 1994 Nov;15(5-6):445-51. Combined antileukemic activity of pIXY 321 and Ara-C against human acute myeloid leukemia cells. Tang C, Huang Y, Ponnathpur VS, Ray S, Mahoney ME, Bullock G, Ibrado AM, Bhalla K. Department of Medicine, Medical University of South Carolina, Charleston 29425. Prolonged administration of conventional (100 mg/m2/day) or low dose Ara-C (20 mg/m2/day) has been associated with significant clinical antileukemic effects in AML and myelodysplastic syndromes. These doses and schedules of Ara-C yield plasma Ara-C concentrations in the range of 10 to 100 nM. Utilizing concentrations and a schedule of Ara-C treatment, representative of Ara-C exposures in these clinical situations, we performed in vitro studies to examine the effects of co-treatment with pIXY 321 on Ara-C induced apoptosis and Ara-C-mediated colony growth inhibition of human myeloid leukemia HL-60 cells. Significantly greater internucleosomal DNA fragmentation, higher percentage of morphologically recognizable apoptotic cells and increased colony growth inhibition were observed following treatment with 100 versus 10 nM Ara-C for 5 days. Simultaneous exposure to 10 ng/ml pIXY 321 resulted in significantly increased colony growth inhibition as well as DNA fragmentation and apoptosis due to 10 nM but not 100 nM Ara-C. These concentrations of Ara-C inhibited c-myc and did not induce c-jun mRNA expression. These effects of Ara-C on c-myc and c-jun expressions were not influenced by co-treatment with pIXY 321. Neither treatment with pIXY 321 or Ara-C alone, nor co-treatment with pIXY 321 and Ara-C, significantly altered the intracellular p26BCL-2 levels in HL-60 cells. These results indicate that co-treatment with pIXY 321 significantly increases low dose Ara-C-induced apoptosis and thereby its antileukemic activity. PMID: 7874002 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 630: Arkh Patol. 1994 Nov-Dec;56(6):44-9. [Nuclear oncoprotein expression in lung precancer and cancer at various stages of tumor progression studies at the level of light- and electron-immunohistochemistry] [Article in Russian] Kogan EA, Shtabskii AB, Sekamova SM, Mazurenko NN, Kiselev FL. 68 cases of lung carcinoma, 3 carcinoids and 15 fibrosing alveolitis with foci of adenomatosis and bronchiolo-alveolar carcinoma were studied. Oncoproteins c-fos, c-jun, c-ets-1, c-myc L and L-myc were identified in the tumour and surrounding tissue. Expression of c-fos was revealed in 79 of 138(59.4%) of proliferative and dysplastic changes of lung epithelium; c-jun in 40 of 61 (65.6%), c-ets-1 in 22 of 41 (53.7%), c-myc in 41 of 96(42.7%) and L-myc in 15 of 61 (24.6%), mainly in altered bronchial epithelium with a positive reaction to the antibodies against neuron specific enolase and S100 protein. More pronounced expression of nuclear oncoproteins, heterogeneity of their location in tissues, frequent cytoplasmic location in tumour cells were typical for lung carcinoma. PMID: 7605217 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 631: FEBS Lett. 1994 Oct 17;353(2):133-7. Serum alleviates the requirement of the granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced Ras activation for proliferation of BaF3 cells. Sakamaki K, Yonehara S. Pharmaceutical Basic Research Laboratory, JT Inc., Yokohama, Japan. Deletion analysis of the beta subunit of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor previously defined two cytoplasmic regions required for distinct signaling. The membrane-proximal region is responsible for induction of c-myc and pim-1, and is indispensable for GM-CSF-dependent proliferation of mouse BaF3 transfectants. The distal region is required for activation of Ras, Raf-1, MAP kinase and p70 S6 kinase as well as induction of c-fos and c-jun, but is dispensable for GM-CSF-dependent proliferation of transfectants under normal culture conditions containing serum. Here we show that signals induced by the distal region of the beta subunit are also required for proliferation. GM-CSF supported proliferation of BaF3 transfectants expressing the normal beta subunit, even in serum-free medium. However, in the absence of seru, GM-CSF did not support proliferation of BaF3 transfectants that have the beta deletion mutants lacking the distal region. Serum-induced activation of Ras, phosphorylation of MAP kinase and expression of c-fos in parental BaF3 cells and antisense oligonucleotide against c-raf blocked DNA synthesis of BaF3 cells. These results indicate that proliferation of BaF3 cells requires signals induced by the proximal as well as the distal region of the beta subunit of the GM-CSF receptor, and that serum alleviates the requirement of signals induced by the distal region. PMID: 7926037 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 632: Cancer Res. 1994 Oct 15;54(20):5280-3. Epidermal growth factor-mediated apoptosis of MDA-MB-468 human breast cancer cells. Armstrong DK, Kaufmann SH, Ottaviano YL, Furuya Y, Buckley JA, Isaacs JT, Davidson NE. Johns Hopkins Oncology Center, Johns Hopkins University School of Medicine, Baltimore, Maryland 21287. MDA-MB-468 human breast cancer cells lack estrogen receptors, overexpress epidermal growth factor (EGF) receptors, and are growth inhibited by EGF. We show that treatment of MDA-MB-468 cells with EGF leads to inhibition of cell proliferation, fragmentation of DNA into nucleosomal oligomers, and the development of apoptotic morphology. This treatment is associated with increased expression of c-myc, c-fos, jun family members, and transforming growth factor beta 1 mRNA and with partial proteolytic cleavage of poly(ADP-ribose) polymerase and lamin B. The observation that EGF can mediate apoptosis in EGF receptor-overexpressing cells has important implications for clinical efforts directed at the EGF receptor. PMID: 7923154 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 633: Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):10217-21. A variant epidermal growth factor receptor exhibits altered type alpha transforming growth factor binding and transmembrane signaling. Moriai T, Kobrin MS, Hope C, Speck L, Korc M. Department of Medicine, University of California, Irvine 92717. Epidermal growth factor (EGF) and type alpha transforming growth factor (TGF-alpha) bind to a specific region in subdomain III of the extracellular portion of the EGF receptor (EGFR). Binding leads to receptor dimerization, auto-and transphosphorylation on intracellular tyrosine residues, and activation of signal transduction pathways. We compared the binding and biological actions of EGF and TGF-alpha in Chinese hamster ovary (CHO) cells expressing either wild-type human EGFR (HER497R) or a variant EGFR that has an arginine-to-lysine substitution in the extracellular domain at codon 497 (HER497K) within subdomain IV of EGFR. Both receptors exhibited two orders of binding sites with radioiodinated EGF (125I-EGF). Similar results were obtained with 125I-TGF-alpha in cells expressing HER497R. In contrast, only one order of low-affinity binding sites was seen with 125I-TGF-alpha in the case of HER497K. Although EGF and TGF-alpha enhanced tyrosine phosphorylation of both receptors, CHO cells expressing HER497K exhibited an attenuated growth response to EGF and TGF-alpha and a reduced induction of the protooncogenes FOS, JUN, and MYC. Moreover, high concentrations of TGF-alpha (5 nM) inhibited growth in these cells but not in cells expressing HER497R. These findings indicate that a region in subdomain IV of EGFR regulates signal transduction across the cell membrane and selectively modulates that binding characteristics of TGF-alpha. PMID: 7937865 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 634: Cardiovasc Res. 1994 Oct;28(10):1519-25. Selective changes in natriuretic peptide and early response gene expression in isolated rat atria following stimulation by stretch or endothelin-1. Bruneau BG, de Bold AJ. University of Ottawa Heart Institute, Ottawa Civic Hospital, Ontario, Canada. OBJECTIVE: Important physiological and pathophysiological conditions are associated with changes in secretion and synthesis of atrial natriuretic factor (ANF) and brain natriuretic peptide (BNP). The aim of this study was to examine the effects of mechanical stretch and endothelin-1 on ANF and BNP secretion and gene expression in isolated adult rat atria, as paradigms for mechanical and endocrine stimulation of atrial tissue. The expression of the early response genes c-fos, c-jun, Egr-1, and c-myc was also studied, since their protein products may be involved in controlling natriuretic peptide gene expression. METHODS: Isolated rat atria were stimulated by stretch (5 g) or endothelin-1 (10(-7) M) for 30 min, 2 h, or 4 h. ANF and BNP secretion was measured by radioimmunoassay, and relative mRNA levels were determined by northern blotting. RESULTS: Atrial stretch resulted in an immediate 1.8-fold increase in ANF release, which returned to basal levels after 160 min. Endothelin-1 caused a gradual increase in ANF release, up to 2.3 times basal levels, and thereafter returned towards basal levels. BNP secretion was increased threefold by endothelin-1, and remained significantly raised for 90 min. BNP mRNA levels were transiently increased by 33% after 2 h of endothelin-1 stimulation. Stretch increased c-fos mRNA levels (+55%) and Egr-1 mRNA levels (+70%) after 2 h, and increased c-myc mRNA levels (+69%) after 4 h. Endothelin-1 increased Egr-1 mRNA levels up to +767% after 4 h. CONCLUSIONS: Endothelin-1 stimulates BNP secretion from rat atria; this is followed by an increase in BNP mRNA levels. Conversely, acute secretion of ANF by stretch or endothelin-1 is not accompanied by changes in ANF mRNA levels. Atrial stretch results in changes in the expression of the early response genes c-fos, Egr-1, and c-myc, while endothelin-1 stimulates Egr-1 expression. The specific changes in natriuretic peptide and early response gene expression reveal distinct mechanisms of modulation of atrial gene expression by mechanical and endocrine stimuli. PMID: 8001040 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 635: Exp Cell Res. 1994 Oct;214(2):561-9. Jun B expression is regulated differently by three mitogenic pathways in thyrocytes. Pirson I, Dumont JE. Institute of Interdisciplinary Research, School of Medicine, Free University of Brussels, Belgium. In dog thyrocytes in primary culture, thyrotropin (TSH), acting through cyclic AMP, induces proliferation and differentiation expression, while tetradecanoylphorbol acetate (TPA) or epidermal growth factor (EGF) induces proliferation and dedifferentiation. In this work, we have investigated the regulation of mRNA expression of the protooncogene jun B in these cells. TSH stimulated jun B expression very transiently with biphasic kinetics similar to those obtained for c-myc mRNA accumulation. Forskolin reproduced these effects, suggesting that they are, as other effects of TSH in this system, mediated by cyclic AMP. As shown by nuclear run-on experiments, jun B is regulated by the cAMP pathway at the transcriptional level. As in other cell types, EGF or TPA caused a more sustained increase in mRNA levels. In thyroid slices, in which DNA synthesis appears to be induced by the wounding process, jun B is also induced, suggesting a correlation with the proliferative status of the cell. Interestingly, two jun B mRNAs of 2.1 and 2.3 kb were induced by all the mitogenic pathways. The kinetics of their accumulation were different; i.e., TPA induced the smaller transcript with some delay after the longer one and cycloheximide induced the progressive shortening of the first appearing heavier mRNA. The 2.3-kb messenger has a longer poly(A) tail, and kinetics in the presence of actinomycin D suggested it could represent a precursor form of the 2.1-kb messenger. It is suggested that the specific kinetics of cyclic-AMP-induced accumulation of jun B mRNA could be related to the dual stimulation of differentiation and proliferation by TSH in dog thyrocytes. PMID: 7925650 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 636: Cell Struct Funct. 1994 Oct;19(5):341-8. tsJT16, a cell cycle ts mutant of rat fibroblast defective in early G0/G1 transition, fails to induce G1-cyclin and cdk2 genes after serum stimulation at the nonpermissive temperature. Ueohzono T, Tanida-Miyake E, Kato N, Ide T. Department of Cellular and Molecular Biology, Hiroshima University School of Medicine, Japan. tsJT16 is a cell-cycle temperature-sensitive (ts) mutant derived from rat fibroblasts whose functional defect appears soon after the growth stimulation from G0 phase. In addition to c-fos, c-myc and ornithine decarboxylase gene, 7 primarily inducible genes, c-jun, KC, JE, 2F1, 2A9, egr-1, and egr-2, were further shown to be expressed after serum stimulation at both permissive and nonpermissive temperatures. However, expression of secondarily inducible genes, cyclin D1 and D3 and cdk2, was ts and was cycloheximide sensitive. Expression of cyclin C was not inhibited by cycloheximide but it was ts. Failure in expression of G1 cyclins and Cdk2 is suggested to be a causal event for inability of growth induction of tsJT16 at the nonpermissive temperature. PMID: 7850896 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 637: Cell Growth Differ. 1994 Oct;5(10):1127-35. Functional characterization of two cholecystokinin-B/gastrin receptor isoforms: a preferential splice donor site in the human receptor gene. Ito M, Iwata N, Taniguchi T, Murayama T, Chihara K, Matsui T. Department of Medicine, Kobe University School of Medicine, Japan. The cholecystokinin-B and gastrin receptor is encoded by a single gene composed of five exons and spanning over 10 kilobases on human chromosome 11p 15.5-->15.4. Exon 4 has two possible alternative splicing donor sites that seem to be conserved in other species such as the canine, rat, Mastomys, and mouse. They could generate two receptor isoforms (short- and long-form), which differ in their putative third cytoplasmic domain of the serpentine G-protein-coupled receptors. In the present study, we examined whether an alternative splicing is operated in a tissue-specific manner and whether two receptor isoforms have functional differences. RNase-protection assay and S1 nuclease mapping demonstrated the preferential expression of the short-form in the human brain as well as the digestive organs, stomach and pancreas. The two putative isoforms of the cholecystokinin-B/gastrin receptor expressed in mouse fibroblasts showed the same characteristics in their ligand-bindings, the major signal transduction such as phosphoinositides production, cytoplasmic Ca2+ increase, tyrosine phosphorylation of focal adhesion kinase, activation of mitogen-activated protein kinase, and the induction of early-responsive genes such as c-fos, c-myc, and c-jun. Moreover, the ligand-dependent trophic effect was seen in both receptor isoforms. Taken together with the absence of tissue-specific expression of two receptor isoforms, these results suggest a species-specific dominant splice donor site in exon 4 of the human receptor gene. PMID: 7848914 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 638: Cell Growth Differ. 1994 Oct;5(10):1069-76. 12(S)-hydroxyeicosatetraenoic acid regulates DNA synthesis and protooncogene expression induced by epidermal growth factor and insulin in rat lens epithelium. Lysz TW, Arora JK, Lin C, Zelenka PS. Department of Surgery, UMD-New Jersey Medical School, Newark 07103-2714. Neonatal rat lens epithelium has a high 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE] synthetic capacity, which decreases as epithelial cell proliferation decreases with age. To determine whether products of the 12-lipoxygenase pathway are involved in lens cell proliferation, we measured the effect of 12-lipoxygenase inhibitors on endogenous 12-HETE production, epidermal growth factor/insulin-stimulated DNA synthesis and protooncogene expression in cultured neonatal rat lens epithelial cells. Incubation of neonatal rat lenses in epidermal growth factor plus insulin, which stimulated endogenous 12-HETE production 8- to 10-fold, also produced a transient induction of c-fos and c-myc mRNAs after 2 to 3 h, followed by a round of DNA synthesis approximately 20 h later. The lipoxygenase inhibitor, cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate, strongly inhibited both the endogenous 12-HETE synthesis and growth factor-stimulated DNA synthesis with a half-maximal inhibition between 10 and 20 microM. Cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate (10 microM) also inhibited the expression of c-fos and c-myc mRNA and, to a lesser extent, c-jun mRNA. The inhibitory effects of cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate on protooncogene expression and DNA synthesis were prevented by 0.3 microM 12(S)-HETE but not by equivalent concentrations of either 5(S)-HETE or 15(S)-HETE. These findings suggest that endogenously synthesized 12(S)-HETE may mediate epidermal growth factor/insulin-stimulated DNA synthesis in neonatal rat lens epithelial cells by regulating protooncogene expression. PMID: 7848908 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 639: J Biol Chem. 1994 Sep 30;269(39):24321-7. Activation of nuclear factor kappa B and oncogene expression by 12(R)-hydroxyeicosatrienoic acid, an angiogenic factor in microvessel endothelial cells. Laniado-Schwartzman M, Lavrovsky Y, Stoltz RA, Conners MS, Falck JR, Chauhan K, Abraham NG. Department of Pharmacology, New York Medical College, Valhalla 10595. 12(R)-Hydroxy-5,8,14(Z,Z,Z)-eicosatrienoic acid (12(R)-HETrE) is an arachidonic acid metabolite formed by the corneal epithelium of several species, porcine leukocytes, and human and rat epidermal cells. It is a potent, stereospecific proinflammatory and angiogenic factor and its synthesis is increased manyfold in inflamed tissues, e.g. cornea and skin. It is possible that the angiogenic activity of 12(R)-HETrE is due to a direct mitogenic effect on microvessel endothelial cells via yet to be elucidated cellular and molecular mechanisms. In the present study, we demonstrated the ability of 12(R)-HETrE to stimulate the growth of quiescent endothelial cells in a time- and concentration-dependent manner with a maximal effect at 0.1 nM. This effect was highly stereospecific since its enantiomer, 12(S)-HETrE, had no effect within the same concentration range. Northern blot analysis and transient transfection experiments with chloramphenicol acetyltransferase constructs of oncogene promoter regions demonstrated significant increases over control (0.5% fetal calf serum) in c-myc-, c-jun, and c-fos mRNA levels and expression in cells treated with 0.1 nM 12(R)-HETrE. Electrophoretic mobility shift assay of nuclear protein extracts from cells treated with 12(R)-HETrE with specific radiolabeled oligonucleotides corresponding to known transcriptional binding sites, including AP-1, AP-2, SP1, TRE, NF kappa B, TFIID, OKT1, CREB, CTF/NF1, and GRE demonstrated a markedly rapid and specific increase in the binding activity of NF kappa B and to a lesser extent, AP-1. No significant increase was observed in the binding of other transcription factors assayed as compared to control (untreated) cells. Since the protooncogenes (c-fos, c-jun, and c-myc) are immediate early response genes that are implicated in the process of cell proliferation and differentiation, and activation of certain transcription factors, in particular NF kappa B, is associated with the immediate response of the cell to an injury, we propose that 12(R)HETrE's mitogenic and angiogenic activities are mediated, in part, via the activation of NF kappa B and expression of these protooncogenes. PMID: 7523372 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 640: Cancer Res. 1994 Sep 15;54(18):4958-66. Differences between drug-sensitive and -resistant human leukemic CEM cells in c-jun expression, AP-1 DNA-binding activity, and formation of Jun/Fos family dimers, and their association with internucleosomal DNA ladders after treatment with VM-26. Kim R, Beck WT. Department of Molecular Pharmacology, St. Jude Children's Research Hospital, Memphis, Tennessee 38101. Although the DNA topoisomerases are critical intracellular targets of a number of clinically important anticancer drugs, the mechanism(s) by which inhibition of these enzymes causes cell death are poorly understood. We found that treatment of human leukemic lymphoblasts (CCRF-CEM) with teniposide (VM-26), under conditions that stabilize DNA-topoisomerase II complexes, caused the formation of internucleosomal DNA ladders. However, it appeared unlikely that the VM-26-stabilized DNA-topoisomerase II-cleavable complexes directly produce these internucleosomal DNA ladders, since similar nucleosomal DNA ladders were observed following either continuous or a short (1 h) exposure of cells to VM-26. Under continuous exposure to VM-26, the internucleosomal DNA ladders were associated with the transient induction of c-jun mRNA in a dose-dependent fashion, reaching maximum expression at 6 h after treatment with VM-26 and being down-regulated to basal levels by 12 h. The induction of c-jun mRNA by VM-26 apparently preceded DNA ladder formation. However, in CEM sublines selected for resistance to VM-26 (CEM/VM-1 and CEM/VM-1-5; approximately 50- and 140-fold resistant, respectively) and which display the phenotype of multidrug resistance associated with altered DNA topoisomerase II (at-MDR), we found that the induction of c-jun mRNA by VM-26 and subsequent DNA ladder formation were progressively attenuated in proportion to the resistance of the cells, apparently due in part to decreased stabilization of DNA-topoisomerase II-cleavable complexes. Further, the attenuated induction of c-jun in the at-MDR cells was found to be associated with a decreased rate of c-jun transcription and an increase in the instability of its mRNA following VM-26 treatment. The attenuation of c-jun mRNA induction was also reflected in decreased production of c-Jun protein in the at-MDR cells. Of interest was the fact that no significant induction of c-fos mRNA by VM-26 was observed in either CEM or at-MDR cells. Furthermore, the induction of c-jun was related to the activation of AP-1 DNA-binding activity in a time- and dose-dependent manner in CEM cells, whereas the activation of AP-1 binding was attenuated in at-MDR cells in proportion to their resistance to VM-26. Using Jun and Fos family member antibody inhibition experiments in gel-mobility shift assays, we found that AP-1-binding activity appeared to be preferentially mediated by c-Jun/Fra-1 heterodimers in both CEM and at-MDR cells.(ABSTRACT TRUNCATED AT 400 WORDS) PMID: 8069863 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 641: FEBS Lett. 1994 Sep 5;351(2):201-6. Interleukin 2-induced activation of JAK3: possible involvement in signal transduction for c-myc induction and cell proliferation. Asao H, Tanaka N, Ishii N, Higuchi M, Takeshita T, Nakamura M, Shirasawa T, Sugamura K. Department of Microbiology, Tohoku University School of Medicine, Sendai, Japan. We have investigated the role of JAK3 in interleukin 2 (IL-2)-induced signal transduction with a human T cell line, ED40515(-), lacking expression of the IL-2 receptor gamma chain and its sublines transfected with wild-type or mutant cDNAs of the IL-2 receptor gamma chain. Our results demonstrated that the membrane-proximal cytoplasmic region, encompassing the src homology region 2 (SH2)-like subdomain, of the gamma chain is essential for association and activation of JAK3. Furthermore, IL-2-induced activation of JAK3 paralleled induction of the c-myc gene and DNA synthesis but not induction of the c-fos and c-jun genes. These results support the hypothesis that JAK3 plays a pivotal role in the IL-2 receptor-mediated signals for cell growth. PMID: 8082765 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 642: J Allergy Clin Immunol. 1994 Sep;94(3 Pt 2):605-11. Activation of early response genes and cell proliferation by human interleukin-3, granulocyte-macrophage colony-stimulating factor, and interleukin-5 receptors: comparison with human interleukin-4 receptor signaling. Chen JX, Watanabe S, Muto A, Miyajima A, Yokota T, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Interleukin (IL)-3, granulocyte-macrophage colony-stimulating factor, and IL-5 receptors (IL-3R, GMR, and IL-5R) are composed of the alpha chain specific to each and the common beta chain, and both the alpha and beta subunits are members of the cytokine receptor superfamily. We previously showed that the high-affinity human GMR reconstituted by cotransfecting the alpha and beta chain cDNA clones transduces signals in response to hGM-CSF to activate transcription of c-fos, c-jun, and c-myc proto-oncogenes in mouse proB cell line BA/F3 or in mouse fibroblast NIH3T3 cells. These results indicated that molecules, such as tyrosine kinase, unique to hematopoietic cells are not essential to transduce signals. In this study, the function of the alpha subunit of GMR was compared with those of IL-3R and IL-5R by cotransfecting human cDNAs encoding the alpha subunit of IL-3R or IL-5R and the common beta subunit into BA/F3 or NIH3T3 cells. We found that the reconstituted human IL-3R, in response to hIL-3, transduced signals to activate transcription of c-fos promoter and induced DNA synthesis in both types of cells in a manner similar to hGMR. Likewise, hIL-5 activates c-fos promoter in transfected NIH3T3 cells expressing hIL-5R. These results indicated that the alpha subunits of IL-3R and IL-5R have properties similar to those of the GMR alpha subunit. In contrast, transfected human IL-4 receptor (hIL-4R) cDNA, which weakly activated c-fos promoter and induced DNA synthesis in BA/F3 cells, failed to elicit these activities in NIH3T3 cells in response to hIL-4.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8083468 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 643: Exp Cell Res. 1994 Sep;214(1):297-302. Cyclic AMP, early response gene expression, and DNA synthesis in rat smooth muscle cells. Hultgardh-Nilsson A, Querol-Ferrer V, Jonzon B, Krondahl U, Nilsson J. Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska Institutet, Stockholm, Sweden. Smooth muscle cells (SMC) isolated from neonatal and adult rats differ markedly in their growth characteristics. The growth of neonatal cells is mainly due to autocrine stimulation, whereas the growth of adult SMC is dependent upon addition of exogenous mitogens. Increasing intracellular cyclic AMP (cAMP) levels effectively inhibits DNA synthesis in adult cells, but is essentially without effect on the rate of DNA synthesis in neonatal cells. In the present study we investigated whether this difference in cAMP sensitivity is due to an effect of cAMP on early response genes. The results show that increasing intracellular levels of cAMP by exposing the cells to the synthetic adenosine analogue N-ethyl-carboxamido adenosine (NECA) results in an accumulation of c-jun and c-fos mRNA in both cell types. NECA also lowered c-myc mRNA levels in neonatal cells, whereas it marginally increased the presence of c-myc mRNA in adult cells. Exposure to NECA also resulted in a limited increase in alpha-actin mRNA levels. NECA did not inhibit DNA synthesis or growth of adult SMC actively proliferating in the presence of 10% serum, suggesting that cAMP interferes with processes taking place during the early G1 phase or in the entry of growth-arrested cells into the G1 phase of the SMC cell cycle. It is concluded that the growth-inhibitory effect of NECA is unlikely to be due to actions of cAMP on early response genes. However, it cannot be completely excluded that an increased synthesis of jun/fos transcription factors may induce the transcription of other, growth-suppressing genes in the cells. PMID: 8082733 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 644: Mol Cell Biol. 1994 Sep;14(9):5858-69. Cell-extracellular matrix interactions can regulate the switch between growth and differentiation in rat hepatocytes: reciprocal expression of C/EBP alpha and immediate-early growth response transcription factors. Rana B, Mischoulon D, Xie Y, Bucher NL, Farmer SR. Department of Biochemistry, Boston University School of Medicine, Massachusetts 02118. Previous investigations have shown that culture of freshly isolated hepatocytes under conventional conditions, i.e., on dried rat tail collagen in the presence of growth factors, facilitates cell growth but also causes an extensive down-regulation of most liver-specific functions. This dedifferentiation process can be prevented if the cells are cultured on a reconstituted basement membrane gel matrix derived from the Englebreth-Holm-Swarm mouse sarcoma tumor (EHS gel). To gain insight into the mechanisms regulating this response to extracellular matrix, we are analyzing the activities of two families of transcription factors, C/EBP and AP-1, which control the transcription of hepatic and growth-responsive genes, respectively. We demonstrate that isolation of hepatocytes from the normal quiescent rat liver by collagenase perfusion activates the immediate-early growth response program, as indicated by increased expression of c-jun, junB, c-fos, and c-myc mRNAs. Adhesion of these activated cells to dried rat tail collagen augments the elevated levels of these mRNAs for the initial 1 to 2 h postplating; junB and c-myc mRNA levels then drop steeply, with junB returning to normal quiescence and the c-myc level remaining slightly elevated during the 3-day culture period. Levels of c-jun mRNA and AP-1 DNA binding activity, however, remain elevated from the outset, while C/EBP alpha mRNA expression is down-regulated, resulting in a decrease in the steady-state levels of the 42- and 30-kDa C/EBP alpha polypeptides and C/EBP alpha DNA binding activity. In contrast, C/EBP beta mRNA production remains at near-normal hepatic levels for 5 to 8 days of culture, although its DNA binding activity decreases severalfold during this time. Adhesion of hepatocytes to the EHS gel for the same period of time dramatically alters this program: it arrests growth and inhibits AP-1 DNA binding activity and the expression of c-jun, junB, and c-myc mRNAs, but, in addition, it restores C/EBP alpha mRNA and protein as well as C/EBP alpha and C/EBP beta DNA binding activities to the abundant levels present in freshly isolated hepatocytes. These changes are not due merely to growth inhibition, because suppression of hepatocyte proliferation on collagen by epidermal growth factor starvation or addition of transforming growth factor beta does not inhibit AP-1 activity or restore C/EBP alpha DNA binding activity to normal hepatic levels. These data suggest that expression of the normal hepatic phenotype requires that hepatocytes exist in a G0 state of growth arrest, facilitated here by adhesion of cells to the EHS gel, in order to express high levels of hepatic transcription factors such as C/EBP alpha. PMID: 8065319 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 645: Arterioscler Thromb. 1994 Aug;14(8):1364-71. Response of atherosclerotic intimal smooth muscle cells to epidermal growth factor in vitro. Mitsumata M, Gamou S, Shimizu N, Yoshida Y. Department of Pathology, Yamanashi Medical University, Japan. Increased proliferation of intimal smooth muscle cells (SMCs) plays an important role in the early phase of atherogenesis. To investigate growth mechanisms of these cells, we used intimal SMCs from rabbits fed an atherogenic diet and examined the sequential events that may facilitate induction of intimal SMC proliferation as well as the possible effects of growth-promoting factors secreted by these cells. In serum-free medium, epidermal growth factor (EGF) stimulated [3H]thymidine uptake by quiescent intimal SMCs at a rate six times higher than quiescent medial SMCs. There was no significant difference between the two cell types in terms of the number of specific EGF receptor per cell, the dissociation constant of EGF, and the time course of EGF binding and internalization. Furthermore, in both types of cells, c-fos, c-jun, and c-myc mRNAs were induced after 1, 1, and 4 hours of EGF treatment, respectively, whereas they required 8 hours of contact with EGF to induce proliferation. Growth response of medical SMCs to EGF was greatly enhanced when rabbit serum, deficient in lipoproteins and free of platelet-derived growth factor, was added to the medium. Moreover, EGF induced a twofold to fourfold increase in DNA synthesis in medial SMCs cocultured with intimal SMCs compared with medial SMCs incubated alone. Likewise, DNA synthesis of medial SMCs grown in medium conditioned by intimal SMCs was six times higher than that observed in medium conditioned by medical SMCs. Adding EGF to the medium conditioned by intimal SMCs increased their DNA synthesis even further.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8049199 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 646: Am J Gastroenterol. 1994 Aug;89(8 Suppl):S86-96. Liver regeneration: a picture emerges from the puzzle. LaBrecque D. Department of Internal Medicine, University of Iowa Hospitals and Clinics, Iowa City. Liver regeneration remains a fascinating enigma. Many pieces of the puzzle have been elucidated, but as each piece is discovered, it is found to be composed of many smaller pieces, some of which are missing, and the multiple interlocking edges with other pieces of the puzzle remain poorly understood. The true initiating event or events remain unclear. The essential requirement for activation of immediate early genes is generally unchallenged. C-jun is essential for normal hepatogenesis in mouse development and it appears to be required for proliferation in response to injury, as well. Yet, nefenopin and cyproterone acetate induce hyperplastic responses in the liver with no induction of c-fos, c-myc, or c-jun. HGF is the single most potent liver mitogen yet discovered. However, levels of HGF do not correlate with the degree of liver regeneration, and high concentrations exist in conditions such as those in chronic hemodialysis patients who have no evidence of regeneration and minimal evidence of liver injury. Numerous conditions exist that induce immediate early genes and yet do not lead to cell proliferation. Thus, the availability of mitogens by themselves is not sufficient to induce regeneration, and the induction of immediate early genes is not sufficient to lead to regeneration. Whereas the isolated parenchymal cell culture system has been extremely valuable in identifying an increasing number of stimulatory and inhibitory substances and identifying the initial steps in their mechanisms of action, this simple system does not take into account the extremely complex interactions of these multiple growth factors in vivo and the interaction of the parenchymal cell with the other cellular and structural components of the liver. All must be accounted for in a complete model of liver regeneration. Parenchymal cell growth itself appears to be controlled by a series of steps, each of which requires the presence of specific growth regulators, which may be stimulators or inhibitors. There is a strictly defined sequence that must be followed, and if one or more of the factors is missing at the essential time point, growth will not progress and the cell will return to the G0 phase or proceed to apoptosis and death. While all this is occurring, the liver must also continue to carry out its essential life-supporting functions. And, finally, the liver must somehow know when to stop. Over-expression of some growth factors, such as TGF alpha, appears to produce tumors, whereas overexpression of others, such as HGF, does not.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 8048418 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 647: J Immunol. 1994 Aug 1;153(3):1310-7. Impairment of ligand binding and growth signaling of mutant IL-2 receptor gamma-chains in patients with X-linked severe combined immunodeficiency. Ishii N, Asao H, Kimura Y, Takeshita T, Nakamura M, Tsuchiya S, Konno T, Maeda M, Uchiyama T, Sugamura K. Department of Microbiology, Tohoku University School of Medicine, Sendai, Japan. The IL-2R gamma-chain is an indispensable subunit for the functional IL-2R. Recently, mutations of the gamma-chain have been reported to be closely associated with X-linked severe combined immunodeficiency (XSCID). The present study reveals that three patients with XSCID have three different mutations in the gamma-chain; a point mutation, a two consecutive-base deletion, and lack of the second exon in mRNA. The point mutation that we have detected is C to T, which results in one amino acid substitution of valine for alanine in the extracellular domain of the IL-2R gamma-chain (named AV mutant). The two-base deletion detected causes a frame shift of the coding region in the SH2 subdomain in the cytoplasmic domain (named tSH mutant). Transfection studies performed with the mutant gamma-chains demonstrated that the AV mutant and tSH mutant failed to bind to IL-2 and to transduce growth signals, respectively. These findings indicate that the gamma-chain gene mutations that accompany XSCID induce loss of the gamma-chain function, possibly resulting in stagnation of the differentiation and development of T cells. Publication Types: Case Reports PMID: 8027558 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 648: Drugs Aging. 1994 Aug;5(2):102-15. Hypertension and age-related changes in the heart. Implications for drug therapy. Isoyama S. First Department of Internal Medicine, Tohoku University School of Medicine, Sendai, Japan. Heart disease in older individuals can be characterised as the result of 2 processes, hypertension and atherosclerosis, which are the major causes of heart failure in the elderly population. The aging heart undergoes changes at the molecular, cellular and organ levels. These age-related changes may then be modulated by pathological conditions, such as hypertension, and by the reduction of blood pressure. One characteristic of the aged heart is a limited capacity for adaptation, by hypertrophy, to increased mechanical load. This age-related attenuation of the hypertrophic response may be attributed to the diminished induction of proto-oncogenes such as c-fos, c-myc and c-jun. This diminution results from aging of the heart per se and may be modulated by extracardiac factors. With regard to the coronary vasculature, the age at which hypertension develops seems to be an important factor for determining the vascularity of hypertrophied hearts. Late-onset hypertension is not accompanied by coronary angiogenesis, and it decreases dilator reserve in spite of the absence of myocardial hypertrophy. In contrast, mechanical overload in infant hearts is accompanied by angiogenesis and normal dilator reserve. In principle, the normalisation of hypertension results in the regression of myocardial hypertrophy and decreased coronary dilator reserve. In aged hearts, it is not clear how hypertension-induced myocardial hypertrophy or coronary vascular changes regress. Although antihypertensive treatment is clearly associated with an improvement of cardiovascular mortality and morbidity in hypertensive elderly individuals, it remains unclear how treatments ameliorate the hypertension-induced alterations. Publication Types: Review PMID: 7981482 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 649: Leuk Res. 1994 Aug;18(8):577-85. Interferon-alpha-2b downregulation of oncogenes H-ras, c-raf-2, c-kit, c-myc, c-myb and c-fos in ESKOL, a hairy cell leukemic line, results in temporal perturbation of signal transduction cascade. Harvey WH, Harb OS, Kosak ST, Sheaffer JC, Lowe LR, Heerema NA. Department of Biology, Earlham College, Richmond, IN 47374-4095. ESKOL, a B-lymphoblastoid cell line consisting of late differentiated cells, resembles hairy cell leukemia (HCL). It is pseudodiploid with a deleted 7q and an unbalanced translocation between chromosomes 4 and 6. It was screened by Northern hybridization for oncogenes, including H-ras, c-raf-2 (c-raf1p1), c-kit, c-myc, c-myb, c-fos, Fim-1, c-jun, ski, and c-mos, which are believed to contribute to B-cell differentiation and maturation. Interferon-alpha-2b (IFN) downregulates the expression of H-ras, c-raf-2, c-kit, c-myc, c-myb, c-fos, as determined by Northern hybridization of RNA isolated from cells harvested at time points during a 30 h time course. Downregulation of oncogenes H-ras, c-raf-2, c-kit, whose proteins are associated with cell surfaces or are cytosolar transducers, occurs before those oncogenes c-myc, c-myb, and c-fos, whose products are DNA binding proteins. This suggests a temporal perturbation of signal transduction by IFN. No change in oncogene expression occurred in non-treated cells nor were these oncogenes expressed in the non-transformed B-lymphoblast cell line, Wil-2, under the same treatment regimen. The basis for the IFN perturbation is not understood; yet the role of these oncogenes as signal transducers in differentiation and proliferation of human hematopoietic progenitors is unfolding, and ESKOL is an excellent system in which to study this phenomenon. PMID: 7520517 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 650: J Clin Invest. 1994 Jul;94(1):210-8. Alpha 1 adrenergic receptor-induced c-fos gene expression in rat aorta and cultured vascular smooth muscle cells. Okazaki M, Hu ZW, Fujinaga M, Hoffman BB. Department of Medicine, Stanford University School of Medicine, California. While growth of blood vessels is important in hypertension, relatively little is known about the contribution of catecholamines. Using isolated rat aorta and cultured smooth muscle cells, we examined adrenergic stimulation of gene expression. Phenylephrine, a selective alpha 1 adrenergic receptor agonist, caused a rapid and transient increase in c-fos mRNA accumulation which was inhibited by prazosin, an alpha 1 receptor antagonist. Similarly, phenylephrine stimulated c-jun and c-myc mRNA accumulation. Chloroethyl-clonidine, a compound which irreversibly blocks alpha 1B receptors, completely blocked the phenylephrine-induced increase in c-fos mRNA. RNase protection experiments demonstrated that rat aorta prominently expressed mRNA for alpha 1B and alpha 1A/D receptors. Phenylephrine-induced c-fos mRNA was partially inhibited by H-7, a protein kinase C inhibitor, and by nifedipine, a Ca2+ channel blocker; these two compounds together had additive effects. In situ hybridization showed that expression of c-fos mRNA induced by phenylephrine was localized to aorta's medial layer. These results suggest that alpha 1 receptor-induced increase in c-fos mRNA in aorta is mediated by a chloroethyl-clonidine-sensitive receptor subtype signaling via increasing intracellular Ca2+ concentrations and activating protein kinase C. PMID: 8040263 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 651: Jpn Heart J. 1994 Jul;35(4):403-18. Responses to hemodynamic stress in the aged heart. Isoyama S, Ito N, Komatsu M, Nitta Y, Abe K, Aoki M, Takishima T. First Department of Internal Medicine, Tohoku University School of Medicine, Sendai, Japan. In aged individuals the incidence of heart failure is higher than in younger subjects. Ischemic events are also common in the aged heart because of changes in the coronary vasculature and myocytes caused by aging. Adaptational responses to increased hemodynamic overload and to ischemia in the aged heart are discussed at the molecular, cellular and organ levels. One characteristic of the aged heart is a limited capacity for adaptation with hypertrophy to increased mechanical load. This age-related attenuation of the hypertrophic response may be attributed to the diminished induction of proto-oncogenes such as c-fos, c-myc and c-jun by hemodynamic stress. This diminution results from the aging of the heart per se and may be modulated by extracardiac factors. An age-related diminution was also observed in the mRNA induction of heat shock proteins by transient ischemia. However, this diminished induction of immediate early genes in the aged heart was not observed after more severe stress. With regard to the coronary vasculature, the age at which pressure-overload begins seems to be one of the important factors which determine the vascularity of hypertrophied hearts. Late-onset pressure-overload decreased dilator reserve in spite of the absence of myocardial hypertrophy. Thus, the responses to stress in the aged heart are quite different from those in the young heart. The limited capacity for adaptation to hemodynamic overload and poor protective mechanisms against stress may be causes of the higher incidence of heart failure in the aged. Publication Types: Review Review, Tutorial PMID: 7967046 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 652: Cardiovasc Res. 1994 Jul;28(7):1062-9. Changes in gene expression following short coronary occlusions studied in porcine hearts with run-on assays. Knoll R, Arras M, Zimmermann R, Schaper J, Schaper W. Max-Planck-Institute, Department of Experimental Cardiology, Bad Nauheim, Germany. OBJECTIVE: Brief coronary occlusions cause upregulation of expression in a wide variety of genes. These changes in tissue mRNA concentration could have been produced by transcriptional or post-transcriptional events. The aim of this study was to discriminate between increased transcription and changes in mRNA stability using run-on assays with isolated myocyte nuclei. METHODS: Myocyte nuclei isolated from ischaemic/reperfused and normal myocardium were incubated with labelled ribonucleotides. The radioactive RNA was then hybridised with specific cDNA probes and slot blots were autoradiographed. RESULTS: There was increased transcriptional activity for the proto-oncogenes c-myc, c-jun, jun-B, and jun-D. There were marked increases in transcriptional activity for sarcoplasmic Ca(2+)-ATPase, calmodulin, phospholamban, and calsequestrin. Strong transcriptional activity was found for the ubiquitin and heat shock protein (hsp27, hsp70) genes, and for PAI-1 and GAPDH. The transcription for the beta myosin heavy chain gene was not altered. CONCLUSIONS: Changes in the tissue concentration of mRNA species following brief coronary occlusion and reperfusion are most often the result of altered transcriptional activity. Increased c-fos mRNA concentrations observed in earlier studies cannot be explained by transcriptional activity of myocytes during reperfusion. Calmodulin is strongly transcribed but tissue concentration stays constant. The overall pattern of gene expression is indicative of damage at the molecular level, and calcium binding proteins (among perhaps many others) are in need of repair. PMID: 7954593 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 653: Stem Cells. 1994 Jul;12(4):352-69. Differentiation primary response genes and proto-oncogenes as positive and negative regulators of terminal hematopoietic cell differentiation. Liebermann DA, Hoffman B. Fels Institute for Cancer Research and Molecular Biology, Temple University, School of Medicine, Philadelphia, Pennsylvania 19140. By genetically manipulating hematopoietic cells of the myeloid lineage, including both normal cells and differentiation inducible leukemic cell lines, evidence was obtained to indicate that myeloid differentiation primary response (MyD) genes and proto-oncogenes, which are known to control cell growth, function as positive and negative regulators of terminal hematopoietic cell differentiation, which is associated with inhibition of cell growth, and, ultimately programmed cell death (apoptosis). Interferon regulatory factor-1 (IRF-1), an MyD gene induced by Interleukin 6 (IL-6) or Leukemia Inhibitory factor (LIF), plays a role in growth inhibition associated with terminal differentiation. Leucine zipper transcription factors of the fos/jun family, also identified as MyD genes, function as positive regulators of hematopoietic cell differentiation, increasing the propensity of myeloblastic leukemia cells to be induced for differentiation in vitro, and reducing the aggressiveness of their leukemic phenotype in vivo. The zinc finger transcription factor EGR-1, an MyD gene specifically induced upon macrophage differentiation, was shown to be essential for and to restrict differentiation along the macrophage lineage. Finally, evidence has been accumulating to indicate that the novel MyD genes--MyD116, MyD118 and gadd45 (a member in the MyD118 gene family)--play a role in growth arrest and apoptosis of hematopoietic cells, as well as other cell types. The proto-oncogenes c-myc and c-myb, known to regulate cellular growth, were shown to function as negative regulators of terminal differentiation. Both c-myc and c-myb are normally expressed in proliferating myeloblasts and suppressed following induction of differentiation. Deregulated and continuous expression of c-myc was shown to block terminal myeloid differentiation at an intermediate stage in the progression from immature blasts to mature macrophages, whereas deregulated and continuous expression of c-myb blocked the terminal differentiation program at the immature myeloblast stage. By manipulating myc function in conditional (differentiation inducible) mutant myeloblastic leukemia cell lines, expressing a chimeric mycer transgene, it was shown that there is a window during myeloid differentiation, after the addition of the differentiation inducer, when the terminal differentiation program switches from being dependent on c-myc suppression to becoming c-myc suppression independent, and where activation of c-myc has no apparent effect on mature macrophages. These myeloblastic leukemia cell lines provide a powerful tool to increase our understanding of the role of c-myc in normal hematopoiesis and in leukemogenesis, while also providing a strategy to clone myc target genes.(ABSTRACT TRUNCATED AT 250 WORDS) Publication Types: Review PMID: 7951003 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 654: Bioessays. 1994 Jul;16(7):489-96. Regulation of the Ras signalling network. Maruta H, Burgess AW. Ludwig Institute for Cancer Research, Melbourne, Australia. The mitogenic action of cytokines such as epidermal growth factor (EGF) or platelet derived growth factor (PDGF) involves the stimulation of a signal cascade controlled by a small G protein called Ras. Mutations of Ras can cause its constitutive activation and, as a consequence, bypass the regulation of cell growth by cytokines. Both growth factor-induced and oncogenic activation of Ras involve the conversion of Ras from the GDP-bound (D-Ras) to the GTP-bound (T-Ras) forms. T-Ras activates a network of protein kinases including c-Mos, c-Raf-1 and MAP kinase. Eventually the activation of MAP kinase leads to the activation of the elongation factor 4E and several transcription factors such as c-Jun, c-Myc and c-Fos. There are several modulators of Ras activity, such as the GTPase activating proteins (GAP1 and NF1), which stimulate the conversion of T-Ras to D-Ras. A series of small NF1 fragments, which bind T-Ras, as well as truncated forms of derivatives of c-Raf-1, c-Jun and c-Myc, are capable of blocking the T-Ras-activated mitogenesis in a competitive manner. These agents offer a unique opportunity to control the proliferation of T-Ras-associated tumors, which represent more than 30% of total human carcinomas. Publication Types: Review Review, Tutorial PMID: 7945277 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 655: Clin Exp Metastasis. 1994 Jul;12(4):329-34. Analysis of c-fos, c-jun, c-erbB1, c-erbB2 and c-myc in primary lung carcinomas and their lymph node metastases. Volm M, van Kaick G, Mattern J. German Cancer Research Center, Heidelberg. The purpose of this study was to identify possible alterations in proto-oncogenes (c-fos, c-jun, c-erbB1, c-erbB2 and c-myc) at the protein level in primary lung carcinomas and simultaneous metastatic lymph nodes of 21 patients. The analysis showed that proteins of c-jun and c-myc were expressed in a significantly higher frequency in metastases than in primary lung tumors. Gross differences were not found between primary tumors and metastatic tumors with regard to the expression of c-erbB1, c-erbB2 and c-fos. The finding of cases with a higher expression of c-jun and c-myc in lymph nodes suggests that metastatic capability may be higher in certain cell populations. PMID: 7913670 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 656: Arkh Patol. 1994 Jul-Aug;56(4):22-31. [Cell oncogene expression in normal, metaplastic, dysplastic epithelium and squamous cell carcinoma of the uterine cervix] [Article in Russian] Petrov SV, Mazurenko NN, Sukhova NM, Moroz IP, Katsenel'son VM, Raikhlin NT, Kiselev FL. Immunohistochemical analysis of the protein expression c-myc, ets 1, ets 2, TPR-met, c-fos, c-jun, c-ras-pan, p53, yes, src in 79 samples of normal, metaplastic squamous epithelium, intraepithelial and invasive squamous cell carcinoma of uterine cervix was performed using polyclonal rabbit antibodies to the synthetic peptides homologous active areas of corresponding oncoproteins. Higher content of myc, fos, ets2, p53, ras is noted in metaplasia, dysplasia and in tumours as compared to the normal tissues. Protein myc is revealed in the cytoplasm at a grave dysplasia and in the nucleus in the intraepithelial carcinoma: this may serve as a criterion at a differential diagnosis of these conditions. Expression of the oncoproteins fos, ets2, p53, src in the metaplastic squamous cell carcinoma was higher than in the true squamous cell (ectocervical) carcinoma. When compared to the advanced carcinomas, increase of ets2, p53, and at some degree that of myc, the increase is noted in the latter. Invasive carcinoma with a high level of oncoproteins showed a tendency to the synchronization of myc and ras expression. Poor prognosis was associated with a low level (before treatment) of the expression of the majority of the oncoproteins studied. PMID: 7848100 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 657: FEBS Lett. 1994 Jun 27;347(2-3):157-62. Changes in cell cycle-associated gene expression in a model of impaired liver regeneration. Albrecht JH, Hoffman JS, Kren BT, Steer CJ. Department of Medicine, Hennepin County Medical Center, Minneapolis, MN 55415. Following partial hepatectomy (PH) there is compensatory regeneration of the remnant liver which eventually restores hepatic mass and function. The response to PH was studied in normal BALB/c and athymic nude mice, a model of impaired liver regeneration. Following PH, nude mice demonstrated diminished peak hepatic [3H]thymidine uptake and delayed liver mass restoration through 60 h post-PH. However, between 72-120 h there was no significant difference in mass restoration between the groups. The expression of genes associated with different stages of the cell cycle was evaluated in both models. In nude mice, there was an increase in peak expression of c-jun transcripts, while c-myc transcript expression was moderately attenuated. Thymidine kinase (TK) and cyclin-dependent kinase 1 (CDK1) mRNA expression was also diminished in athymic nude mice. The results suggest that while the defect in the regenerative response of the nude mouse after PH affects events in several phases of the cell cycle, mass restoration of the liver is only delayed and not attenuated. PMID: 8033995 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 658: Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):5937-41. Induction of tyrosine hydroxylase gene expression by a nonneuronal nonpituitary-mediated mechanism in immobilization stress. Nankova B, Kvetnansky R, McMahon A, Viskupic E, Hiremagalur B, Frankle G, Fukuhara K, Kopin IJ, Sabban EL. Department of Biochemistry and Molecular Biology, New York Medical College, Valhalla 10595. Stress stimulates the sympathoadrenal system, causing activation of the catecholamine biosynthetic enzymes. Here we examine the changes of gene expression of tyrosine hydroxylase (TH; EC 1.14.16.2), the initial enzyme of catecholamine biosynthesis, with stress. A single immobilization of rats led to a large transient elevation in TH mRNA and a small elevation in TH immunoreactive protein and activity. Repeated daily immobilizations triggered more sustained changes in TH mRNA levels. After two immobilizations, the levels remained elevated even 3 days later. The rise in TH mRNA was followed by increased immunoreactive protein but only a small elevation in activity. With seven repeated immobilizations, the animals did not appear to adapt and still manifested a further rise in TH mRNA. TH activity was markedly elevated and returned to control levels 7 days after the immobilization. The rise in TH mRNA with a single immobilization occurred even in adrenals of hypophysectomized rats that underwent splanchnic nerve section. Immobilization for 30 min was sufficient to increase TH mRNA. The effect was abolished by the transcriptional inhibitor actinomycin D. Mobility gel-shift assays revealed increased binding of c-Fos and c-Jun to the AP-1 transcription factor site after a single immobilization, and the binding was not further elevated with repeated stress. This study shows that a single immobilization can activate TH gene expression by a nonneuronal nonpituitary-mediated pathway associated with increased binding of AP-1 transcription factors. PMID: 7912437 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 659: J Immunol. 1994 Jun 15;152(12):5680-90. Signal transduction mediated by the reconstituted IL-2 receptor. Evidence for a cell type-specific function of IL-2 receptor beta-chain. Minami Y, Oishi I, Liu ZJ, Nakagawa S, Miyazaki T, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. The binding of IL-2 to its specific receptor (IL-2R) triggers various cellular events including the induction of nuclear proto-oncogenes (c-fos, c-jun and c-myc genes) and the proliferation of hemopoietic cells. In the present study, we have established NIH 3T3 fibroblasts in which the three IL-2R subunits, the alpha-chain (IL-2R alpha), the beta-chain (IL-2R beta), and the gamma-chain (IL-2R gamma), are constitutively expressed. The resulting cell lines express high affinity IL-2R on their cell surface at levels comparable with those of IL-2-responsive lymphoid cells. We observed that the high affinity IL-2R in NIH 3T3 fibroblasts can mediate the IL-2-stimulated tyrosine phosphorylation of p42/p44 (mitogen-activated protein kinase) and the induction of the c-fos, c-jun and c-myc genes. In NIH 3T3 fibroblasts the high affinity IL-2R bearing a deletion of a region rich in acidic amino acids (the "acidic" region) in the IL-2R beta-chain failed to induce the tyrosine phosphorylation of MAP kinase as well as the expression of the all three nuclear proto-oncogenes. On the other hand, our previous studies had demonstrated that the high affinity IL-2R bearing the same mutant IL-2R beta-chain retained the ability to induce c-myc gene in response to IL-2 in a murine IL-3-dependent pro-B cell line, BAF/B03. Hence, these results reveal the underlying complexity of signal transduction among different cell types. The inability of the reconstituted high affinity receptor to mediate the IL-2-induced proliferation of NIH 3T3 fibroblasts suggests that induction of the three nuclear proto-oncogenes and the tyrosine phosphorylation of mitogen-activated protein kinase in NIH 3T3 fibroblasts are not sufficient to induce cellular proliferation. PMID: 8207200 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 660: J Virol. 1994 Jun;68(6):3527-35. Analysis of AP-1 function in cellular transformation pathways. Suzuki T, Murakami M, Onai N, Fukuda E, Hashimoto Y, Sonobe MH, Kameda T, Ichinose M, Miki K, Iba H. Department of Tumor Virus Research, University of Tokyo, Japan. To understand the role of endogenous AP-1 activity in cellular transformation induced by oncogenes, we have made use of a fos mutant (supfos-1) and a jun mutant (supjun-1), either of which can function as a transdominant inhibitor of AP-1-mediated transcriptional regulation. Chicken embryo fibroblasts (CEF) infected with a series of transforming retroviruses were doubly infected with retrovirus carrying supfos-1 or supjun-1, and suppression of cellular transformation was monitored in terms of reversion to normal cellular morphology or acquisition of anchorage-dependent growth. Cellular transformation induced by several exogenously expressed transforming genes of the fos or jun family was efficiently suppressed, as expected. CEF transformed by v-src, v-yes, v-fps, c-Ha-ras, and N-terminally truncated c-raf were also induced to revert to the normal phenotype by these transdominant mutants, suggesting that functional transcription factor AP-1 activity is essential for the cellular transformation induced by these oncogenes. The suppression is not attributable to nonspecific inhibition of cellular proliferation, because CEF transformed by v-ros or v-myc were not induced to revert to the normal phenotype. We next analyzed changes in all known components of chicken AP-1 induced by v-src, c-Ha-ras, or activated c-raf transformation. The levels of both Fra-2 and c-Jun expression were elevated two- to fourfold, and hyperphosphorylation of Fra-2 was also observed. We further showed that Fra-2-c-Jun heterodimer is mainly responsible for the elevated AP-1 DNA-binding activity in these transformed cells, and we propose that this heterodimer play a crucial role in the transformation induced by these oncogenes. PMID: 8189491 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 661: Hum Reprod. 1994 Jun;9 Suppl 1:174-80. Regulation of gene expression in T-47D human breast cancer cells by progestins and antiprogestins. Murphy LC, Alkhalaf M, Dotzlaw H, Coutts A, Haddad-Alkhalaf B. Department of Biochemistry and Molecular Biology, University of Manitoba, Winnipeg, Canada. The molecular mechanisms by which progestins and antiprogestins inhibit human breast cancer cell growth are essentially unknown. The mechanisms by which they mediate growth inhibition in human breast cancer cells and the expression of the putative autocrine/paracrine growth factors, epidermal growth factor and transforming growth factors alpha and beta-1 were studied under conditions in which progestin and antiprogestin inhibit the growth of T-47D human breast cancer cells in culture. Under the same conditions, the expression of genes such as c-myc, c-jun and c-fos, which are known to have important roles in growth and differentiation, has been measured. The results indicate that progestins and antiprogestins differentially regulate expression of these genes. The data are consistent with the conclusion that the mechanism of growth inhibition of these two agents differs, although an initial interaction with the progesterone receptor is a necessary first step in initiating the as yet ill-defined cascade of events leading to growth inhibition. Publication Types: Review Review, Tutorial PMID: 7962462 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 662: J Cell Physiol. 1994 Jun;159(3):441-9. Enhancement of differentiation induction of mouse myelomonocytic leukemic cells by extracellular ATP. Yamaguchi M, Hirayoshi K, Okuma M, Nagata K. Department of Cell Biology, Chest Disease Research Institute, Kyoto, Japan. We examined the effect of ATP on the terminal differentiation of mouse myelomonocytic leukemic M1 cells to macrophages. Although ATP alone did not induce M1 cell differentiation, addition of ATP with the differentiation inducer, interleukin 6 (IL-6), enhanced the induction of differentiation by IL-6 about two-fold. Comparison among several adenine nucleotides revealed that the order of effectiveness on differentiation enhancement was ATP > ADP > AMP > or = adenosine. Using reactive blue 2, a P2 receptor antagonist, we confirmed that the effect of ATP on the stimulation of differentiation was mediated through the P2 purinergic receptor. Examination of cytosolic [Ca2+] elevation by ATP and comparison of potency of differentiation enhancement among several ATP analogs demonstrated that the effect of differentiation enhancement was mediated through P2y purinergic receptors expressed on M1 cell surface. Within 3 h of exposure, ATP alone slightly increased the expression of differentiation-related immediate early response genes, c-myc and JunB, and ATP also enhanced the IL-6-induced expression of these genes. Induction of JunB expression by ATP analogs correlated with their potency of differentiation enhancement, which suggested that induction of JunB by ATP is one of signaling pathways involved in the exertion of its differentiation-enhancing effect. PMID: 7514609 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 663: Proc Natl Acad Sci U S A. 1994 May 24;91(11):4649-53. Adipocyte differentiation selectively represses the serum inducibility of c-jun and junB by reversible transcription-dependent mechanisms. Wang H, Scott RE. Department of Pathology, University of Tennessee College of Medicine, Memphis 38163. Nonterminally differentiated 3T3 T adipocytes are resistant to growth stimulation by 10% (vol/vol) fetal bovine serum even though they can be induced to proliferate with extremely high serum concentrations. We now report that in adipocytes 10% fetal bovine serum also fails to typically induce c-jun or junB. Rather, after 10% fetal bovine serum treatment, c-jun and junB expression is markedly repressed after a brief initial slight induction. Gel mobility shift studies confirm that AP-1 DNA binding activity is inhibited in adipocytes. Repression in c-jun and junB inducibility in adipocytes results from transcriptional mechanisms, can be reversed by treatment with protein synthesis inhibitors or higher serum concentrations, and does not affect c-fos or c-myc expression. These data suggest that adipocyte differentiation selectively and transcriptionally represses the inducibility of c-jun and junB so as to decrease the cell's ability to proliferate in response to 10% fetal bovine serum. PMID: 8197114 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 664: Am J Physiol. 1994 May;266(5 Pt 1):C1231-9. Gene expression during phorbol ester-induced differentiation of cultured human megakaryoblastic cells. Dorn GW 2nd, Davis MG, D'Angelo DD. Department of Internal Medicine, University of Cincinnati, Ohio 45267-0542. Platelet protein makeup is determined during transformation of megakaryoblasts to mature megakaryocytes, the immediate precursor of circulating platelets. To better understand the molecular mechanisms of megakaryocyte formation, gene expression was characterized by Northern analysis and RNA fingerprinting of cultured human CHRF-288 megakaryoblastic cells undergoing phorbol ester-stimulated megakaryocytic differentiation or serum-stimulated megakaryoblast proliferation. Protooncogenes c-fos and c-jun were coordinately upregulated in both proliferating and differentiating cells, whereas c-myc transcripts were upregulated during proliferation only. In contrast, mRNAs for transforming growth factor-beta 1 (TGF-beta 1) and thromboxane receptors were coordinately upregulated during differentiation but differentially regulated during proliferation. RNA fingerprinting revealed multiple transcripts specific to either proliferating or differentiated cells. Three of these were identified by homology to known DNA sequence: CDw44 adhesion molecule (upregulated during differentiation), glutathione sulfhydryl peroxidase (downregulated during differentiation), and plectin cytoskeletal protein (upregulated during differentiation). Thus, although megakaryoblast proliferation and megakaryocyte differentiation both involve DNA and protein synthesis, each growth response is characterized by a distinct pattern of gene expression. PMID: 8203487 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 665: Leuk Res. 1994 May;18(5):319-25. Induction of proto-oncogene and cytokine expression in human peripheral blood monocytes and the monocytic cell line THP-1 after stimulation with mycoplasma-derived material MDHM. Quentmeier H, Schumann-Kindel G, Muhlradt PF, Drexler HG. DSM-German Collection of Microorganisms and Cell Cultures, Department of Human and Animal Cell Cultures, Braunschweig. Mycoplasma fermentans-derived high-molecular-weight material (MDHM) was originally described to induce differentiation of murine thymocytes to cytolytic effector T-cells by stimulating IL-6 release from adherent cells. This study shows that human peripheral blood monocytes (PBMo) also respond to MDHM with increases in IL-1 beta, IL-6 and TNF alpha expression, both at the mRNA and protein level. The induced expression of IL-1 beta and TNF alpha mRNA in the monocytic THP-1 cell line increased as quickly as in primary cells. In contrast to PBMo, THP-1 and 14 other monocytic/myeloid leukemia-derived cell lines did not secrete measurable amounts of the cytokines upon treatment with MDHM. IL-1 beta and IL-6 genes contain AP-1 binding sites as regulatory elements, the AP-1 protein being composed of c-jun and c-fos gene products. In THP-1 cells c-jun mRNA expression increased after incubation with MDHM while positive c-fos expression remained unaffected. Although these data suggest AP-1 regulated cytokine mRNA expression, results from PBMo are not in accordance with this notion. In the primary cells MDHM-induced elevation of cytokine mRNA levels was preceded by a downregulation of c-fos expression while positive c-jun expression was not modulated. c-myc mRNA expression, constitutively high in THP-1 cells, was induced in MDHM-stimulated PBMo. In conclusion, MDHM-stimulated induction of cytokine mRNA expression was accompanied by different proto-oncogene responses in PBMo and THP-1 cells. These differences may represent different regulatory pathways of the two cell systems. Alternatively, these data support the notion that neither AP-1 nor the c-myc protein are involved in the MDHM-induced increase in IL-1 beta, IL-6 or TNF alpha mRNA levels. Furthermore, the present results demonstrate clearly that mycoplasma products can have a profound impact on the activation status of eukaryotic cells. PMID: 8182922 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 666: J Card Surg. 1994 May;9(3 Suppl):537-42. Myocardial stunning: association with altered gene expression. Wechsler AS, Entwistle JW 3rd, Ding M, Yeh T Jr, Jakoi ER. Department of Surgery, Medical College of Virginia, Richmond 23298-0645. Regional and global myocardial ischemia and reperfusion have been demonstrated to induce expression of the stress response protein heat shock 70 (HSP70) and of immediate early genes, c-jun, c-fos, and c-myc. Because of the models that have been utilized, it has not been possible to discriminate whether this response is the consequence of ischemia, reperfusion, or abnormal hemodynamic stress superimposed on stunned myocardium. In a nonworking isolated and blood-perfused rat heart model, we evaluated the mRNAs for c-fos, c-myc, and hsp70. The heart was subjected to varying periods of ischemia and reperfusion. Significant increases in hsp70 and c-fos were observed, which increased with longer periods of ischemia. No significant increase in c-myc was measured. In addition, mRNA encoding the Ca2+/glucose responsive stress protein GRP78 was evaluated. No increase in this early response gene was noted despite the use of a model associated with cellular calcium loading. Based on these observations, we suggest that the induction of hsp70 and c-fos is the consequence of ischemia and reperfusion and not dependent upon an early hypertrophy response such as would be observed in afterload mismatching or on calcium loading. Further investigations are necessary to isolate the effects of ischemia from those of reperfusion. PMID: 8069049 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 667: Leukemia. 1994 May;8(5):740-8. Signal transduction of steel factor and granulocyte-macrophage colony-stimulating factor: differential regulation of transcription factor and G1 cyclin gene expression, and of proliferation in the human factor-dependent cell line MO7. Hallek M, Ando K, Eder M, Slattery K, Ajchenbaum-Cymbalista F, Griffin JD. Dana-Faber Cancer Institute, Boston, Massachusetts. Steel factor (SF) synergizes with a variety of hemopoietins to support the growth and differentiation of human progenitor cells. The human factor-dependent cell line MO7 has been used as a model to study the interaction of SF with other growth factors such as GM-CSF, because both factors support the proliferation of this cell line and are synergistic in combination. Previous studies have shown that this effect is not readily explained by the synergistic activation of early, cytosolic signal transduction intermediates such as tyrosine kinases, Raf-1, MAP2 kinase, or phospholipase C gamma. In an attempt to further explore the biological and biochemical mechanisms of the synergy between SF and GM-CSF, we examined the effects of these growth factors on the regulation of nuclear proto-oncogenes, cell cycle control genes, and G1-->S transition of MO7 cells. Individually, GM-CSF was a much more potent growth factor for MO7 cells than SF, particularly under serum-free conditions. Only GM-CSF, but not SF, was able to stimulate G1-->S transition of MO7 cells after factor deprivation for 24 h. Northern blot analyses showed also differential effects of GM-CSF and SF on the expression of some nuclear proto-oncogenes and G1 cyclins. GM-CSF (10 ng/ml), but not SF (20 ng/ml) increased the expression of c-myc and cyclin D2 mRNA, whereas both factors caused transient increases of c-fos and cyclin D3 mRNAs. When added simultaneously, GM-CSF and SF induced an at least additive increase of c-fos mRNA expression; this effect required the presence of fetal calf serum. No additive effects of GM-CSF and SF on c-myc, cyclin D2 or D3 mRNA expression were observed. C-jun and c-myb mRNAs were constitutively expressed in the MO7 cell line, but not further increased after stimulation with GM-CSF or SF for 15 min to 48 h. The inability of SF to induce growth promoting genes such as c-myc and cyclin D2 may explain why this cytokine does not support sustained proliferation of MO7 cells. These observations suggest that SF and GM-CSF exert different effects on the expression of genes involved in regulatory pathways of cell proliferation, but the molecular mechanism of synergy remains to be elucidated. PMID: 7514243 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 668: Mutat Res. 1994 May 1;307(1):43-51. X-ray-induced transcriptional activation of c-myc and XRCC1 genes in ataxia telangiectasia cells. Shung B, Miyakoshi J, Takebe H. Department of Biology, College of Natural Sciences, Chonnam University, Yongbong-Dong, Kwangju, South Korea. The transcriptional level of c-myc, c-jun and XRCC1 genes after X-irradiation was compared in human cells originating from subjects presumably with different DNA repair abilities. The mRNA amount of the beta-actin gene was used as an internal standard of transcription. The relative mRNA level of c-myc and XRCC1 genes was significantly increased 15 min after X-irradiation with doses of 2-8 Gy in ataxia telangiectasia (AT) cells (AT5BIVA and TAT2SF), in contrast to little change in xeroderma pigmentosum (XP2OS(SV) and XP2YO(SV)) and normal cells (WI38VA13 and GM0637). The increased mRNA level of the XRCC1 gene in AT5BIVA and of the c-myc and XRCC1 genes in TAT2SF cells was maintained for up to 8 h after X-irradiation with 2 Gy. For the c-jun mRNA level after X-irradiation with 2-8 Gy, no significant change was observed in all cell lines tested. These results indicate that AT cells show a high transcriptional response of certain genes in response to X-irradiation, and suggest that the transcriptional activation of c-myc and XRCC1 genes after X-irradiation may be related to the hyper-radiosensitivity of AT cells. PMID: 7513822 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 669: Biochim Biophys Acta. 1994 Apr 12;1226(1):89-96. Oncogene overexpression and de novo drug-resistance in human prostate cancer cells. Yamazaki H, Schneider E, Myers CE, Sinha BK. Biochemical & Molecular Pharmacology Section, NCI, NIH, Bethesda, MD 20892. We have isolated a variant [PC3(R)] of the human prostate PC3 tumor cell line which showed resistance to several anticancer drugs. Studies to evaluate the mechanisms of resistance to anticancer drugs in the PC3(R) cell line indicated that mdr1 was not overexpressed. Studies also indicated that activities of topo I and topo II were not different in these cell lines, nor was there any difference in the formation of drug-induced KCl-SDS precipitable complexes, indicating that topoisomerases were not involved in the development of resistance in PC3(R) cells. While the activity of glutathione S-transferase and total glutathione levels were also similar in these cell lines, the glutathione peroxidase activity in PC3(R) cells was 5-fold lower than in PC3 cells. Furthermore, proto-oncogene expression for c-jun, c-myc, and H-ras was significantly higher in resistant cells than in sensitive cells, indicating that the amplification of early response genes may play a role in the emergence of de novo resistance in PC3(R) cells. PMID: 8155744 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 670: Jpn J Cancer Res. 1994 Apr;85(4):364-71. Proto-oncogene expression in a human chondrosarcoma cell line: HCS-2/8. Zhu JD, Pan HO, Suzuki F, Takigawa M. Chester Beatty Laboratories, London, UK. HCS-2/8 is a stable human chondrosarcoma cell line with many chondrocytic characteristics and has the capacity to form chondrosarcomas in nude mice. The cells display both biochemically and morphologically definable changes in sparse, subconfluent, confluent and over-confluent phases of in vitro culture. Such features of HCS-2/8 cells may reflect the processes of both proliferation and differentiation of chondrocytes in vivo. We examined the correlations of these changes of HCS-2/8 cells with their transcript levels of 21 proto-oncogenes by Northern analysis. We found no detectable transcripts of 9 proto-oncogenes (c-sis, c-met, c-src, c-lyn, c-fgr, c-ros, c-pim, Blym and N-myc), but detected transcripts of 12 other proto-oncogenes (int-2, erbB, c-abl, c-raf-1, c-fyn, K-ras, H-ras, c-mos, c-myc, c-myb, c-fos, and c-jun). In the over-confluent phase, the levels of c-fos and c-raf-1 were increased several dozen times and about 5 times, respectively, while the level of c-abl was about 1/5th of that in the sparse, subconfluent and confluent phases of culture. The level of int-2 increased about 10-fold in the confluent and over-confluent phases of in vitro culture. The transcript levels of c-mos and K-ras were high in the sparse phase, low in the subconfluent and confluent phases and high in the over-confluent phase. The levels of the other 6 proto-oncogenes in HCS-2/8 cells were constant in all phases of in vitro culture. PMID: 8200849 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 671: Mol Carcinog. 1994 Apr;9(4):210-9. In vitro and in vivo acceleration of the neoplastic phenotype of a low-tumorigenicity rat bladder carcinoma cell line by transfected transforming growth factor-alpha. Kawamata H, Kameyama S, Oyasu R. Department of Pathology, Northwestern University Medical School, Chicago, IL 60611. We conducted an experiment to determine whether expression of transforming growth factor-alpha (TGF-alpha) enhances tumorigenicity in a low-tumorigenicity rat bladder carcinoma cell line and whether it is sufficient to induce a tumorigenic phenotype in a nontumorigenic rat bladder cell line. D44c cells (which are nontumorigenic) were derived from a minute nodule from a bladder treated with N-methyl-N-nitrosourea (MNU); G1-200 cl-17 cells (which have low tumorigenicity) were isolated from D44c cells exposed to MNU in vitro. Neither cell line expressed TGF-alpha mRNA. The cells were cotransfected with pSV2neo and pSR alpha-rTGF-alpha. The latter plasmid contains the rat TGF-alpha cDNA under the transcriptional control of the SR alpha promoter. In the low-tumorigenicity G1-200 cl-17 cells, the expression of TGF-alpha mRNA and the subsequent synthesis of TGF-alpha protein activated epidermal growth factor receptors (EGFRs) and markedly enhanced tumorigenicity in nude mice (i.e., shortened the latency period before tumor appearance, accelerated the rate of growth, and increased the size of the tumors) as well as anchorage-independent growth in vitro. In nontumorigenic D44c cells, however, transfected TGF-alpha did not induce either anchorage-independent growth or tumorigenicity in nude mice, in spite of overexpression of EGFR mRNA and the constitutive expression of c-jun and junB mRNA. These results suggest that the increased signal transduction mediated by TGF-alpha enhanced tumorigenicity in a cell that was already tumorigenic but was not sufficient to induce tumorigenicity in a nontumorigenic cell. PMID: 8148054 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 672: Hepatology. 1994 Apr;19(4):962-72. Hepatocyte growth factor in transgenic mice: effects on hepatocyte growth, liver regeneration and gene expression. Shiota G, Wang TC, Nakamura T, Schmidt EV. Massachusetts General Hospital Cancer Center, Charlestown 02129. Attention has recently been focused on hepatocyte growth factor as a major candidate factor in liver regeneration because it is the most potent known mitogen for hepatocytes in vitro. However, hepatocyte growth factor also displays diverse activities in vitro as scatter factor, as an epithelial morphogen, as a pluripotent mitogen and as a growth inhibitor. Consequently, we developed transgenic mice that expressed hepatocyte growth factor under the control of albumin regulatory sequences to examine its in vivo role in hepatocyte growth. Hepatocytes of these mice expressed increased levels of hepatocyte growth factor as an autocrine growth factor. Hepatocyte growth factor was a potent stimulus for liver repair; the livers of hepatocyte growth factor-transgenic mice recovered completely in half the time needed for their normal siblings after partial hepatectomy. This transgenic model also enabled us to study the chronic effects of hepatocyte growth factor expression. During several months of observation, the labeling index of hepatocytes in albumin-hepatocyte growth factor mice was doubled, and liver DNA content was increased compared with that in wild-type mice. To identify intermediate signaling pathways for hepatocyte growth factor that might regulate this increased growth response, we examined transgenic mice for changes in expression of genes that are known to be regulated during liver regeneration. We found that levels of c-myc and c-jun mRNA were increased in the hepatocyte growth factor-transgenic mice. In additional experiments the increased c-myc expression was the consequence of increased transcription rates as seen in nuclear run-on and myc-CAT reporter gene experiments. We conclude that hepatocyte growth factor increases growth and repair processes when expressed for long periods in the liver and that c-myc and c-jun may be important intermediaries in the hepatocyte growth response caused by hepatocyte growth factor. PMID: 8138271 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 673: Semin Cancer Biol. 1994 Apr;5(2):157-65. Oncogenes and epithelial cell transformation. Reichmann E. Institute for Molecular Pathology, Vienna, Austria. More than 80% of the tumors occurring in man are carcinomas. Despite the prevalence of these epithelial tumors most in vitro studies of oncogenesis have employed mesenchymal rather than epithelial cells. As a result, a detailed understanding of the molecular mechanisms of carcinoma formation and an exact definition of the malignant transformed epithelial phenotype are lacking. Taking into account that the genesis of a carcinoma involves alterations in multiple genes, giving rise to complex phenotypic changes, the question arises whether each single, mutational step can be correlated with distinct alterations of the epithelial phenotype. To approach this question, oncogenes that are both critically positioned in growth factor receptor-driven signaling pathways and relevant in carcinoma development, such as ras, src, myc and fos, have been expressed in epithelial cells. The results indicate that epithelial cells transformed by oncogenes in vitro alter their gene expression programs, thereby losing certain features of cell polarity and cell adhesion. However, as with true tumor cells, these changes occur in distinct combinations, to different degrees and are influenced by the local environment of the cells. Publication Types: Review Review, Tutorial PMID: 8061331 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 674: J Submicrosc Cytol Pathol. 1994 Apr;26(2):147-62. Protooncogene, growth factor, growth factor receptor, and estrogen and progesterone receptor gene expression in the immature rat uterus after treatment with estrogen and tamoxifen. Bhattacharyya N, Ramsammy R, Eatman E, Hollis VW, Anderson WA. Department of Biology, Howard University, Washington, D.C. 20059. The mechanism(s) by which estrogen regulates cell growth in target cells and the cascade of biochemical changes associated with growth have not yet been fully determined. Equally undetermined is an understanding of the mechanism(s) by which tamoxifen blocks estrogen-regulated growth. This study, therefore, attempts to define and correlate the physiological processes in the rat uterus following estrogen and tamoxifen administration with temporal events manifested by mRNA expression of protooncogenes (m-myc, c-ras, c-fos, c-jun), growth factors and/or inhibins (IGF-1, IGF-2, IGF-2 Exon 1 and Exon 2), EGF, TGFB-1, -2, -3), growth factor or inhibin receptors (EGFr, TGFB-2r, TGFB-3r), and estrogen-induced differentiative proteins including estrogen receptor (ER), progesterone receptor (PR). In this study, mRNA was isolated from hormone and antagonist-treated rat uteri at 0', 15', 30', 1 h, 6 h, 24 h, 48 h and 72 h. Expression studies were analysed by dot/Northern blot hybridization with cDNA or oligonucleotide probes for the RNAs mentioned above. Nuclear runoff transcriptional assays were also performed. Our data suggest that fos, myc, ras and jun protooncogenes were expressed from 15' to 48 h after treatment with either estrogen or tamoxifen. Tamoxifen treatment resulted in diminished expression, but incomplete inhibition of the protooncogene mRNAs. Estrogen treatment resulted in rapid elevation of both EGF and EGFr mRNA levels, both of which were suppressed after tamoxifen treatment. Tamoxifen may exert its antiestrogenic effects by inhibiting EGF and EGFr, and myc protooncogene activity on the one hand, and by overexpression of the TGFB isotypes and their receptors on the other hand. With the proliferation cycle short circuited, tamoxifen-treated cells hypertrophied and differentiated to terminal cells by 24-48 h. Working hypotheses for the mechanisms of action of estrogen and tamoxifen are presented based on our data. PMID: 8019941 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 675: J Cell Biochem. 1994 Apr;54(4):432-9. Control of fibroblast senescence and activation of programmed cell death. Wang E, Lee MJ, Pandey S. Bloomfield Center for Research in Aging, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis, Jewish General Hospital, Montreal, Quebec, Canada. We have characterized a nuclear phosphoprotein of 57 kda, statin, found only in nonproliferating cells of both quiescent and senescent natures. Emerging results suggest that statin may function as a sequester to block the early G1 phase phosphorylation for the RB protein. A second protein, terminin, undergoes senescence-specific posttranslational modification from 90 to 60 kda, and further death-specific conversion from 60 to 30 kda. We also found that apoptotic mouse 3T3 fibroblasts express c-fos, c-myc, c-jun, and cdc2, as well as the upregulation of RB phosphorylation and BrdU incorporation, just before final DNA fragmentation and death. It seems that en route to death, cells re-enter the cell-cycle transverse and experience early G1 and part of S Phase; however, this cycling event is an abortive one. In contrast, senescent fibroblasts are resistant to the initiation of the death program, since they are unable to enter cell cycle traverse. Long-term serial passaging of normal human fibroblasts may be inadvertently selecting those, while termed as senescent, are also specialized survivors, and thus a good culture model to study both the control of permanent departure from cell cycle traverse and the mechanism underlying the survival or antideath cellular program. Publication Types: Review Review, Tutorial PMID: 8014192 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 676: Neurochem Res. 1994 Apr;19(4):489-99. Glutamate receptor-driven activation of transcription factors in primary neuronal cultures. Condorelli DF, Dell'Albani P, Amico C, Lukasiuk K, Kaczmarek L, Giuffrida-Stella AM. Institute of Biochemistry, Faculty of Medicine, University of Catania, Italy. We have used primary neuronal cultures prepared from fetal cerebral hemispheres to investigate the effects of different glutamate receptor agonists and antagonists on the expression of transcription factor encoding genes, such as c-fos, fosB, c-jun, junB, junD, c-myc, and zif/268. The addition of glutamate (100 microM) to the culture medium rapidly activated c-fos, fosB, c-jun, junB and zif/268 gene expression, reaching the maximal level at 30-60 minutes for zif/268 and at 60 minutes for the other genes. The onset of fosB mRNA accumulation was slightly delayed in comparison to the other genes. No clear induction was found for junD and c-myc. Different glutamate receptor agonists, such as NMDA, kainate, quisqualate, trans-(+/-)-1-aminocyclopentane-1,3-dicarboxylic acid (t-ACPD) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) were able to increase c-fos, c-jun, and zif-268 mRNA levels with rapid and transient kinetics similar to those observed after glutamate treatment. Similar results were obtained for junB and fosB after kainate and quisqualate stimulation. Pretreatment with MK-801, a non competitive NMDA antagonist, produced an almost complete inhibition of glutamate-driven expression of transcription factor genes, thus suggesting that NMDA receptor plays a major role in glutamate induced-gene expression. On the contrary the kainate/AMPA receptor antagonist, DNQX, did not influence glutamate induced-gene expression. Under the conditions used in the present study, NMDA was effective in inducing the simultaneous activation of several IEGs even when added to the culture medium containing millimolar concentration of magnesium. When experiments were performed in Krebs solution, NMDA was effective in stimulating zif/268 and c-fos mRNAs only in the absence of Mg2+, while glutamate activated c-fos and zif/268 both in the presence and absence of magnesium ions. As expected, NMDA effect was fully inhibited by MK-801. The level of AP-1 DNA binding activity, as measured by electrophoretic mobility shift assay, increased after addition of glutamate and NMDA to cultured neurons and such increase was antagonized by the pretreatment with MK-801. PMID: 7520539 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 677: Int J Cancer. 1994 Apr 1;57(1):98-103. Comparison of retinoic acid and phorbol myristate acetate as inducers of monocytic differentiation. Matikainen S, Hurme M. Department of Bacteriology and Immunology, University of Helsinki, Finland. Several human myeloid leukemia cell lines growing in vitro can be induced to differentiate to more mature monocyte/macrophage-like cells by treatment with protein kinase C-activating phorbol esters, such as PMA. In addition to PMA, cells of the THP-1 myeloid leukemia cell line acquire macrophage-like characteristics after treatment with all-trans retinoic acid (RA). To analyze the signal transduction mechanisms induced by RA, we first compared the effects of PMA and RA on the expression of genes which are known to be regulated during monocytic differentiation. Both RA and PMA effectively down-regulated c-myc expression, while c-myb expression decreased only after PMA treatment. Expression of the beta 2-integrin genes, CD11a and CD11b, was clearly increased after both of these treatments. Their effects on the src-family tyrosine kinase genes were different: hck expression was similarly induced by these agents but lyn expression was stronger and more rapid after RA treatment. RA also enhanced lyn mRNA production rapidly in HL-60, indicating that the activation of lyn gene expression is common in monocytic and granulocytic maturation of myeloid leukemia cells. To examine whether the AP-1 enhancer activity is involved in RA-induced monocytic differentiation, THP-1 cells were transiently transfected with a chloramphenicol acetyl transferase (CAT)-reporter gene containing 5 copies of the AP-1 binding sites. In contrast to PMA, RA did not induce any CAT activity in these cells, thus suggesting that the RA-induced changes in the expression of those genes described above were not dependent on the AP-1 enhancer activity. PMID: 7512079 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 678: Biochem Biophys Res Commun. 1994 Mar 15;199(2):977-83. Effect of erythropoietin on DNA synthesis, proto-oncogene expression and phospholipase C activity in rat vascular smooth muscle cells. Gogusev J, Zhu DL, Herembert T, Ammarguellat F, Marche P, Drueke T. INSERM U 90, Hopital Necker, Paris, France. The administration of recombinant human erythropoietin (rHuEpo) to anemic chronic renal failure patients may be associated with an increase in blood pressure, possibly by direct effects on peripheral blood vessels. The experiments of the present study were designed to explore the hypothesis that rHuEpo might exert mitogenic effects on vascular smooth muscle cells (VSMCs), and that pre-existing hypertension might be a predisposing condition. Cultured aortic VSMCs from spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats were studied for DNA synthesis, phospholipase C activity, and cell growth related proto-oncogene expression in the presence of rHuEpo. In cells from both rat strains, rHuEpo dose-dependently increased DNA synthesis and stimulated phospholipase C activity, as indicated by 3H-thymidine incorporation and inositol phosphate formation, respectively. Exposure of VSMCs to rHuEpo for various periods gradually increased the levels of c-myc and JunB mRNAs and transiently induced c-fos mRNA expression as determined by Northern analysis. The hormone-induced DNA synthesis was markedly enhanced in VSMCs from SHR compared to those from WKY. In contrast, rHuEpo-induced phospholipase C activity and proto-oncogene expression did not differ between the two strains. Taken together, these results suggest that rHuEpo may function as a vascular smooth muscle cell growth promoting factor through activation of the phospholipase C cascade and a modulation of proto-oncogene expression. It could thereby contribute to vascular hypertrophy and arterial hypertension. PMID: 8135847 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 679: J Cell Biochem. 1994 Mar;54(3):265-72. Cell cycle arrest by prostaglandin A1 at the G1/S phase interface with up-regulation of oncogenes in S-49 cyc- cells. Hughes-Fulford M. Research Service, Veterans Administration Medical Center, San Francisco, California 94121. Our previous studies have implied that prostaglandins inhibit cell growth independent of cAMP. Recent reports, however, have suggested that prostaglandin arrest of the cell cycle may be mediated through protein kinase A. In this report, in order to eliminate the role of c-AMP in prostaglandin mediated cell cycle arrest, we use the -49 lymphoma variant (cyc-) cells that lack adenylate cyclase activity. We demonstrate that dimethyl prostaglandin A1 (dmPGA1) inhibits DNA synthesis and cell growth in cyc- cells. DNA synthesis is inhibited 42% by dmPGA1 (50 microM) despite the fact that this cell line lacks cellular components needed for cAMP generation. The ability to decrease DNA synthesis depends upon the specific prostaglandin structure with the most effective form possessing the alpha, beta unsaturated ketone ring. Dimethyl PGA1 is most effective in inhibiting DNA synthesis in cyc- cells, with prostaglandins PGE1 and PGB1 being less potent inhibitors of DNA synthesis. DmPGE2 caused a significant stimulation of DNA synthesis. S-49 cyc- variant cells exposed to (30-50 microns) dmPGA1, arrested in the G1 phase of the cell cycle within 24 h. This growth arrest was reversed when the prostaglandin was removed from the cultured cells; growth resumed within hours showing that this treatment is not toxic. The S-49 cyc- cells were chosen not only for their lack of adenylate cyclase activity, but also because their cell cycle has been extensively studied and time requirements for G1, S, G2, and M phases are known. Within hours after prostaglandin removal the cells resume active DNA synthesis, and cell number doubles within 15 h suggesting rapid entry into S-phase DNA synthesis from the G1 cell cycle block.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8200906 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 680: Tanpakushitsu Kakusan Koso. 1994 Mar;39(3):287-90. [Cytokine signal transduction and expression of cell cycle related genes] [Article in Japanese] Shibuya H, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. Publication Types: Review Review, Tutorial PMID: 8153360 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 681: Cancer Res. 1994 Mar 1;54(5):1374-80. Photodynamic therapy mediated induction of early response genes. Luna MC, Wong S, Gomer CJ. Clayton Ocular Oncology Center, Childrens Hospital Los Angeles, California 90027. Photodynamic therapy (PDT) generates reactive oxygen species which initiate the cytotoxic events of this tumor treatment. We demonstrate that PDT mediated oxidative stress induced a transient increase in the early response genes c-fos, c-jun, c-myc, and egr-1 in murine radiation-induced fibrosarcoma cells. Incubation of exponentially growing cells with porphyrin based photosensitizers in the dark also induced an increase in mRNA levels of early response genes. However, the xanthine photosensitizer, rose bengal, produced increased c-fos mRNA levels only following light treatment. Nuclear runoff experiments confirmed that the induction of c-fos mRNA is controlled in part at the level of transcription. Likewise, a chloramphenicol acetyltransferase reporter construct containing the major c-fos transcriptional response elements was inducible by porphyrin and PDT. Signal transduction pathways associated with PDT mediated c-fos activation were examined by treating cells with protein kinase inhibitors. Staurosporine and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine inhibited PDT mediated c-fos activation while N-(2-guanidinoethyl)-5-isoquinoline-sulfonamide had no effect. In addition, quinacrine, which can inhibit phospholipase activity, blocked PDT induced c-fos mRNA expression. These results suggest that photosensitizer mediated oxidative stress acts through protein kinase-mediated signal transduction pathway(s) to activate early response genes. PMID: 8118827 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 682: Mol Biol Cell. 1994 Mar;5(3):375-88. Cloning of mid-G1 serum response genes and identification of a subset regulated by conditional myc expression. Tavtigian SV, Zabludoff SD, Wold BJ. Myriad Genetics, Salt Lake City, Utah 84108. The emergence of cells from a quiescent G0 arrested state into the cell cycle is a multistep process that begins with the immediate early response to mitogens and extends into a specialized G1 phase. Many immediate early serum response genes including c-fos, c-myc, and c-jun are transcriptional regulators. To understand their roles in regulating cell cycle entry and progression, the identities of their regulatory targets must be determined. In this work we have cloned cDNA copies of messenger RNAs that are either up- or down-regulated at a mid-G1 point in the serum response (midserum-response [mid-SR]). The mid-SR panel is expected to include both direct and indirect targets of immediate early regulators. This expectation was confirmed by the identification of several transcriptional targets of conditional c-myc activity. In terms of cellular function, the mid-SR class is also expected to include execution genes needed for progression through G1 and into S-phase. DNA sequence data showed that the mid-SR panel included several genes already known to be involved in cell cycle progression or growth transformation, suggesting that previously unknown cDNAs in the same group are good candidates for other G1 execution functions. In functional assays of G0-->S-phase progression, c-myc expression can bypass the requirement for serum mitogens and drive a large fraction of G0 arrested cells through G1 into S-phase. However, beyond this general similarity, little is known about the relation of a serum-driven progression to a myc-driven progression. Using the mid-SR collection as molecular reporters, we found that the myc driven G1 differs qualitatively from the serum driven case. Instead of simply activating a subset of serum response genes, as might be expected, myc regulated some genes inversely relative to serum stimulation. This suggests that a myc driven progression from G0 may have novel properties with implications for its action in oncogenesis. PMID: 8049528 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 683: Arkh Patol. 1994 Mar-Apr;56(2):16-21. [Expression and co-expression of cellular oncogenes in the course of tumor progression in neuroendocrine lung tumors] [Article in Russian] Kogan EA, Shtabskii AB, Sekamova SM, Sukhova NM, Mazurenko NN. The tumours studied were as follows: benign carcinoids, well differentiated carcinomas (atypical carcinoids) and poorly differentiated tumours (small-cell carcinoma with neuroendocrine differentiation). Oncoproteins c-myc, c-fos, c-jun, L-myc, c-sis, c-ras, c-src, c-ets-1, c-met were studied immunohistochemically in the material obtained from 25 patients during the operations. A higher expression of c-fos, c-jun, c-ets-1 and c-met is observed at early stages of progression and that of c-myc and L-myc at later stages. Enhancement of c-myc and L-myc expression correlated with the invasiveness and lymphogenic metastasizing. Co-expression and negative correlation between some oncogenes are established. The data obtained may be used for prognosis and therapy. PMID: 8037585 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 684: Hum Cell. 1994 Mar;7(1):1-5. [Structure and function of IL-2 receptor subunits] [Article in Japanese] Sugamura K. Department of Bacteriology, Tohoku University School of Medicine, Sendai, Japan. IL-2 regulates growth and differentiation of various types of cells in the immune system via its interaction with IL-2 receptor (IL-2R). The high and intermediate-affinity IL-2Rs, which consist of the alpha beta gamma heterotrimer complex and the beta gamma heterodimer complex, respectively, harbor the function of the intracellular signal transduction, indicating that the beta and gamma chains are indispensable for the signal transduction but not the alpha chain. The reconstitution studies of IL-2Rs with alpha, beta and gamma chain genes demonstrated that each subunit has potential for altering the affinity of the receptor, and the cytoplasmic domains of the beta and gamma chains participate in signal transduction in terms of cell growth, activation of alpha tyrosine kinase and enhancement of c-myc, c-fos and c-jun transcription. The region containing the SH2 homologous sequence of the gamma chain should have a critical function for signal transduction. On the other hand, common subunits are known to be shared among receptors for IL-3, IL-5 and GM-CSF, and receptors for IL-6, LIF, OSM and LIF. We have demonstrated that the monoclonal antibody specific for the IL-2R gamma chain completely inhibited not only IL-2-dependent cell growth but also IL-4-dependent, IL-7-dependent, and IL-9-dependent cell growth, suggesting that the gamma chain is possibly shared among receptors for IL-2, IL-4, IL-7 and IL-9. Impairment of the gamma chain function is considered to be closely related to human XSCID characterized by profound T cell defect.(ABSTRACT TRUNCATED AT 250 WORDS) Publication Types: Review Review, Tutorial PMID: 8025015 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 685: J Neurosci Res. 1994 Feb 15;37(3):303-12. Protein synthesis and mRNA in isolated growth cones from differentiating SH-SY5Y neuroblastoma cells. Meyerson G, Parrow V, Gestblom C, Johansson I, Pahlman S. Department of Pathology, University of Uppsala, University Hospital, Sweden. The human neuroblastoma cell line, SH-SY5Y, differentiates into a neuronal, sympathetic phenotype in the presence of phorbol ester and serum. Growth cones prepared from differentiating SH-SY5Y cells have characteristics similar to those of growth cones from embryonic rat brain. In addition, SH-SY5Y growth cones contain ribosomes. In this study we show, by metabolic labeling of isolated growth cones, that local protein synthesis occurred in these structures. The pattern of labeled proteins was very similar to that of the corresponding cell body fraction. RNA was shown to be transported to the growth cone compartment, and by in situ hybridization. beta-actin mRNA could be visualized in intact neuritic growth cones. Comparison by Northern blot hybridizations of RNA prepared from growth cones and cell bodies, respectively, showed that mRNAs coding for growth-associated protein 43, microtubule-associated protein 2, actin, neuropeptide tyrosine, and glyceraldehyde-3-phosphate dehydrogenase were present in both fractions. In contrast, mRNAs coding for the nuclear proteins c-jun and N-myc were virtually absent in the growth cone, but readily detectable in the cell body preparation. The selective distribution of mRNAs to the growth cones was not restricted to stable, abundant mRNA species, since mRNA coding for the insulin-like growth factor I receptor was stable, but not present in growth cones. Thus, differentiating SH-SY5Y cells can sort and transport RNA to the growth cone compartment, suggesting that this system of clonal cells could be useful to unravel mechanisms involved in the compartmentalization of mRNA. PMID: 8176754 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 686: Hepatology. 1994 Feb;19(2):489-97. In vivo response of hepatocytes to growth factors requires an initial priming stimulus. Webber EM, Godowski PJ, Fausto N. Department of Pathology and Laboratory Medicine, Brown University School of Medicine, Providence, Rhode Island 02912. Although growth factor effects have been studied in cultured hepatocytes, little information exists as to whether these factors can trigger hepatocyte replication in vivo. In this study we infused epidermal growth factor, transforming growth factor-alpha and hepatocyte growth factor directly into the portal vein of rats for 24 hr to see whether they could induce DNA synthesis in normal livers or in livers subjected to one-third hepatectomy. Infusion of transforming growth factor-alpha or epidermal growth factor at doses up to 80 micrograms/24 hr had little effect on hepatic DNA synthesis in normal liver, whereas the monomeric and heterodimeric forms of hepatocyte growth factor generally produced increases of less than threefold in hepatic DNA synthesis. In contrast, after one-third hepatectomy infusion of epidermal growth factor, transforming growth factor-alpha or hepatocyte growth factor produced dose-dependent increases in hepatic DNA synthesis. At a dose of 40 micrograms/24 hr, epidermal growth factor increased DNA synthesis threefold, whereas transforming growth factor-alpha or hepatocyte growth factor increased DNA synthesis to greater than six times that in rats that had undergone hepatectomy alone. Furthermore, infusion of these growth factors, with or without one third-hepatectomy, induced the expression of transforming growth factor-alpha mRNA in the liver. The pattern of protooncogene expression induced by one-third hepatectomy was studied to determine the effect of this procedure in sensitizing the liver to the growth factors. Compared with the well-characterized two-thirds hepatectomy system, there was a similar but smaller increase in c-myc expression but no induction of c-jun expression.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8294105 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 687: Semin Diagn Pathol. 1994 Feb;11(1):3-14. The molecular basis of skeletal muscle differentiation. Dias P, Dilling M, Houghton P. Department of Molecular Pharmacology, St Jude Children's Research Hospital, Memphis, TN 38101. In recent years, significant advances have been made in understanding the molecular mechanisms involved in the regulation of skeletal-muscle differentiation. This review focuses on the role of the MyoD family of myogenic transcription factors that includes MyoD, myf-5, myogenin, and MRF4 (herculin or myf-6) in myogenesis. Members of this family share sequence homology for the basic-helix-loop-helix (bHLH) regulatory motif. The basic domain is required for DNA binding, whereas the HLH domain is required for dimerization. The bHLH motif confers both properties of transcriptional activation of muscle specific genes and inhibition of cell growth through collaboration with the E2A gene products (E12 and E47) and the retinoblastoma gene product (pRB). The functions of the MyoD family can be suppressed through inhibition of their expression or activity by various factors. These include peptide growth factors (FGF and TGF-beta), immediate early gene products (Fos, Jun and Myc), other oncogene products (Ras, Src), the Id protein, and innervation. The use of gene-knockout animal models has shown that there is some degree of functional redundancy in that inactivation of either MyoD or myf-5 has no effect on muscle development, whereas inactivation of both genes results in an absolute lack of muscle cells. In contrast, the inactivation of myogenin alone results in mice with gross deficiency of mature muscle. Publication Types: Review PMID: 8202645 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 688: J Biol Chem. 1994 Jan 21;269(3):1599-602. The adapter protein Shc interacts with the interleukin-2 (IL-2) receptor upon IL-2 stimulation. Ravichandran KS, Burakoff SJ. Division of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115. Binding of interleukin-2 (IL-2) to the IL-2 receptor (IL-2R) stimulates Src family kinases, tyrosine phosphorylation of several proteins, conversion of Ras to its active GTP-bound form, and eventually c-fos, c-jun, and c-myc induction. The IL-2R beta chain plays a crucial role in IL-2R signaling. Within the cytoplasmic domain of the beta chain, a region essential for mitogenesis and another involved in binding the Src family kinase Lck have been defined. The beta chain itself is tyrosine-phosphorylated upon IL-2 stimulation. Since the adapter protein Shc acts upstream of Ras and is involved in T cell receptor-mediated Ras activation, we examined the role of Shc in IL-2 signaling. Shc was found to be tyrosine-phosphorylated upon IL-2 stimulation in CTLL-20 cells. After its phosphorylation, Shc interacted with another adapter protein, Grb2, and, via Grb2, with the Ras GTP/GDP exchange factor mSOS. After IL-2 stimulation, Shc also associated with the IL-2R beta chain. Thus, during IL-2 signaling, the interaction of Shc with the IL-2R beta chain and its simultaneous association with Grb2 and mSOS may couple IL-2R stimulation to Ras signaling. PMID: 8294403 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 689: Cancer Res. 1994 Jan 15;54(2):592-7. Inhibition by interleukin 4 of leukemia inhibitory factor-, interleukin 6-, and dexamethasone-induced differentiation of mouse myeloid leukemia cells: role of c-myc and junB proto-oncogenes. Kasukabe T, Okabe-Kado J, Hozumi M, Honma Y. Department of Chemotherapy, Saitama Cancer Center Research Institute, Japan. Interleukin 4 (IL-4) inhibited the differentiation of mouse myeloid leukemia M1 cells induced by leukemia inhibitory factor (LIF), interleukin 6, or dexamethasone and conversely enhanced the induction of M1 cell differentiation by 1 alpha,25-dihydroxyvitamin D3. IL-4 blocked LIF-induced differentiation of M1 cells when it was added to the culture medium within 10 h after LIF, but IL-4 did not block differentiation when it was added 12 h after LIF. These results indicate that IL-4 inhibited a critical intermediate step in myeloid leukemia cell differentiation. LIF markedly stimulated the expression of junB mRNA within 2 h but suppressed the expression of c-myb and c-myc after 2- and 12-h treatment, respectively. IL-4 did not significantly affect LIF-induced junB expression or suppression of c-myb expression. However, it interfered significantly with the LIF-induced suppression of c-myc gene expression. Similar results were obtained when interleukin 6 was used to induce differentiation of M1 cells. Dexamethasone and 1 alpha,25-dihydroxyvitamin D3 did not induce junB gene expression but suppressed the expression of c-myb and c-myc. IL-4 also interfered with dexamethasone-induced suppression of c-myc gene expression. On the other hand, IL-4 enhanced 1 alpha,25-dihydroxyvitamin D3-induced down-regulation of c-myc gene expression, consistent with its enhancement of differentiation. These results indicate that the change in c-myc expression induced by IL-4 in M1 cells is closely associated with the effect of IL-4 on the induction of differentiation of M1 cells. PMID: 8275499 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 690: Am J Surg. 1994 Jan;167(1):14-9; discussion 19-20. Transforming growth factor-beta inhibits rat intestinal cell growth by regulating cell cycle specific gene expression. Ko TC, Beauchamp RD, Townsend CM Jr, Thompson EA, Thompson JC. Department of Surgery, University of Texas Medical Branch Galveston 77555-0533. Transforming growth factor-beta 1 (TGF-beta 1) inhibits the growth of intestinal cells, but the mechanisms involved are unknown. Using a rat intestinal crypt cell line (IEC-6), we determined the site of action in the cell cycle that TGF-beta 1 acts to suppress proliferation. We also examined the effect of TGF-beta 1 on the expression of proliferation-associated "immediate early" genes (zif268, jun-B, c-myc) during the early G1 phase and the cdc2 gene during the transition from the G1 phase to the S phase of the cell cycle. Cell cycle progression was determined by incorporation of 3H-thymidine, and gene expression was analyzed by Northern blot analysis. We found that TGF-beta 1 acts to inhibit proliferation of rat intestinal crypt cells by blocking cell cycle progression at the middle G1 phase. The genes activated during G1 can be divided into TGF-beta 1 insensitive (zif268, jun-B, and c-myc) and TGF-beta 1 sensitive (the cdc2 gene). TGF-beta 1 suppresses the induction of the cdc2 gene during the G1/S transition without inhibiting the activation of immediate early genes during the early G1 phase. PMID: 8311125 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 691: Clin Endocrinol (Oxf). 1994 Jan;40(1):117-26. Oncoprotein immunoreactivity in human pituitary tumours. Raghavan R, Harrison D, Ince PG, James RA, Daniels M, Birch P, Caldwell GI, Kendall-Taylor P. Department of Neuropathology, Newcastle General Hospital, Newcastle upon Tyne, UK. OBJECTIVE AND DESIGN: The immediate early gene locus AP-1, incorporating the cellular oncogenes c-fos and c-jun (and their oncoprotein products Fos and Jun respectively) play a key role in regulating cell growth and differentiation. The myc-gene is also known to promote cell growth. In order to investigate the possible role of these oncogenes in human pituitary adenomas, Fos, Jun and Myc oncoprotein immunoreactivities were assessed in surgically resected pituitary adenomas in relation to in-vivo characteristics (hormone secretion, size and invasiveness) and an in-vitro index of cell proliferation (Ki-67 immunoreactivity). Thirty-three human pituitary adenomas and 16 normal pituitary glands were examined. MEASUREMENTS: Oncoprotein immunoreactivity was recorded as present (+) or absent (-), and Ki-67 labelling indices were scored quantitatively. Tumour size was scored from CT scan appearances and radiographic evidence of bone erosion was noted. RESULTS: Oncoprotein immunoreactivity was present in a total of 32/33 cases. Myc immunoreactivity was restricted to the only ACTH-secreting tumour in the series (1/33). Ki-67 immunoreactivity was present in 24/32 cases and labelling indices varied from 0.1 to 3.2%. CONCLUSIONS: Oncoprotein immunoreactivity did not correlate with hormonal profile, bone erosion or the size of the proliferating compartment estimated by Ki-67 labelling indices. Although oncoprotein expression is common in human pituitary adenomas, its significance remains to be elucidated. PMID: 8306470 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 692: Hepatology. 1994 Jan;19(1):23-31. The hepatitis B virus preS2/St transactivator utilizes AP-1 and other transcription factors for transactivation. Lauer U, Weiss L, Lipp M, Hofschneider PH, Kekule AS. Max-Planck-Institut fur Biochemie, Martinsried, Germany. Integrated hepatitis B virus DNA cloned from hepatitis B virus-associated hepatocellular carcinoma frequently contains 3'-truncated middle surface genes (preS2/St), which were recently found to have a transcriptional transactivator function. Because preS2/St, among others, is able to transactivate the promoters of the cellular oncogenes c-myc and c-fos, it has been speculated that integrated preS2/St genes might contribute to hepatitis B virus-associated liver carcinogenesis. In this study, we investigated the mechanism of target gene stimulation by preS2/St. It was found that deletion of a fragment containing the binding site for transcription factor AP-1 (Jun-Fos) substantially decreases inducibility of the human c-myc promoter by preS2/St. A subsequent investigation of AP-1 activation by preS2/St revealed the following: (a) insertion of multimeric AP-1 binding sites confers inducibility to an otherwise unstimulatable test promoter; (b) transactivation of AP-1 sites is dramatically increased when Jun and Fos are overexpressed by cotransfected expression plasmids; and (c) inhibitors of AP-1 activation also impair transactivation by preS2/St. Besides AP-1, preS2/St was also able to utilize the unrelated transcription factors NF-kappa B and AP-2 for transactivation, suggesting that the gene product of preS2/St acts indirectly through one or several general cellular pathways rather than as a bona fide transcription factor. Because AP-1 conveys induction of a large panel of tumor-relevant genes, its preS2/St-dependent activation implies a possible causative role in hepatitis B virus-associated hepatocarcinogenesis. PMID: 8276360 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 693: Annu Rev Med. 1994;45:297-323. Molecular events in the pathogenesis of hepadnavirus-associated hepatocellular carcinoma. Robinson WS. Stanford University School of Medicine, California 94305. Chronic hepadnavirus infection is associated with hepatocellular carcinoma (HCC) in natural hosts such as humans, woodchucks, and Beechey ground squirrels. Several possible oncogenic mechanisms have been identified, including a potential role of the hepadnavirus x (hbx) gene, which transactivates transcription regulated by certain cis-acting sequences, e.g. regulatory sequences of the hepatitis B virus (HBV) and heterologous regulatory sequences of other viruses and cellular genes. The oncogenic potential of hbx is suggested by the observation of HCCs in hbx transgenic mice, the oncogenic transformation of cells expressing hbx in culture, and the transactivation of oncogenes c-myc and c-jun by hbx. Cis-activation of cellular oncogenes N-myc and c-myc by viral promoter insertion has been a common finding in woodchuck hepatitis virus (WHV)-associated HCCs of woodchucks. No such cis-activation of any cellular gene has been shown in virus-associated HCCs of ground squirrels or humans. Amplification and overexpression of the c-myc gene has been a common finding in HCCs of ground squirrels, and is rare in woodchuck or human HCCs. Point mutations in the p53 gene and allelic deletion of p53 have been common findings in human HCCs, but have not been found in HCCs in woodchucks and have been found rarely in ground squirrels. How each of these genetic changes in the different hosts contributes to HCC remains to be determined, but apparently different changes in different HCCs of hepadnavirus-infected hosts suggest that several separate genetic events may contribute to the development of HCC. These events may differ in each host, and some may not result from a direct virus-specific mechanism. Chronic hepadnavirus infection is often associated with chronic necroinflammatory liver disease and cirrhosis, a pathologic process common to several other risk factors for HCC. This suggests that this pathologic process (necroinflammatory disease) may be hepatocarcinogenic regardless of the inciting agent. Thus hepadnavirus infection may play an important role in the development of HCC by causing chronic hepatitis and HCC with the same mechanisms by which other risk factors for HCC cause chronic necroinflammatory liver disease and HCC. Publication Types: Review Review, Tutorial PMID: 8198385 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 694: Tumour Biol. 1994;15(6):311-7. Tissue differences in the expression of mRNAs of Ha-ras, c-myc, fos and jun in human uterine endometrium, myometrium and leiomyoma under the influence of estrogen/progesterone. Fujimoto J, Hori M, Ichigo S, Nishigaki M, Tamaya T. Department of Obstetrics and Gynecology, Gifu University School of Medicine, Japan. Ha-ras expression level in uterine endometrium (EM) in the proliferative phase (PP) was significantly higher than that in the secretory phase (SP). c-myc expressions were detected in EM, uterine myometrium (MM) and uterine leiomyoma (LM) without any cyclic change; fos expressions in LM, MM, and EM were detectable in PP, but not in SP. jun expression level in LM was significantly higher than that in MM and EM in PP, but did not alter during the menstrual cycle. Estrogen elevated the levels of Ha-ras and fos mRNAs in the three tissues, jun mRNA in MM and EM, and c-myc mRNA in LM and MM. These results suggest that there is a tissue difference in the expression of Ha-ras, c-myc, fos and jun among EM, MM and LM, under the influence of estrogen/progesterone. PMID: 7997802 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 695: Crit Rev Eukaryot Gene Expr. 1994;4(1):55-116. Steroid hormone-induced expression of oncogene encoded nuclear proteins. Hyder SM, Stancel GM, Loose-Mitchell DS. Department of Pharmacology, University of Texas, Houston Health Science Center, Medical School 77225. In this article we have attempted to review the literature on the regulation of nuclear protooncogene expression by steroid hormones and other small molecules that interact with receptors of the steroid/thyroid superfamily. Until about 5 years ago, there were relatively few reports of steroidal regulation of cellular oncogenes, but hundreds of papers on this topic have appeared since then. This demonstrates the intense interest in this area that has developed recently. It now been demonstrated that all the major classes of steroid hormones control expression of nuclear protooncogenes in one or more systems. Given the actions of these proteins as transcription factors and their central role in cellular communications systems, it seems likely that they play a key role in mediating the biological effects of steroids on processes such as proliferation and differentiation. To date, most of the work in this general area has focused primarily on the regulation of three genes: c-fos, c-jun, and c-myc. However, a quick glance at the table of nuclear protooncogenes in the introduction of this article indicates that over 40 nuclear protooncogenes are now recognized. For the large majority of these, regulatory effects of steroids and related molecules have not yet been reported. Hence, we predict that reports in this general area of research will continue to appear at a very rapid rate over the next few years. In addition, we have tried to provide enough background information for readers to get an overview of the regulation of nuclear protooncogene expression by nonsteroidal factors. We felt this information was important to emphasize that steroid hormones represent only one of the many classes of regulatory molecules that control expression of nuclear protooncogenes. Thus, an important area for future research will be to understand how these multiple regulatory systems interact to control expression of this important class of cellular oncogenes and the biological processes that they mediate. Publication Types: Review PMID: 7987047 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 696: Growth Factors. 1994;10(3):171-80. Butyrate synchronization of hepatocytes: modulation of cycling and cell cycle regulated gene expression. Gupta S, Alpini G, Vemuru RP, Hurston E, Shafritz DA. Marion Bessin Liver Research Center, Department of Medicine, Albert Einstein College of Medicine, Bronx, NY 10461. To develop a model for studies of liver growth control, we characterized cell cycle synchronization of liver-derived cells with sodium butyrate. Exposure of cultured HTC (rat hepatoma) cells to 5 mM butyrate arrested cell growth in a reversible manner. Flow cytometric analysis revealed that butyrate-treated HTC cells were restricted in G0/G1, as well as S/G2M phases. After release from butyrate arrest, HTC cells underwent synchronous cycles of DNA synthesis and transited through S phase. Inhibition of cell growth by butyrate was associated with a complex pattern of cell cycle regulated gene expression, including a decoupling of c-fos and c-jun gene expression. Transcription of c-fos, as well as c-jun increased with butyrate arrest, whereas steady rate mRNA levels of c-jun only were increased, suggesting additional regulation of c-fos. In addition, butyrate-arrested cells exhibited a transcriptionally determined accumulation of H3 histone, C-Ha-ras and ornithine decarboxylase mRNAs, suggesting that cell cycle-related check points following the onset of S phase were modulated. An increase in c-myc mRNA levels in butyrate-arrested cells was post-transcriptionally regulated. After release from butyrate-arrest, the abundance of immediate early, as well as S phase regulated, gene expression changed coordinately with S phase cell transitions. Thus, exposure of HTC cells to butyrate modulates cell cycle regulated gene expression, inhibits cycling, and results in accumulation of cells in specific compartments. Synchronization of liver cells with butyrate should, therefore, provide a useful model for defining cell cycle-related events in response to various mitogenic stimuli. PMID: 7946406 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 697: J Cell Sci Suppl. 1994;18:105-8. Signal transduction by the macrophage-colony-stimulating factor receptor (CSF-1R). Roussel MF. St Jude Children's Research Hospital, Memphis, Tennessee. The macrophage-specific colony-stimulating factor 1 (CSF-1 or M-CSF) is required throughout the G1 phase of the cell cycle to regulate both immediate and delayed early responses necessary for cell proliferation. These are triggered by the binding of the growth factor to the colony-stimulating factor 1 receptor and the activation of its intrinsic tyrosine-specific protein kinase. Phosphorylation of the colony-stimulating factor 1 receptor on specific tyrosine residues enables it to bind directly to cytoplasmic effector proteins, which in turn relay receptor-induced signals through multiple-signal transduction pathways. The activity of p21ras as well as transcription factors of the ets gene family appears to be required for colony-stimulating factor 1 to induce the c-myc gene, and the latter response is essential to ensure cell proliferation. Genes within the fos/jun or activator protein 1 family are targeted via a parallel and independently regulated signal transduction pathway. The continuous requirement for colony-stimulating factor 1 after the immediate early response is initiated indicates that expression of additional delayed early response genes, although contingent on previously induced gene products, might also depend on colony-stimulating factor 1-induced signals. Among the growth factor-regulated delayed early response genes are D-type G1 cyclins, which play an important role in cell-cycle progression. Publication Types: Review Review, Tutorial PMID: 7883784 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 698: Arch Dermatol Res. 1994;286(8):476-80. Changes in oncogene mRNA expression during human keratinocyte differentiation. Sharpe GR, Fisher C, Redfern CP. University Department of Dermatology, Royal Liverpool University Hospital, UK. The nuclear proto-oncogenes are involved in transcriptional regulation and control many cell processes. The role of changes in proto-oncogene expression in controlling the balance between proliferation and differentiation was studied in cultured keratinocytes. Normal human keratinocytes were grown in the serum-free medium MCDB153 with an extracellular calcium concentration of 70 microM. After treatment with different differentiation conditions, cellular RNA was size-fractionated on agarose gels and transferred to nylon membranes which were subsequently hybridized with c-myc, c-jun, and H-ras 32P-labelled probes. Relative RNA loading was assessed using probes for beta-actin and ribosomal 18s RNA. Inducing differentiation by increasing the calcium concentration of the medium from 70 microM to 1.5 mM resulted in a marked decrease in c-myc RNA levels to 26% of control levels within 8 h. After 48 h in 1.5 mM calcium, c-myc levels had recovered to approximately 50% of control levels. There was a gradual reduction in c-jun levels to 56% of control levels by 4 days. Treatment with 10 nM TPA, which also induces keratinocyte differentiation, reduced c-myc RNA levels to 70% of control levels during the first 4 h, but thereafter c-myc levels remained approximately constant for a further 20 h. TGF beta (2 ng/ml), which inhibits keratinocyte growth without inducing differentiation, did not alter c-myc RNA levels over a 4-day period. There were no changes in c-myc levels following the addition of retinoic acid and none of the conditions altered H-ras levels.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 7864662 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 699: Oncol Res. 1994;6(4-5):203-10. Tumor-promoting phorbol ester-induced cell death and gene expression in a human prostate adenocarcinoma cell line. Young CY, Murtha PE, Zhang J. Department of Urology, Mayo Clinic/Foundation, Rochester, MN 55905. 12-O-tetradecanoyl phorbol ester (TPA) has profound cytotoxic effects on a human prostate cancer cell line, LNCaP. The TPA effect may be mediated via a protein kinase C (PKC) pathway, since staurosporine, a potent PKC inhibitor, could reverse the cell-killing effect. Our studies, based on cellular fragmentation, chromatin condensation, and nuclear fragmentation, suggest that the cell-killing effect is due to apoptosis. Moreover, we also examined expression of early growth response genes and androgen-induced genes in association with TPA-induced apoptosis. Northern blot analysis demonstrated that androgen induction of human glandular kallikrein-1 (hKLK2) mRNA was repressed by TPA in a concentration-dependent manner. A time course study showed that both hKLK2 and c-myc mRNAs were repressed by TPA as early as four hours. In contrast, the steady state mRNA levels for c-fos, c-jun, nerve growth factor induced gene A, and the orphan steroid receptor nur77 were rapidly induced within the first two hours of the treatment. Furthermore, transient co-transfection experiments demonstrated that c-fos and c-jun could repress androgen receptor-mediated gene induction. The above studies suggest that (1) the repression of androgen induction of gene expression by TPA-activated PKC is at least in part due to overexpression of c-jun and c-fos and (2) PKC may be a negative growth regulator in prostate cells. PMID: 7841543 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 700: Cytotechnology. 1994;16(3):167-78. Ras oncogene enhances the production of a recombinant protein regulated by the cytomegalovirus promoter in BHK-21 cells. Yano T, Teruya K, Shirahata S, Watanabe J, Osada K, Tachibana H, Ohashi H, Kim EH, Murakami H. Laboratory of Cellular Regulation Technology, Graduate School of Genetic Resources Technology, Kyushu University, Fukuoka, Japan. In order to enhance recombinant protein productivity in animal cells, we developed the oncogene activated production (OAP) system. The OAP system is based on the premise that oncogenes are able to enhance promoter activity. To this end, we constructed reported plasmids by fusing various promoters to the human interleukin-6 (hIL-6) cDNA, and the effector plasmids by inserting individual oncogenes, for example c-myc, c-fos, v-jun, v-myb and c-Ha-ras, downstream from the human cytomegalovirus immediate early (CMV) promoter. Results of transient expression experiments with BHK-21 cells suggest that the CMV promoter is the most potent promoter examined and that the ras product is able to transactivate the beta-actin, CMV and SR alpha promoters. Recombinant BHK-21 cells producing hIL-6 under the control of the CMV promoter were contransfected with the ras oncogene and dihydrofolate reductase gene, then selected with 50 nM methotrexate to coamplify the ras oncogene. We were able to rapidly establish a stable and highly productive clone which exhibited a 35-times higher production rate as compared to the control value. PMID: 7766145 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 701: Crit Rev Oncog. 1994;5(4):359-71. Retrodifferentiation and cell death. Hass R. Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115. The reversibility of a differentiation program termed dedifferentiation, redifferentiation, or retrodifferentiation opens a spectrum of new possibilities for cellular development. During differentiation and retrodifferentiation, the expression of gene products associated with a differentiated phenotype and cell cycle regulation demonstrate inverse patterns. This effect requires a coordinated network that simultaneously controls cell growth and differentiation. In particular, crosstalk between induction of differentiation and G0/G1 cell cycle exit can be initiated and sustained by activated serine/threonine kinases and tyrosine kinases. Phosphorylation signals are relayed to certain genes or transcription factors such as Fos/Jun, EGR-1, NF-kappa B, MyoD, or the Myc/Max gene family. However, the precise regulation of these transcription factors to confer signals to differentiation-associated and cell cycle-regulatory genes remains unclear. Cell cycle exit into a transient G0'-arrest cycle or a terminal G0 phase is determined by a network of phosphorylation signals involving the retinoblastoma protein and a variety of factors such as the E2F family, cyclins, and cyclin-dependent kinases. In this context, a variety of differentiation-induced cell lines, including monocytic, neuronal, or muscle cells, can progress through the G0'-arrest cycle, whereby a certain population retains the capacity to retrodifferentiate and reenter the cell cycle. In contrast, the rest of the differentiated population enters the irreversible G0 phase (terminal commitment) that finally results in programmed cell death. The expression of growth arrest-specific (gas and gadd) genes is associated with the G0'-arrest cycle, and other factors, including c-myc, p53, mdm2, and bcl2/bclx, contribute to the regulation of the cell death program. Although the precise signaling cascade determining retrodifferentiation or cell death remains unclear, a coordinated inter- and intracellular regulation could establish a certain biological balance between these exclusive pathways. Consequently, a retrodifferentiation process may provide a potential for cell type conversion or transdifferentiation, whereby retrodifferentiated cells can be induced to develop via a different pathway according to tissue-specific requirements. Publication Types: Review Review, Tutorial PMID: 7711113 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 702: Eur J Biochem. 1993 Dec 15;218(3):883-91. Insulin-like growth factor-1 modulates steroid hormone effects on osteocalcin synthesis in human MG-63 osteosarcoma cells. Pirskanen A, Jaaskelainen T, Maenpaa PH. Department of Biochemistry & Biotechnology, A. I. Virtanen Institute, University of Kuopio, Finland. The 1,25-dihydroxyvitamin-D3-(calcitriol)-induced medium osteocalcin and cellular osteocalcin mRNA concentrations are increased in a dose-dependent and time-dependent manner by prior treatment of the human MG-63 osteosarcoma cells with insulin-like growth factor-I (IGF-I). In addition, IGF-I reverses the inhibitory effects of dexamethasone and enhances the effects of retinoic acid on osteocalcin synthesis. The stimulatory effect of IGF-I on osteocalcin synthesis is accompanied by stabilization of osteocalcin mRNA and a decrease of AP-1 binding to the osteocalcin promoter. The binding of the vitamin-D receptor (VDR) to its cognate response element is not affected by IGF-I. In contrast to its effects on osteocalcin synthesis, both baseline and calcitriol-stimulated alkaline phosphatase activities are decreased by IGF-I treatment. Furthermore, IGF-I has mitogenic effects on MG-63 cells. The proliferation of the cells and the levels of c-jun mRNA are greatly increased during IGF-I treatment. Calcitriol reduces or eliminates both these effects. The low concentrations of IGF-I used in this study suggest that IGF-I is an important normal regulator of the synthesis of osteocalcin, a bone calcium-binding protein participating in bone mineralization, by modulating the effects of steroid hormones on its synthesis. PMID: 8281940 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 703: Biochem Pharmacol. 1993 Dec 3;46(11):2007-20. Altered gene expression in human leukemia K562 cells selected for resistance to etoposide. Ritke MK, Yalowich JC. Department of Pharmacology, University of Pittsburgh School of Medicine, PA 15261. Sublines of K562 human leukemia cells were selected for resistance (30- to 80-fold) to etoposide by continuous exposure to 0.5 microM VP-16. Two etoposide-resistant cell lines, K/VP.5 and K/VP.5-1, showed a 5-fold reduction in levels of topoisomerase II alpha protein compared with K562 cells. Northern analysis indicated a 2.5-fold reduction in topoisomerase II alpha mRNA in etoposide-resistant cell lines, due in part to a 1.7-fold decrease in topoisomerase II mRNA stability with no change in transcription rate. Immunoblotting assays of electrophoresed cell lysates from VP-16-treated cells revealed less drug-induced covalent topoisomerase II/DNA adducts in resistant than in sensitive cells, suggesting a functional alteration in resistant cell topoisomerase II. Recent reports of specific topoisomerase II DNA binding sites near the promoter sites of growth response genes and alterations of gene expression in cells treated with topoisomerase II inhibitory drugs led to experiments to determine if the apparent functional alterations of topoisomerase II were accompanied by changes in the regulation of these genes. Therefore, the expression of several growth response genes was compared by northern analysis in parental K562 and both VP-16-resistant cell lines. Basal levels of c-myc were comparable for all three cell lines, but levels of c-jun and c-fos were elevated 2- to 4-fold in VP-16-resistant cell lines. Increased levels of c-fos and c-jun were not a result of altered rates of transcription, as determined by nuclear run-off assays. Exposure of both sensitive and resistant cells to 200 microM VP-16 for 5 hr resulted in no further changes in topoisomerase II mRNA levels but caused an additional 2- to 3-fold elevation in the level of c-jun mRNA, indicating that altered basal levels of this gene were not due to deregulation of this gene. Acquired VP-16 resistance in K/VP.5 and K/VP.5-1 cells was accompanied by reduced levels and altered activities of DNA topoisomerase II as well as changes affecting the expression of genes important for growth and differentiation. PMID: 8267650 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 704: Oncogene. 1993 Dec;8(12):3447-57. Overexpression of cyclin D1 in rat fibroblasts causes abnormalities in growth control, cell cycle progression and gene expression. Jiang W, Kahn SM, Zhou P, Zhang YJ, Cacace AM, Infante AS, Doi S, Santella RM, Weinstein IB. Columbia-Presbyterian Cancer Center, Columbia University, College of Physicians and Surgeons, New York, New York 10032. Cyclin D1, a putative G1 cyclin, has been implicated in cell cycle control. The human cyclin D1 gene is located on chromosome 11q13 where DNA rearrangement and amplification have been detected in several types of human cancer. Previous studies demonstrated that the cyclin D1 gene is not only rearranged or amplified but also overexpressed in some of these human tumors and tumor-derived cell lines. To further address the roles of cyclin D1 in cell cycle control and tumorigenesis, we have stably overexpressed the human cyclin D1 cDNA in Rat6 embryo fibroblasts by using retrovirus mediated transduction. The cyclin D1 protein was overproduced about 10-fold and was localized predominately in the nucleus. Cyclin D1 overexpressing cells displayed a decrease in the duration of the G1 phase, decreased cell size, and induced tumors when injected into athymic (nude) mice. In addition, overexpression of cyclin D1 in Rat6 cells perturbed the expression of several cellular growth-related genes including c-myc, c-jun, and cyclin A, but not cyclin D3. Taken together, these results indicate that deregulated expression of the cyclin D1 gene can cause disturbances in cell cycle control and gene expression and also enhance tumorigenesis. PMID: 8247550 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 705: Oncogene. 1993 Dec;8(12):3323-32. v-raf confers CSF-1 independent growth to a macrophage cell line and leads to immediate early gene expression without MAP-kinase activation. Buscher D, Dello Sbarba P, Hipskind RA, Rapp UR, Stanley ER, Baccarini M. Department of Immunbiology, Fraunhofer Institute for Toxicology and Molecular Biology, Hannover, Germany. The BAC-1.2F5 macrophage cell line depends on CSF-1 for proliferation and survival. Phosphorylation and activation of the RAF-1 kinase are among the early events in CSF-1 signal transduction. To characterize the role of RAF-1 in CSF-1-induced proliferation, we overexpressed oncogenically activated RAF-1, cellular RAF-1 and RAF-1 kinase-defective mutant proteins in BAC-1.2F5 cells. We were unable to establish stable cell lines expressing either kinase-negative or full length RAF-1 proteins, implying that expression of these molecules is not tolerated in BAC-1.2F5 cells. Oncogenically activated RAF-1 induces CSF-1-independent growth in the absence of autocrine growth factor production. Autonomous growth is not associated with dedifferentiation, since v-raf-expressing macrophages perform the same immunological functions as control cells. Intriguingly, autonomous growth correlates with the suppression of CSF-1-mediated MAP-Kinase activation and with the low constitutive expression of a number of CSF-1-inducible genes, including fos, jun, ets2, and myc, but also the genes for the inflammatory cytokines TNF alpha and IL-1 beta. Many of these genes have AP-1 binding sites in their promoters, and the v-raf-expressing cells contain constitutive AP-1 binding activity. These data indicate that RAF-1, but not MAP-Kinase, is a key component in CSF-1 mitogenic signal transduction, and are consistent with a working hypothesis in which RAF-1 mediates transcriptional activation of genes via AP-1. PMID: 8247534 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 706: Oncogene. 1993 Dec;8(12):3229-37. Early response gene signalling cascades activated by ionising radiation in primary human B cells. Wilson RE, Taylor SL, Atherton GT, Johnston D, Waters CM, Norton JD. CRC Department of Carcinogenesis, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, UK. We have used a panel of 13 protein kinase C-responsive immediate early gene probes to dissect the cellular signalling pathways activated by ionising gamma radiation in primary human B cells. Of these 13 genes, a delayed transient induction was observed for only 8: c-fos, c-jun, jun-B, jun-D, c-myc, ergI/krox 24 and two 'anonymous' genes, 3L3 and 19A. Expression of c-myc and c-fos mRNAs was paralleled by the appearance of their encoded proteins suggesting that these oncoproteins may couple radiation signalling to cellular responses. Of three protein kinase C-coupled transcription factors examined by gel retardation assay, (AP1, NF kappa B, EgrK/Krox24) only NF kappa B and, to a lesser extent, AP1 was stimulated in response to irradiation. These observations are not obviously compatible with a simple model invoking protein kinase C in radiation signalling in primary B cells and suggest that the pleiotropic effects of ionising radiation on this cell type are mediated through a distinct cellular signalling cascade. PMID: 8247526 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 707: Mol Cell Biol. 1993 Dec;13(12):7447-56. Expression of trkA cDNA in neuroblastomas mediates differentiation in vitro and in vivo. Matsushima H, Bogenmann E. Department of Pediatrics, Jikei University School of Medicine, Tokyo, Japan. The human trkA cDNA was transfected into a malignant human neuroblastoma (NB) cell line (HTLA230) to investigate its role in NB growth and differentiation. This cell line lacks expression of both endogenous trkA and gp75NGFR genes. Transfectants expressing the trkA mRNA and surface-bound receptors transcriptionally activate immediate-early genes (c-fos, c-jun, and jun-B) following nerve growth factor (NGF) stimulation. NGF treatment induces growth arrest as well as down-regulation of the amplified N-myc oncogene. Genes selectively expressed in mature neurons (SCG-10, ret proto-oncogene, GAP-43, etc.) are transcriptionally activated, and neurite outgrowth further demonstrates differentiation of transfectants following NGF stimulation. trkA-expressing NB cells remain tumorigenic in nude mice; however, subcutaneous treatment of tumor-bearing mice with NGF induces Schwannian and neuronal cell differentiation similar to the induction seen in human ganglioneuroblastomas. Thus, trkA expression in HTLA230 cells is sufficient to generate a functional NGF receptor complex that leads to growth-arrested and differentiated NB cells in vitro and in vivo in the presence of NGF. Hence, NGF may play a crucial role in NB cell differentiation and regression in vivo. PMID: 8246962 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 708: Exp Hematol. 1993 Dec;21(13):1640-7. Effect of combined treatment with interleukin-3 and interleukin-6 on 4-hydroperoxycyclo-phosphamide-induced programmed cell death or apoptosis in human myeloid leukemia cells. Bullock G, Tang C, Tourkina E, Ibrado AM, Lutzky J, Huang Y, Mahoney ME, Bhalla K. Division of Hematology/Oncology, Medical University of South Carolina, Charleston 29425. In autologous bone marrow transplantation in patients with acute myeloid leukemia (AML), 4-hydroperoxycyclophosphamide (4-HC) is a commonly used ex vivo purging agent for leukemic blasts. In the present report, we demonstrate that exposure to high concentrations of 4-HC for 1 hour, as used in ex vivo bone marrow purging, produces internucleosomal DNA fragmentation characteristic of apoptosis, or programmed cell death (PCD), in human myeloid leukemia HL60 cells. Lower concentrations of 4-HC (10, 20, or 50 microM/L) failed to cause this effect, while higher concentrations (> or = 200 microM/L) produced random DNA fragmentation. 4-HC-mediated internucleosomal DNA fragmentation was associated with a marked induction in c-jun and significant reductions in bcl-2 and c-myc oncogene expressions. A combined treatment with interleukin-3 (IL-3) plus IL-6 for 18 hours before an additional, 1-hour concurrent treatment with 4-HC (100 microM/L) significantly increased 4-HC-induced DNA fragmentation as well as colony growth inhibition of HL60 cells. The effects of cotreatment with IL-3 plus IL-6 were also associated with a further, modest decrease in bcl-2 and c-myc and augmentation of c-jun expression. These findings highlight an alternative mechanism of 4-HC-induced leukemic cell death that can be potentially enhanced by cotreatment with IL-3 plus IL-6. PMID: 8243566 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 709: Am J Vet Res. 1993 Dec;54(12):2010-4. Specific expression of cellular oncogenes c-myc and c-myb in T-cell lines established from three types of bovine lymphosarcomas. Ishiguro N, Matsui T, Shinagawa M. Department of Veterinary Public Health, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Japan. Expression of cellular oncogenes in 3 lymphoid cell lines, BTL-PC3 (BoCD2-, BoCD4-, BoCD8-, BoWC1+), BLS1 (BoCD2+, BoCD4-, BoCD8-, BoWC1+) and BLT2 (BoCD2-, BoCD4-, BoCD8-, BoWC1-), which have been established from calf, skin, and thymic types of lymphosarcomas, respectively, were analyzed by DNA-RNA (northern blot) hybridization. To determine specific expression of oncogenes involved in malignant transformation of the lymphoid cells, cellular RNA was isolated from bovine tumor cell lines, BTL-PC3, BLS1, and BLT2, and from Madin Darby bovine kidney cells used as a control for bovine cell lines. The RNA was hybridized against 5 viral oncogene probes (v-jun, v-myc, v-erbB, v-erbA and v-fes), 6 human cellular oncogene probes (N-ras, c-Blym-1 c-erbB-2, c-fos, c-myb and c-abl), human p53 tumor suppressor gene, and bovine LDH-A gene probes. Line BTL-PC3 expressed 2.4-kilobase (kb) c-myc and 4.0- and 3.6-kb c-myb transcripts, and line BLT2 expressed a 3.8-kb c-myb transcript, but line BLS1 expressed no message for the oncogenes tested. Specific transcripts of p53 were found in BTL-PC3 and BLT2 lines, but not in BLS1. Madin Darby bovine kidney cell line expressed multiple cellular oncogenes, c-jun, c-myc, and c-fos, and p53 genes. Southern blot hybridization did not reveal abnormal DNA rearrangements associated with the expressed oncogenes (c-myc and c-myb) in the 3 bovine tumor lines. (ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8116930 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 710: Environ Health Perspect. 1993 Dec;101 Suppl 5:163-8. Compensatory regeneration, mitogen-induced liver growth, and multistage chemical carcinogenesis. Ledda-Columbano GM, Coni P, Simbula G, Zedda I, Columbano A. Istituto di Patologia Sperimentale, Universita di Cagliari, Italy. Liver cell proliferation has often been implicated to play a major role during different steps of the carcinogenic process. Most of the experimental studies indicating a close association between cell proliferation and liver cancer development have made use of a compensatory type of proliferative stimulus. However, liver growth may also be caused by direct hyperplasia after administration of primary mitogens. Our recent studies examined the possible differences between these two types of cell proliferation. Our studies indicate that a) increased expression of proto-oncogenes such as c-fos, c-jun, and c-myc is not necessary for entry into the cell cycle during mitogen-induced liver growth; b) mitogen-induced liver growth does not support initiation of chemical hepatocarcinogenesis; c) repeated proliferative stimuli induced by primary mitogens do not stimulate the growth of initiated cells to a focal and/or nodular stage; and d) mitogen-induced liver growth, unlike compensatory regeneration, is followed by a particular mode of cell death, namely, apoptosis. This type of cell death may be responsible for the elimination of carcinogen-initiated cells. Publication Types: Review Review, Tutorial PMID: 8013404 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 711: Biochem Biophys Res Commun. 1993 Nov 30;197(1):278-86. Regulation of c-MYC protein expression in the developing rat cerebellum by phosphoinositide turnover. Takahashi M, Toyoshima S, Miyazawa A, Horikoshi T, Yoshioka T. Department of Psychiatry, School of Medicine, Yokohama City University, Japan. Using developing rat cerebellum, we examined the correlation between the turnover of phosphoinositide (PI) and c-myc expression. The 32P incorporation into polyphosphoinositides changed remarkably with advancing age. It reached a maximum value on PND 7, and then decreased gradually until PND 14. Immunostaining by anti-PIP2 antibody changed in parallel. The expression of c-myc mRNA was also changed developmentally, showing a peak on PND 7; whereas c-MYC protein expression showed a peak on PND 10. Together, these results suggest that c-myc expression is regulated by PI turnover during the early developing stage of rat cerebellum. PMID: 8250935 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 712: Blood. 1993 Nov 15;82(10):3133-40. High-dose mitoxantrone induces programmed cell death or apoptosis in human myeloid leukemia cells. Bhalla K, Ibrado AM, Tourkina E, Tang C, Grant S, Bullock G, Huang Y, Ponnathpur V, Mahoney ME. Division of Hematology/Oncology, Medical University of South Carolina, Charleston 29425. Mitoxantrone has been shown in vitro to exhibit a steep dose-response relationship with respect to the clonogenic survival of acute myeloid leukemia cells. In this report, we show that 1-hour exposure of human myeloid leukemia HL-60 and KG-1 cells to mitoxantrone concentrations ranging between 0.1 and 10.0 mumol/L induced internucleosomal DNA fragmentation of approximately 200-bp integer multiples, characteristic of cells undergoing programmed cell death (PCD) or apoptosis. Mitoxantrone-mediated PCD was associated with a steep inhibition of the clonogenic survival of the leukemic cells. In addition, intracellularly, mitoxantrone-induced PCD was associated with a marked induction of c-jun and significant repression of c-myc and BCL-2 oncogenes. Pretreatment with the protein kinase C stimulator phorbol myristate acetate enhanced mitoxantrone-induced internucleosomal DNA fragmentation, whereas protein kinase C inhibitors staurosporine and H7 had no effect. These findings suggest that PCD is a potential mechanism underlying the steep dose-response relationship of mitoxantrone to the inhibition of clonogenic survival of acute myeloid leukemia cells. PMID: 8219202 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 713: Nucleic Acids Res. 1993 Nov 11;21(22):5092-100. Specific cleavage of transcription factors by the thiol protease, m-calpain. Watt F, Molloy PL. CSIRO Division of Biomolecular Engineering, Sydney Laboratory, North Ryde, NSW, Australia. The intracellular nonlysosomal calcium-dependent cysteine protease, m-calpain, is shown to specifically cleave the bHLHzip transcription factor USF leaving the binding and dimerisation domains intact. The resultant protein is capable of efficient DNA binding but is no longer able to activate transcription. A surprisingly high proportion of other transcription factors tested, AP1 (c-Fos/c-Jun), Pit-1, Oct-1, CP1a and b, c-Myc, ATF/CREB, AP2 and AP3 but not Sp1, were similarly cleaved by m-calpain to produce specific partial digestion products. These properties make m-calpain a particularly useful protease for proteolytic studies of transcription factors and also raise the possibility that m-calpain may be involved in vivo in regulation of turnover or transcriptional activity of a number of transcription factors. PMID: 8255762 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 714: Anticancer Res. 1993 Nov-Dec;13(6A):2021-5. Correlation between successful heterotransplantation of lung tumors in nude mice, poor prognosis of patients and expression of Fos, Jun, ErbB1, and Ras. Volm M, Mattern J. German Cancer Research Center, Heidelberg. In order to examine whether the expression of the oncoproteins might be important for the malignancy of tumors, the relationship between the take rate of 88 human squamous cell lung carcinomas in nude mice and the expression of protooncogene products was analyzed. The expression of c-fos, c-jun, c-ras, c-erbB1, c-neu and c-myc at the protein level was investigated by immunohistochemistry. Tumor take was assumed if within three months growing nodules were detected and confirmed histologically. The take rate of squamous cell lung carcinomas in nude mice was 49%. Sixty-eight percent of the tumors were positive for Fos, 40% for Jun, 67% for Ras, 77% for ErbB1, 35% for Neu and 39% for Myc. Tumors with an (over)expression of the proteins encoded by the oncogenes c-fos, c-jun, c-erbB1 and c-ras had a significantly higher take rate in nude mice than tumors without an (over)expression of the oncogene products. In contrast, the expression of the c-neu and the c-myc genes at the protein level had no influence on the take rate of the tumors in nude mice. Interestingly, only patients with tumors with an (over)expression of the proteins encoded by the oncogenes c-fos, c-jun, c-erbB1 and c-ras had significantly shorter survival times than patients whose tumors did not show an (over) expression of the oncogene products. These results demonstrate that the aggressiveness of the tumors visible in the higher take rate of the tumors in nude mice and in the shorter survival times of patients can be detected by measurement of the expression of c-fos, c-jun, c-erbB1 and c-ras at the protein level. PMID: 8297109 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 715: Trends Biochem Sci. 1993 Nov;18(11):433-7. DNA damage and the DNA-activated protein kinase. Anderson CW. Biology Department, Brookhaven National Laboratory, Upton, NY 11973-5000. DNA-activated protein kinase (DNA-PK) is a nuclear serine/threonine protein kinase that is activated in vitro by DNA fragments. The cellular targets of DNA-PK are nuclear, DNA-binding, regulatory proteins including Sp1, Fos, Jun, Myc, the tumor suppressor protein p53, and RNA polymerase II. These characteristics suggest a role for DNA-PK in coordinating nuclear processes and as a modulator of checkpoint mechanisms activated by DNA damage. Publication Types: Review Review, Tutorial PMID: 8291090 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 716: Hepatology. 1993 Nov;18(5):1193-201. Induction of metallothionein and its localization in the nucleus of rat hepatocytes after partial hepatectomy. Tohyama C, Suzuki JS, Hemelraad J, Nishimura N, Nishimura H. National Institute for Environmental Studies, Ibaraki, Japan. Metallothioneins, a group of cysteine-rich heavy-metal binding proteins, are induced in the regenerating rat liver in response to the stimuli evoked by partial hepatectomy. We have investigated the expression of metallothionein genes and proto-oncogenes (c-fos, c-jun and c-myc), as well as specific localization of metallothionein in the liver cells after partial hepatectomy. Metallothionein mRNA was detected as early as 3 hr and reached a maximal level by 6 hr. Expression of the proto-oncogenes apparently preceded the elevation of metallothionein protein because the latter was maximal 18 hr after partial hepatectomy, followed by a decrease until 70 hr. Hepatocytes of the intact rat liver have metallothionein in the cytoplasm only. Interestingly, metallothionein was localized predominantly in the nucleus as early as 6 hr after partial hepatectomy, and the staining intensity of metallothionein became maximal at 15 hr, followed by detection in both the cytoplasm and nucleus at 24 hr or longer. The use of a confocal laser scanning microscope with both tissue sections and isolated nuclei has clearly shown that metallothionein immunofluorescence exists inside hepatocyte nuclei after partial hepatectomy. Expression of the proto-oncogenes c-fos and c-jun is elevated after partial hepatectomy, and the resultant heterodimer of gene products may contribute to the observed metallothionein gene induction. However, the observation that metallothionein protein levels were elevated until 18 hr after partial hepatectomy suggests that an alternative pathway for the induction of metallothionein gene expression may also be present.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8225226 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 717: EMBO J. 1993 Nov;12(11):4181-9. Signal transduction by the high-affinity GM-CSF receptor: two distinct cytoplasmic regions of the common beta subunit responsible for different signaling. Sato N, Sakamaki K, Terada N, Arai K, Miyajima A. DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, CA 94304. The high-affinity receptors for granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin 3 (IL-3) and IL-5 consist of two subunits, alpha and beta. The alpha subunits are specific to each cytokine and the same beta subunit (beta c) is shared by these three receptors. Although none of these receptor subunits has intrinsic kinase activity, these cytokines induce protein tyrosine phosphorylation, activation of Ras, Raf-1 and MAP kinase, and transcriptional activation of nuclear proto-oncogenes such as c-myc, c-fos and c-jun. In this paper, we describe a detailed analysis of the signaling potential of the beta c subunit by using a series of cytoplasmic deletion mutants. The human beta c consists of 881 amino acid residues. A C-terminal deletion mutant of beta c at amino acid 763 (beta 763) induced phosphorylation of Shc and activation of Ras, Raf-1, MAP kinase and p70 S6 kinase, whereas a deletion at amino acid 626 (beta 626) induced none of these effects. The beta 763 mutant, as well as the full-length beta c, induced transcription of c-myc, c-fos and c-jun. Deletions at amino acid 517 (beta 517) and 626 (beta 626) induced c-myc and pim-1, but no induction of c-fos and c-jun was observed. GM-CSF increased phosphatidylinositol 3 kinase (PI3-K) activity in anti-phosphotyrosine immunoprecipitates from cells expressing beta 763 as well as beta c, whereas it was only marginally increased from cells expressing beta 517 or beta 626. Thus, there are at least two distinct regions within the cytoplasmic domain of beta c that are responsible for different signals, i.e. a membrane proximal region of approximately 60 amino acid residues upstream of Glu517 is essential for induction of c-myc and pim-1, and a distal region of approximately 140 amino acid residues (between Leu626 and Ser763) is required for activation of Ras, Raf-1, MAP kinase and p70 S6 kinase, as well as induction of c-fos and c-jun. PMID: 8223433 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 718: Neurosci Lett. 1993 Oct 29;161(2):161-4. Selectively high expression of the transcription factor AP1 in telencephalic structures of epileptic E1 mice. Yoneda Y, Ogita K, Kabutoz H, Mori A. Department of Pharmacology, Setsunan University, Osaka, Japan. In ddY mouse brain, the transcription factors AP1 and CREB were rich in the cerebral cortex, hippocampus and cerebellum but relatively poor in the striatum, hypothalamus and midbrain. In contrast, the transcription factor Myc was rather poorly distributed in mouse brain under the conditions employed. Among these 3 transcription factors examined, DNA binding activities of only AP1 were invariably higher in telencephalic structures, such as the cortex, hippocampus and striatum, of the epileptic El mice than in those of parent ddY mice. These results suggest that the transcription factor AP1 may be at least in part responsible for molecular mechanisms underlying pathogenesis of a variety of abnormal symptoms observed in epileptic El mice. PMID: 8272259 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 719: Proc Natl Acad Sci U S A. 1993 Oct 15;90(20):9611-5. Tumor necrosis factor-induced c-myc expression in the absence of mitogenesis is associated with inhibition of adipocyte differentiation. Ninomiya-Tsuji J, Torti FM, Ringold GM. Affymax Research Institute, Palo Alto, CA 94304. Tumor necrosis factor (TNF) inhibits and reverses differentiation of mouse adipogenic TA1 cells. We have found that TNF induces c-myc in a sustained manner in both preadipocytes and adipocytes; in contrast, serum induces c-myc transiently and only in preadipocytes. This TNF-mediated c-myc induction is not coupled with cell proliferation but is correlated with TNF-mediated inhibition of adipocyte differentiation. We prepared an inducible c-myc transformant of TA1 cells by transfection of the mouse c-myc gene under the control of the metallothionein-I promoter. These cells are unable to differentiate to adipocytes in the presence of Zn2+/Cd2+, and in differentiated TA1 cells, Zn2+/Cd2+ causes reduction of adipocyte-specific gene expression as does TNF. Lastly, exposure of TA1 cells to antisense c-myc oligonucleotide partially blocked the TNF-mediated reduction of adipocyte-specific gene expression. Thus, TNF-mediated c-myc expression is distinct in character from that involved in mitogenic responses but appears to play an important role in inhibition and reversal of adipocyte differentiation. PMID: 8415749 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 720: J Biol Chem. 1993 Oct 15;268(29):21777-82. Ethanol inhibits the autophosphorylation of the insulin-like growth factor 1 (IGF-1) receptor and IGF-1-mediated proliferation of 3T3 cells. Resnicoff M, Sell C, Ambrose D, Baserga R, Rubin R. Department of Pathology and Cell Biology, Jefferson Medical College, Philadelphia, Pennsylvania 19107. The effect of ethanol on cell proliferation was studied in Balb/c 3T3 cells and in stably transfected 3T3 cells constitutively overexpressing the human insulin-like growth factor 1 (IGF-1) receptor (p6 cells). Ethanol inhibited growth of both cell lines when they were cultured in serum-free medium supplemented with individual growth factors, i.e. platelet-derived growth factor and IGF-1 for 3T3 cells, and IGF-1 only for p6 cells. Increases in cell number were prevented in both cell lines even when ethanol was present exclusively during the period of IGF-1 stimulation. The inhibitory effect of ethanol was concentration-dependent, with a 30% inhibition observed at 10 mM ethanol. IGF-1 receptor tyrosine autophosphorylation was completely prevented by ethanol both in intact cells and in immunopurified IGF-1 receptor preparations. The binding of IGF-1 to its receptor on intact cells was unaffected by ethanol. Ethanol also inhibited the stimulation of IGF-1 receptor autophosphorylation and the corresponding growth of p6 cells induced by IGF-2. Transcription of c-myc, c-fos, and c-jun in response to IGF-1 was inhibited by ethanol. These findings demonstrate that ethanol at low concentrations markedly inhibits IGF-1 receptor autophosphorylation and IGF-1-mediated cell growth. PMID: 8408032 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 721: EMBO J. 1993 Oct;12(10):3921-9. A transcription factor with homology to the AP-1 family links RNA transcription and DNA replication in the lytic cycle of Epstein-Barr virus. Schepers A, Pich D, Hammerschmidt W. Institut fur Klinische Molekularbiologie und Tumorgenetik, GSF-Forschungszentrum fur Umwelt und Gesundheit, Munchen, Germany. oriLyt, the lytic origin of DNA replication of Epstein-Barr virus (EBV), ensures viral DNA amplification during the productive or lytic phase of the virus' life cycle. To understand the contribution of cis- and transacting elements involved in DNA replication of oriLyt, a detailed mutational analysis was undertaken which defined BZLF1, a viral transcriptional activator, as an essential replication factor. The BZLF1 protein belongs to the extended fos/jun family of transcription factors and binds to specific BZLF1 binding motifs within oriLyt, as well as to consensus AP-1 sites. Recombinant, chimeric transcription factors identified the transcriptional activation domain of BZLF1 as being necessary to mediate DNA replication, a function which could not be substituted by any other transcription factor tested, including jun, E2, myc or VP16. PMID: 8404860 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 722: Mol Biol Cell. 1993 Oct;4(10):983-92. Differential regulation of early response genes and cell proliferation through the human granulocyte macrophage colony-stimulating factor receptor: selective activation of the c-fos promoter by genistein. Watanabe S, Muto A, Yokota T, Miyajima A, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Granulocyte macrophage colony-stimulating factor (GM-CSF) binds to the high-affinity GM-CSF receptor (GMR) consisting of alpha and beta subunits and induces rapid tyrosine phosphorylation, activation of early response genes, and proliferation of hematopoietic cells. The alpha subunit is the primary cytokine binding component and the beta subunit is required for high-affinity binding as well as for signal transduction. Using tyrosine kinase inhibitors and cytoplasmic deletion mutants of the beta subunit, we obtained evidence that there are at least two distinct pathways downstream of the GMR in BA/F3 cell, one which is essential for proliferation, leads to the c-myc gene activation, and is sensitive to herbimycin and genistein. Activation of this pathway depends on the cytoplasmic region between amino acid positions 455 and 517 of the beta subunit. The second pathway, which leads to activation of c-fos and c-jun genes, is only partially sensitive to herbimycin, is resistant to genistein and depends on the region between amino acid positions 626 and 763 of the beta subunit. Unexpectedly, the c-fos mRNA induction was augmented by genistein. The enhanced expression of c-fos mRNA by genistein also occurred with stimulation with cAMP, PMA, or EGF in NIH3T3 cells. It thus seems likely that genistein affects a common pathway downstream of these signals. PMID: 8298195 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 723: Semin Immunol. 1993 Oct;5(5):345-64. Transmembrane signaling by the interleukin-2 receptor: progress and conundrums. Mills GB, Schmandt R, Gibson S, Leung B, Hill M, May C, Shi YF, Branch DR, Radvanyi L, Truitt KE, et al. Oncology Research, Toronto General Hospital, Ontario, Canada. Activation of the multicomponent interleukin-2 receptor (IL-2R) complex leads to a rapid increase in tyrosine phosphorylation of a number of cellular proteins including the IL-2R beta and IL-2R gamma chains of the IL-2R and the RAF-1 serine threonine kinase. In addition, phosphatidylinositol 3-kinase (PI-3K) protein and activity can be immunoprecipitated with anti-phosphotyrosine and anti-IL-2R beta antibodies from IL-2-activated but not resting T lymphocytes. We have demonstrated that the SH2 (SRC homology 2) domains of the 85 kDa subunit of PI-3K are sufficient to mediate binding of the PI-3K complex to tyrosine phosphorylated, but not non-phosphorylated IL-2R beta, suggesting that tyrosine phosphorylation is an integral component of the activation of PI-3K by the IL-2R. Since none of the members of the IL-2R complex contains an intrinsic tyrosine kinase domain, IL-2-induced tyrosine phosphorylation must be the consequence of activation of intracellular tyrosine kinases. SRC family members including lck, lyn and fyn have been demonstrated to associate with IL-2R beta through binding of the kinase domain to the acidic domain of IL-2R beta. However, we have demonstrated that the serine rich (SD) region of the cytosolic domain of IL-2R beta is also required for association of a tyrosine kinase with the IL-2R complex and that IL-2 can induce proliferation and tyrosine phosphorylation in cell lines which lack the known SRC family kinases expressed by T lymphocytes. Thus members of other kinase families besides SRC may also be involved in mediating IL-2 signal transduction. Biochemical studies and studies of cells expressing mutant IL-2 receptors indicate that IL-2-induced tyrosine kinase activation initiates a complex signaling cascade. The cascade includes SRC family kinase members such as lck, fyn, and lyn, activation of Raf-1 and PI-3K, and ras, and increased expression of the fos, fra-1, and jun protooncogenes. In addition, ligation of the IL-2R leads to rapid increases in myc expression and more delayed increases in the expression of the cdc2 and cdk2 kinases and the cyclins through a tyrosine phosphorylation independent pathway. Whether other biochemical processes initiated by IL-2R ligation, including activation of the MAP2, p70S6 and p90RSK serine threonine kinases, activation of NF-kappa B, and increased expression of Raf-1, Pim-1, bcl-2, IL-2R alpha and IL-2R beta, are consequences of the IL-2-induced tyrosine kinase cascade remains to be determined.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 8260651 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 724: Mol Pharmacol. 1993 Sep;44(3):560-8. Rat CYP1A1 negative regulatory element: biological activity and interaction with a protein from liver and hepatoma cells. Sterling K, Weaver J, Ho KL, Xu LC, Bresnick E. Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, New Hampshire 03755-3835. Rat CYP1A1 promoter activity was suppressed by the presence of a cis negative regulatory element (NRE) at position -843 to -746 in transiently transfected rat H4IIE and human HepG2 hepatoma cells. Removal of the NRE from the promoter-fusion gene constructs caused an increase in the basal promoter activity of 2-6-fold. Co-transfection of the NRE-containing or non-NRE-containing CYP1A1 promoter-fusion gene constructs with a cloned rat NRE, i.e., pNRE, into HepG2 cells caused a 2-fold or greater reduction in constitutive and induced promoter activities. 2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced expression of the endogenous human CYPA1 was also inhibited by transfection of pNRE into HepG2 cells. Deletion of the sequence from base pairs (bp) -658 to -269 in the NRE-containing construct caused a dramatic decrease of constitutive expression in transiently transfected HepG2 cells, compared with an identical construct that lacked the NRE. Deletion of the sequences between bp -658 and -158 in the CYP1A1 promoter did not affect reporter gene activity, indicating a second site of interaction. At least three different rat liver nuclear proteins bound to the rat NRE, as determined by gel mobility shift and DNase I footprinting assays. A 32-bp sequence within the rat NRE, with significant sequence identity to the 26-bp c-myc, fos/jun-octamer-binding, NRE, was protected from DNAse I cleavage by rat liver nuclear extracts. These data suggested a role for this region in the negative regulation of rat CYP1A1. PMID: 8396716 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 725: Circ Res. 1993 Sep;73(3):413-23. Molecular characterization of angiotensin II--induced hypertrophy of cardiac myocytes and hyperplasia of cardiac fibroblasts. Critical role of the AT1 receptor subtype. Sadoshima J, Izumo S. Molecular Medicine Division, Beth Israel Hospital, Boston, Mass. 02215. Increasing evidence suggests that angiotensin II (Ang II) may act as a growth factor for the heart. However, direct effects of Ang II on mammalian cardiac cells (myocytes and nonmyocytes), independent of secondary hemodynamic and neurohumoral effects, have not been well characterized. Therefore, we analyzed the molecular phenotype of cultured cardiac cells from neonatal rats in response to Ang II. In addition, we examined the effects of selective Ang II receptor subtype antagonists in mediating the biological effects of Ang II. In myocyte culture, Ang II caused an increase in protein synthesis without changing the rate of DNA synthesis. In contrast, Ang II induced increases in protein synthesis, DNA synthesis, and cell number in nonmyocyte cultures (mostly cardiac fibroblasts). The Ang II-induced hypertrophic response of myocytes and mitogenic response of fibroblasts were mediated primarily by the AT1 receptor. Ang II caused a rapid induction of many immediate-early genes (c-fos, c-jun, jun B, Egr-1, and c-myc) in myocyte and nonmyocyte cultures. Ang II induced "late" markers for cardiac hypertrophy, skeletal alpha-actin and atrial natriuretic factor expression, within 6 hours in myocytes. Ang II also caused upregulation of the angiotensinogen gene and transforming growth factor-beta 1 gene within 6 hours. Induction of immediate-early genes, late genes, and growth factor genes by Ang II was fully blocked by an AT1 receptor antagonist but not by an AT2 receptor antagonist. These results indicate that: (1) Ang II causes hypertrophy of cardiac myocytes and mitogenesis of cardiac fibroblasts, (2) the phenotypic changes of cardiac cells in response to Ang II in vitro closely mimic those of growth factor response in vitro and of load-induced hypertrophy in vivo, (3) all biological effects of Ang II examined here are mediated primarily by the AT1 receptor subtype, and (4) Ang II may initiate a positive-feedback regulation of cardiac hypertrophic response by inducing the angiotensinogen gene and transforming growth factor-beta 1 gene. PMID: 8348686 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 726: Mol Endocrinol. 1993 Sep;7(9):1121-32. Glucocorticoids induce a G1/G0 cell cycle arrest of Con8 rat mammary tumor cells that is synchronously reversed by steroid withdrawal or addition of transforming growth factor-alpha. Goya L, Maiyar AC, Ge Y, Firestone GL. Department of Molecular and Cell Biology, University of California, Berkeley 94720. Con8 mammary tumor cells are an epithelial cell line derived from the 7,12-dimethylbenz(alpha)anthracene-induced 13762NF rat mammary adenocarcinoma. The synthetic glucocorticoid dexamethasone suppresses the growth of Con8 cells, and after 5 days of treatment with this steroid, Con8 cells undergo less than 0.5 population doublings. This growth arrest is accompanied by a 30-fold elevation in c-jun transcript levels, no change in c-fos expression, and a moderate increase in total AP-1 transcriptional activity. Dexamethasone inhibited DNA synthesis within one cell cycle, and flow cytometry of propidium iodide-stained nuclei demonstrated that dexamethasone growth-suppressed cells had a DNA content indicative of a specific cell cycle block in either G1 or G0. Consistent with a G1/G0 arrest of the cell cycle, dexamethasone did not prevent Con8 cells from entering the S phase after release from synchronization at the G1/S boundary by a double thymidine block. Analysis of [3H]thymidine incorporation and autoradiography of [3H]thymidine-labeled nuclei revealed that after either dexamethasone withdrawal or the addition of transforming growth factor-alpha (TGF alpha), Con8 cells synchronously reinitiate cell cycle progression. Northern blot analysis demonstrated that an induction of transcripts for the G1 marker genes c-myc and cyclin D1 occurs before cells enter the S-phase. After dexamethasone withdrawal, c-myc and cyclin D1 expression transiently peak at 2 and 4 h, respectively. In contrast, c-myc expression peaked at 0.5-1 h, whereas cyclin D1 expression was induced at 2 h and maintained at a high level after the addition of TGF alpha. Our results demonstrate that glucocorticoids induce a specific block of the cell cycle progression of a rat mammary tumor cell, and that after synchronous progression through the cell cycle, the temporal expression pattern for c-myc and cyclin D1 is distinct for dexamethasone release vs. the addition of TGF alpha to glucocorticoid-suppressed cells. PMID: 8247014 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 727: Growth Regul. 1993 Sep;3(3):172-9. Platelet derived growth factor/tyrosine kinase receptor mediated proliferation. Benito M, Lorenzo M. Departamento de Bioquimica y Biologia Molecular, Facultad de Farmacia, Universidad Complutense, Madrid, Spain. PDGF heterodimer of A and B chains, a complete mitogen for 3T3 mouse fibroblasts, exemplifies those growth factors interacting with membrane associated tyrosine kinase receptors. Its binding to the PDGF-receptors results in receptor dimerization and subsequent activation of tyrosine kinase activity in the cytoplasmic protein domain, autophosphorylation of the receptor being the first event in the transduction cascade. Before the ligand-receptor complex is internalized and degraded, receptor stimulation is transmitted to the general transduction network, in which several tyrosine kinase substrates are activated by phosphorylation and changes the cytoplasmic biochemistry. These changes include cytoplasmic alkalinization, increases in the intracellular concentration of cyclic-AMP and Ca2+ and activation of protein kinase C through the degradation of phosphoinositides. The known substrates recruited by the PDGF-receptor association are phosphatidylinositol-3'-kinase, ras-GTPase-activating protein, phospholipase C-gamma, serine-threonine kinase Raf-1 and src and src-related tyrosine kinases. Upon binding of PDGF to its receptor, transactivation of transcriptional and nuclear factors such as c-fos and c-myc genes and dephosphorylation of c-jun occurs, V-sis, the oncogen of the simian sarcoma virus (SSV), is highly homologous to the c-sis/PDGF-B gene that encodes the homodimer of the B-chain of the PDGF receptor. Cells transformed by SSV have been studied as a model system for the autocrine stimulation of the PDGF receptor. Publication Types: Review Review, Tutorial PMID: 8220110 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 728: Exp Cell Res. 1993 Sep;208(1):175-88. Blockage of EGF receptor signal transduction causes reversible arrest of normal and immortal human mammary epithelial cells with synchronous reentry into the cell cycle. Stampfer MR, Pan CH, Hosoda J, Bartholomew J, Mendelsohn J, Yaswen P. Life Sciences Division, Lawrence Berkeley Laboratory, Berkeley, California 94720. We demonstrate that blockage of EGF receptor signal transduction is sufficient by itself to cause a rapid, efficient, and reversible G0-like growth arrest of normal human mammary epithelial cells (HMEC) of finite lifespan as well as two immortally transformed cell lines derived from normal HMEC following in vitro transformation with benzo[a]pyrene. For normal HMEC, the significant level of endogenous production of TGF alpha requires utilization of blocking antibodies to the EGF receptor to achieve cessation of growth in mass culture, whereas removal of EGF is sufficient to arrest the immortal cell lines. In the growth-arrested cells, protein synthesis remains depressed; reexposure to EGF leads to a rapid increase in protein synthesis. Inhibition of DNA synthesis is not detectable until approximately 12 h after removal of EGF/TGF alpha and is pronounced by 24 h. Reexposure to EGF produces high levels of synthesis of the early response genes, c-myc, c-fos, c-jun, and MGSA, within 1 h. DNA synthesis increases only after 10 h, with a sharp peak after 15-20 h. Reexposure of the growth-arrested normal HMEC for 1 h with EGF allows a majority of the cells capable of cycling to subsequently enter the S phase. Little is currently known about cell cycle control in normal human epithelial cells. The efficient and gentle method of achieving reversible G0 growth arrest reported here may facilitate studies on the cell cycle of this cell type. Additionally, results from normal HMEC can be compared with those from syngeneic immortalized cell populations to determine possible cell cycle parameters altered as a result of immortal transformation. PMID: 7689475 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 729: Mol Cell Biol. 1993 Sep;13(9):5888-97. Differential modulation of plasminogen activator gene expression by oncogene-encoded protein tyrosine kinases. Bell SM, Connolly DC, Maihle NJ, Degen JL. Division of Basic Science Research, Children's Hospital Research Foundation, Cincinnati, Ohio 45229. Urokinase-type plasminogen activator (uPA) gene transcription is increased > or = 50-fold in chicken embryo fibroblasts (CEF) following transformation by the protein tyrosine kinase pp60v-src. Protein phosphorylation appears to play a critical role in uPA gene expression in these cells; protein kinase C-activating phorbol esters cooperate with pp60v-src to synergistically increase uPA mRNA, whereas cyclic AMP (cAMP)-dependent protein kinase-activating agents (e.g., 8-bromo cAMP) repress uPA mRNA levels. To explore the relationship between transforming oncogenes and uPA gene expression, uPA mRNA levels were measured in CEF infected with selected avian retroviruses. We report that v-ras and the transforming protein tyrosine kinases v-src, v-yes, and v-ros all increase cellular uPA mRNAs. However, transformation with the protein tyrosine kinase encoded by v-erbB, or the nuclear proteins encoded by v-jun, v-ski, or v-myc, did not increase uPA mRNA detectably. Ras and all of the protein tyrosine kinases analyzed, including the v-erbB product, but none of the nuclear oncoproteins sensitized cells to phorbol ester induction of uPA gene expression. Thus, increased uPA gene expression is not simply a secondary consequence of cell transformation but, rather, is regulated or comodulated by only a subset of oncogene products. Analysis of cells expressing site-directed mutants of pp60v-src showed that the induction of the uPA gene is dependent on protein tyrosine kinase catalytic activity, myristylation, and plasma membrane localization. However, these properties together are not sufficient; an additional feature in the src homology 2 domain is also required. The major sites of serine phosphorylation, serines 12 and 17, and the autophosphorylation site, tyrosine 416, are not essential for uPA gene induction. However, the reduction of uPA mRNA in pp60v-src-transformed cells by 8-bromo cAMP is dependent on tyrosine 416. PMID: 7689154 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 730: J Biol Chem. 1993 Aug 25;268(24):18018-29. Interleukin-1-inducible expression of gro-beta via NF-kappa B activation is dependent upon tyrosine kinase signaling. Joshi-Barve SS, Rangnekar VV, Sells SF, Rangnekar VM. Department of Surgery, University of Kentucky, Lexington 40536. Interleukin-1 (IL-1) induces programmed growth arrest in human melanoma cells, A375-C6. IL-1 action in these cells is associated with induction of a cell type-specific immediate-early (IE) gene expression program characterized by strong, rapid, and sustained induction of gro-alpha and gro-beta, but transient induction of c-jun, IRG-9, and NAK-1, and lack of induction of c-myc. With the exception of gro-alpha and gro-beta, these IE genes are also associated with growth-stimulatory responses in the melanoma cells, suggesting that the gro-genes may play key roles in the growth arrest action of the cytokine. To elucidate the early intracellular signals associated with IL-1 action, we are studying the second messenger signals and transcription factors required for induction of gro-genes. Here, we present evidence that IL-1-inducible gro-gene expression is dependent on tyrosine kinase signaling. Using gel retardation and transient expression assays, we show that IL-1 causes protein tyrosine phosphorylation-dependent activation of NF-kappa B enhancer binding protein, which then induces transcription of the gro-genes via an NF-kappa B site located 76 base pairs upstream from the cap site. IL-1-activated protein tyrosine phosphorylation is also required for gro-gene induction in human cervical carcinoma cells, HeLa; human fibroblast cells, WI-38; and mouse fibroblast cells, L929. Thus, in diverse cell types, IL-1 induces gro-genes via tyrosine kinase-dependent signals. PMID: 7688736 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 731: FEBS Lett. 1993 Aug 16;328(3):225-9. Rapid alteration of c-myc and c-jun expression in leukemic cells induced to differentiate by a butyric acid prodrug. Rabizadeh E, Shaklai M, Nudelman A, Eisenbach L, Rephaeli A. Basil and Gerald Felsenstein Medical Research Center, Petach-Tikva, Israel. The novel prodrug of butyric acid (BA), pivaloyloxymethyl butyrate, has been shown, in vitro, to induce differentiation and inhibit leukemic cell proliferation. The prodrug affects the cells in vitro at lower concentration and at least 100 times faster than does (BA). We have compared the ability of BA with that of its prodrug AN-9 to modulate the expression of the early regulating genes, c-myc and c-jun, in HL-60 cells. Exposure of HL-60 cells to the prodrug resulted in a decrease of c-myc and an increase of c-jun expression. The prodrug elicited this effect at lower concentrations and at least 100 times faster than BA. Since changes in the expression of c-myc and c-jun occur minutes after exposure of the cells to the prodrug, these genes are likely to play a major role in the early stages of the differentiation pathway. PMID: 8348968 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 732: Cell Growth Differ. 1993 Aug;4(8):689-97. Erratum in: Cell Growth Differ 1993 Nov;4(11):955. Interleukin 6 induces DNA binding activity of AP1 in M1 myeloblastic cells but not in a growth resistant cell derivative. Melamed D, Resnitzky D, Haimov I, Levy N, Pfarr CM, Yaniv M, Kimchi A. Department of Molecular Genetics and Virology, Weizmann Institute of Science, Rehovot, Israel. The effects that three different growth inhibitory cytokines exert on expression and function of members of the Jun family were studied in this work. M1 myeloblastic cells were chosen for this purpose because of their high growth sensitivity to interleukin 6 (IL-6), transforming growth factor beta 1 and alpha- and beta-interferons. It is reported here that IL-6 elevated the junB and c-jun mRNA levels and induced the formation of a novel DNA-protein complex with high sequence specificity to 12-O-tetradecanoylphorbol-13-acetate response element (TRE) oligonucleotides. This IL-6 induced TRE binding complex was abolished by anti-Jun specific antibodies and was efficiently competed by an oligonucleotide that comprises the mouse homologue of a previously described human c-myc negative DNA element. It persisted in cells for at least 48 h after IL-6 treatment and failed to be induced by alpha- and beta-interferons or by transforming growth factor beta 1, which affected differently the pattern of jun mRNA expression. To further explore regulatory and functional aspects of this induced TRE binding activity, an IL-6 resistant M1 clone was isolated and further analyzed. This clone carried a postreceptor deficiency that abrogated completely the growth inhibitory responses to IL-6 but did not interfere with the induction of two differentiation related cell surface markers. Interestingly, the IL-6 resistant clone had lost two molecular responses to IL-6, induction of TRE binding activity and suppression of the c-myc gene. The data correlate the IL-6 induced AP1 activity with the suppression of c-myc and growth inhibition. PMID: 8398910 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 733: Am J Physiol. 1993 Aug;265(2 Pt 1):G331-8. Decreased expression of protooncogenes c-fos, c-myc, and c-jun following polyamine depletion in IEC-6 cells. Wang JY, McCormack SA, Viar MJ, Wang H, Tzen CY, Scott RE, Johnson LR. Department of Physiology, University of Tennessee College of Medicine, Memphis 38163. Direct exposure of small intestinal mucosal cells to luminal polyamines stimulates proliferation. This study tests the hypothesis that the protooncogenes c-fos, c-myc, c-jun, and junB are involved in the mechanism by which polyamines modulate mucosal growth. Studies were conducted in the IEC-6 cell line, derived from rat small intestinal crypt cells. Cells were grown in Dulbecco's minimal essential medium containing 5% dialyzed fetal bovine serum (dFBS) in the presence of absence of alpha-difluoromethylornithine (DFMO), a specific inhibitor of ornithine decarboxylase, which is the rate-limiting enzyme for polyamine synthesis. Cellular polyamine levels, cell growth, and relative abundance of c-fos, c-myc, c-jun, and junB mRNAs, were measured at 1, 2, 4, 6, 8, and 12 days after initial plating. The intracellular polyamines, spermidine and spermine, and their precursor, putrescine, in DFMO-treated cells decreased significantly at 2 days and remained depleted thereafter. Although DFMO profoundly decreased growth and final cell number, both control and DFMO-treated cells entered a plateau phase by 6 days. In control cells, c-myc and c-jun mRNA levels significantly increased on days 4-6 and then returned to a basal level of expression, which was maintained thereafter. c-fos mRNA in quiescent cells after 24 h serum deprivation was significantly stimulated by 5% dFBS, although a steady-state level of c-fos mRNA was undetectable in control cells. Treatment with DFMO not only prevented increased expression of c-myc and c-jun protooncogenes at 4 days, but also significantly reduced steady-state levels of c-myc and c-jun mRNA between 6 and 12 days.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 8368314 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 734: Leukemia. 1993 Aug;7 Suppl 2:S102-7. Reconstitution of functional human GM-CSF receptor in mouse NIH3T3 fibroblasts and BA/F3 proB cells. Yokota T, Watanabe S, Mui AL, Muto A, Miyajima A, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Granulocyte-macrophage colony-stimulating factor (GM-CSF) plays a critical role in growth and differentiation of myeloid cells. We previously reconstituted high affinity human GM-CSF receptor (hGM-CSFR) in a proB cell line BA/F3 by cotransfecting alpha and beta chain cDNA clones and showed that the reconstituted receptor could transduce growth promoting signals. The high affinity hGM-CSFR was also reconstituted in mouse NIH3T3 cells, but its ability to transduce signals in fibroblasts remained unanswered. In the present study, we further characterized signal transduction by the reconstituted hGM-CSFR both in NIH3T3 cells and BA/F3 cells. We found that the reconstituted hGM-CSFR transduces signals in NIH3T3 fibroblasts and BA/F3 cells in response to human GM-CSF to activate transcription of c-fos, c-jun and c-myc protooncogenes. hGM-CSF also induces protein tyrosine phosphorylation and DNA synthesis in both cell types. The ability of hGM-CSFR to transduce signals was affected by inhibitors of tyrosine kinase. These results indicated that the hGM-CSFR is functional in fibroblasts, that signal transduction via the hGM-CSFR in fibroblasts involves tyrosine kinase(s) and that association of hGM-CSFR with factor(s) specific to hematopoietic cell lineage is not essential to transduce growth promoting signals. PMID: 8361210 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 735: C R Acad Sci III. 1993 Aug;316(8):772-9. Transforming growth factor beta 1-mediated growth inhibition in chick embryo fibroblasts: reversion by virally-expressed nuclear oncogenes. Piu F, Jurdic P, Brun G, Samarut J, Castellazzi M. Laboratoire de Biologie Moleculaire et Cellulaire, Ecole Normale Superieure, UMR 49, CNRS, 46, Lyon, France. Transforming growth factor beta 1 (TGF-beta 1) inhibits growth of primary cultures of chick embryo fibroblasts by affecting G1 and strongly increasing the generation time. This inhibition is reversed by the nuclear oncogenes v-jun, v-fos, v-myc, but not v-erbA and v-ets. It is also reversed by v-myb from either avian myeloblastosis virus or avian E26 retrovirus. Taken together, these results strongly suggest that independent, functional interferences may take place between the TGF-beta 1-induced growth inhibitory pathway and the oncogen-driven stimulatory pathway(s) at the level of the AP-1, Myc, and Myb transcription factors. PMID: 8044700 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 736: Surgery. 1993 Aug;114(2):147-53; discussion 153-4. Comment in: Surgery. 1995 Mar;117(3):355. Glutamine is essential for epidermal growth factor-stimulated intestinal cell proliferation. Ko TC, Beauchamp RD, Townsend CM Jr, Thompson JC. Department of Surgery, University of Texas Medical Branch, Galveston 77555-0533. BACKGROUND. Glutamine stimulates growth of intestinal mucosa in vivo, but the mechanisms involved are unknown. The purpose of this study was to determine whether glutamine is essential for proliferation of enterocytes stimulated by epidermal growth factor (EGF). In addition, we determined which specific mitogenic actions of EGF require glutamine. METHODS. A nontransformed rat intestinal mucosal cell line (IEC-6) was stimulated with EGF (20 ng/ml) without and with glutamine (0.1 to 10 mmol/L). DNA, RNA, and protein synthesis were quantitated by determining incorporation of tritiated thymidine, tritiated uridine, and 14C-leucine, respectively. Cell numbers and messenger RNA levels of early growth response genes (zif268, jun-B, c-myc) were also determined. RESULTS. Glutamine was required for EGF stimulation of DNA, RNA, and protein synthesis and cell replication; however, EGF-stimulated expression of zif268, jun-B, and c-myc occurred in the absence of glutamine. CONCLUSIONS. This study showed that glutamine is essential for EGF-stimulated intestinal mucosal cell proliferation. The mitogenic effects of EGF can be divided into the glutamine-independent, such as the signal transduction pathway leading to the induction of early growth response genes, and the glutamine-dependent, including DNA, RNA, and protein synthesis. PMID: 7688149 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 737: Cancer Lett. 1993 Jul 30;71(1-3):125-32. Transforming growth factor beta 1-induced delay of cell cycle progression and its association with growth-related gene expression in mouse fibroblasts. Kim TA, Cutry AF, Kinniburgh AJ, Wenner CE. Department of Biochemistry, Roswell Park Cancer Institute, Buffalo, NY 14263. TGF beta-induced cell cycle progression is relatively slower than that induced by EGF or PDGF-BB. Further, TGF beta delays EGF or PDGF-induced 5-phase entry in C3H 10T1/2 mouse fibroblasts. In accordance with this delay, the induction of mRNA level of 'immediate early genes' such as c-myc, c-fos, c-jun and junB by TGF beta has slower kinetics compared with those of EGF. TGF beta induces c-sis gene, suggesting possible involvement of secondary growth stimulation by PDGF-like proteins. However, anti-PDGF-AB antibody, which was inhibitory to FDGF-BB-induced [3H]thymidine incorporation, did not block TGF beta-induced DNA synthesis. These results first demonstrate that the delay of cell cycle progression by TGF beta is closely associated with the altered regulation of growth-related gene expression in fibroblasts. PMID: 8364887 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 738: Toxicology. 1993 Jul 28;81(2):155-64. Cd(2+)-induced c-myc mRNA accumulation in NRK-49F cells is blocked by the protein kinase inhibitor H7 but not by HA1004, indicating that protein kinase C is a mediator of the response. Tang N, Enger MD. Department of Zoology and Genetics, Iowa State University, Ames 50011. Cd2+ is a toxic cation that, at sublethal and marginally lethal levels, modifies cell growth and metabolism. Cd2+ exposure of NRK-49F cells results in inhibition of early EGF-induced DNA synthesis, but induction of delayed DNA synthesis; in stimulation of anchorage independent growth; in accumulation of specific oncogene mRNAs; and in an hypertrophic response. Determining whether specific signal transduction pathways (STPs) are involved in specific gene deregulation by cadmium in NRK-49F cells is important to defining possible mechanisms by which Cd2+ elicits these physiological responses. In this study it is shown that Cd2+ induces delayed myc (8-10 h) and jun (12 h) mRNA accumulation, as well as both early (0.5-1 h) and late (12 h) fos but not TGF beta mRNA accumulation. The times of appearance of Cd(2+)-induced c-fos, c-myc and c-jun expression are dose dependent. The Cd2+ induced accumulation of these specific mRNAs is insensitive to cycloheximide and therefore not due to preinduction of TGF beta or other gene-activating growth factors, but rather to direct induction of oncogene expression and/or mRNA stabilization. Accumulation of c-myc mRNA is shown further to be inhibited by the protein kinase inhibitor H7 but not HA1004, indicating a role for one or more protein kinases C in the STPs by which Cd2+ induces oncogene expression. Thapsigargin, a compound which stimulates increased cytosolic [Ca2+], induces c-myc expression also by an H7 sensitive, HA1004 insensitive pathway. These results suggest that Cd2+ acts through one or more defined signal transduction pathways involving specific protein kinases C to induce the accumulation of c-fos, c-myc and c-jun messenger RNAs. PMID: 8378941 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 739: Mech Dev. 1993 Jul;42(1-2):49-58. Changes in protooncogene expression correlated with general and sex-specific differentiation in murine primordial germ cells. Coucouvanis EC, Jones PP. Department of Biological Sciences, Stanford University, CA 94305. Primordial germ cells (PGCs) of the mouse undergo key developmental transitions during embryonic days 12-15. On day 12 they complete migration into the gonads. They cease mitotic proliferation on day 13 and subsequently enter sex-specific pathways of development. The molecular mechanisms controlling these transitions are poorly understood, yet they are crucial to production of normal gametes later in life. We have used the polymerase chain reaction (PCR) to directly compare levels of expression of several protooncogenes proposed to be involved in control of cell proliferation and differentiation in proliferating and differentiating PCGs of both sexes over a 4 day time course. We report here that mRNA levels for nuclear protooncogenes c-myc, c-fos, and c-jun increase dramatically in both sexes from little or no detectable expression on day 12 to high expression on days 13-15. We observe c-kit message on day 12 in combined PGCs of both sexes, in female but not male PGCs on day 13, and in both sexes on day 14, c-kit mRNA is undetectable on day 15 in either sex, c-mos is not expressed at detectable levels on day 12 in either sex, but increases gradually in female PGCs to very high levels on day 15. In male PGCs, c-mos is expressed at high levels on days 13-15. Our results are consistent with a role for protooncogenes c-myc, c-fos and c-jun in mediating the initial differentiation of PGCs of both sexes that occurs upon colonization of the gonad. Because c-kit and c-mos are expressed differentially in male and female day 13-15 germ cells, they may play roles in initiating or mediating progress along the sex-specific pathways of development that PGCs embark upon at this time. PMID: 8369223 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 740: Cardiovasc Res. 1993 Jul;27(7):1209-13. Heparin does not inhibit oncogene induction in rabbit aorta following balloon denudation. Hamon M, Bauters C, Wernert N, Courtin P, Delcayre C, Adamantidis M, Lablanche JM, Bertrand ME, Dupuis B, Swynghedauw B. Department of Pharmacology, University of Lille, France. OBJECTIVE: Smooth muscle cell proliferation and migration are the predominant responses to intimal and medial injury after percutaneous transluminal coronary angioplasty. The in vivo inhibitory effect of heparin on these responses is well documented. To test the hypothesis that the antiproliferative effect of heparin in vivo may be related to an inhibition of proto-oncogene expression, the effects of pretreatment with heparin on the expression of the c-myc, c-fos and c-jun proto-oncogenes were examined in a rabbit model of balloon denudation. METHODS: Animals were randomised 5 h before balloon denudation to receive a subcutaneous injection of unfractionated heparin (7500 IU.kg-1, n = 7) or saline (n = 6). Total RNA extracted from the aorta 1 h after balloon denudation was analysed by northern blot technique. A histological study was also performed in saline treated (n = 4) and heparin treated (n = 4) animals 28 d after balloon denudation. RESULTS: The histological study showed that the degree of neointimal thickening was significantly less in heparin treated animals. However, the level of expression of the proto-oncogenes we studied was similar in both groups. CONCLUSIONS: Heparin inhibits neointimal thickening after balloon denudation. This inhibition is not associated with an overall decrease in the level of expression of the c-myc, c-fos, or c-jun proto-oncogenes in the arterial wall, suggesting that the antiproliferative effect of heparin may be due to an effect on other events in the cell cycle. PMID: 8252580 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 741: Clin Exp Metastasis. 1993 Jul;11(4):325-9. Expression of oncoproteins in primary human non-small cell lung cancer and incidence of metastases. Volm M, Drings P, Wodrich W, van Kaick G. German Cancer Research Center, Heidelberg. In the current study the relationship between the incidence of metastatic spread and expression (at the protein level) of various proto-oncogenes was investigated in 217 human non-small cell lung carcinomas. Tumors with an overexpression of proteins encoded by the oncogenes c-jun and c-myc showed a significantly increased formation of metastases (c-jun: P = 0.008; c-myc: P = 0.018). No significant correlations were found between the expression of the c-fos, c-erbB1, c-neu and c-ras products and metastatic spread. PMID: 8100491 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 742: Biochim Biophys Acta. 1993 Jun 30;1177(3):307-17. The competence progression model in CHO-K1 cells: the relationship between protein kinase C and immediate early gene expression in the insulin mitogenic signal. Ross S, Englesberg E. Department of Biological Sciences, University of California, Santa Barbara. CHO-K1 cells grow in a defined medium with insulin, at physiological concentrations, as the only hormone. IGF-I can substitute for insulin. Quiescent cells require a 9-10-h lag, subsequent to the addition of insulin, to synthesize DNA. The phorbol ester, 12-O-tetradeconoylphorbol 13-acetate (TPA), cannot support growth of these cells, is a more effective inducer than insulin of c-fos, c-myc, c-jun, jun-B, Krox-20, Krox 24, fra-1 and JE, and induces fra-1, JE and c-myc with different kinetics from those of insulin. The addition of insulin + TPA to quiescent cells produces a synergistic effect on DNA synthesis but not on the expression of immediate early genes. Pretreatment of these cells with TPA or insulin decreases the required lag time for DNA synthesis by 3 h in a protein-synthesis-independent manner. These results, together with other experiments, demonstrate that [1] the insulin signal is independent of PKC, [2] insulin acts as a weak competence and a strong progression factor, while TPA behaves as a strong competence factor, and [3] the 9-10-h lag is made up of a 3-h period which is independent of protein synthesis, advancing the cells to a post-G(o) state of 'competence'. PMID: 8323980 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 743: Hepatology. 1993 Jun;17(6):1109-16. Differences in the steady-state levels of c-fos, c-jun and c-myc messenger RNA during mitogen-induced liver growth and compensatory regeneration. Coni P, Simbula G, de Prati AC, Menegazzi M, Suzuki H, Sarma DS, Ledda-Columbano GM, Columbano A. Istituto di Patologia Sperimentale, University of Cagliari, Italy. The steady-state levels of c-fos, c-jun and c-myc messenger RNA were investigated in rat liver tissue after proliferative stimuli of different nature-namely, compensatory regeneration induced by partial hepatectomy or carbon tetrachloride administration-and direct hyperplasia induced by four different hepatomitogens: lead nitrate, ethylene dibromide, cyproterone acetate and nafenopin. We show here that whereas c-fos and c-jun expression increased soon after partial hepatectomy or carbon tetrachloride administration, an increased expression of c-jun in the absence of c-fos expression occurred during direct hyperplasia induced by lead nitrate and ethylene dibromide. When hyperplasia was induced by cyproterone acetate and nafenopin, the mitogenic response of the liver was not associated with an increased expression of c-jun or c-fos, despite the fact that the timing of the cell cycle was similar to that observed after partial hepatectomy. Finally, when c-myc expression was analyzed, it was found that proliferative conditions associated with an increased expression of this gene were characterized by an increased expression of c-jun. On the contrary, the hyperplasia induced by cyproterone acetate and nafenopin, which is characterized by a lack of increase in the expression of c-fos and c-jun, was also not associated with an increased c-myc expression. Similar results were obtained in these experiments with the mitogen nafenopin, a peroxisome proliferator. In fact, liver hyperplasia induced by this compound was not preceded or accompanied by an increased expression of c-fos and c-myc. This study suggests that depending on the nature of the proliferative stimulus, an increased expression of c-fos, c-jun and c-myc may not be necessary for in vivo induction of liver cell proliferation. PMID: 8514261 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 744: Carcinogenesis. 1993 Jun;14(6):1121-4. Overexpression of oncoproteins in non-small cell lung carcinomas of smokers. Wodrich W, Volm M. German Cancer Research Center, Heidelberg. Non-small cell lung carcinoma specimens of 173 previously untreated patients were analyzed for the expression of proteins encoded by the oncogenes c-myc, c-fos, c-jun, c-erbB-1, c-erbB-2, c-H-ras, c-K-ras and c-N-ras. Forty-six per cent of the tumors were positive for the c-MYC protein, 60% for c-FOS, 50% for c-JUN, 80% for c-ERBB-1, 55% for c-ERBB-2, 12% for c-H-RAS, 5% for c-K-RAS and 71% for c-N-RAS. Proteins encoded by c-fos and c-jun are overexpressed more frequently in carcinomas of smokers (c-fos: P < 0.005; c-jun: P < 0.01). When we grouped the patients according to their tumor histology the results became more evident. Squamous cell lung carcinomas of smokers showed a higher incidence of c-FOS (P = 0.01), c-JUN (P < 0.01) and c-ERBB-1 (P = 0.01) proteins than squamous cell lung carcinomas of non-smokers. The expression rate and the intensity of staining proved not to be influenced either by the number of cigarettes smoked daily or by cessation of smoking. In adenocarcinomas, however, we only found a trend for a more frequent overexpression of c-fos (P = 0.07) and c-jun (P = 0.14) encoded proteins in carcinomas of smokers and no correlation between the expression of c-erbB-1 products and smoking. No correlation was found between the expression of c-MYC, c-ERBB-2, c-H-RAS, c-K-RAS and c-N-RAS proteins and the smoking habits of the patients, neither in squamous cell carcinomas nor in adenocarcinomas of the lung. PMID: 8389672 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 745: J Immunol. 1993 Jun 1;150(11):4822-32. Induction of early response genes by cross-linking membrane Ig on B lymphocytes. Mittelstadt PR, DeFranco AL. Department of Microbiology and Immunology, University of California, San Francisco 94143-0552. Cross-linking of membrane Ig (mIg) on B lymphocytes induces protein tyrosine phosphorylation and phosphoinositide hydrolysis, events that are thought to mediate the diverse biologic responses of B cells to Ag binding. mIg stimulation also induces the expression of the putative transcriptional regulators c-myc, c-fos, egr-1, and jun-B. In this report, normal murine B cells and two murine B lymphoma cell lines were examined for the induction of mRNA expression of seven early response genes first identified in fibroblasts. Expression of four of the seven genes (nur77, nup475, pip92, and 3CH134), encoding two putative transcriptional regulators, a protein of unknown function, and a putative protein phosphatase, was induced after cross-linking of mIg in resting B cells isolated from mouse spleen. In the 2PK-3 and WEHI-231 B lymphoma cell lines three and two, respectively, of these four genes were induced. Expression of these genes could be induced in 2PK-3 cells by activating the phosphoinositide-signaling pathway independently of the tyrosine phosphorylation pathway by signaling through an M1 muscarinic acetylcholine receptor introduced by transfection. Additionally, in all but one case, these early response genes could be induced by directly activating protein kinase C with phorbol esters. In the cell line 2PK-3, the gene 3CH134 was not induced by phorbol ester treatment, but was induced by elevation of intracellular calcium. Thus, a subset of the early response genes identified in serum-stimulated fibroblasts is also induced by Ag-receptor stimulation in B lymphocytes, and this induction appears to be mediated by the phosphoinositide signaling pathway and, for the most part, protein kinase C. PMID: 8388422 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 746: Cell Struct Funct. 1993 Jun;18(3):151-60. Cytoskeletal active drugs modulate signal transduction in the protein kinase C pathway. Pavlath GK, Shimizu Y, Shimizu N. Department of Molecular and Cellular Biology, University of Arizona, Tucson 85721. The cytoskeletal network of cells is postulated to play a role in the signal transduction pathways of growth promoting stimuli. We show here that cytoskeletal active drugs modulate the mitogenic signal transduction pathway of the tumor promoter TPA in 3T3-L1 cells. Compounds which act on microtubules (vinblastine sulfate) and microfilaments (cytochalasin B) have opposite effects on DNA synthesis. Vinblastine sulfate leads to stimulation, whereas cytochalasin B causes potent inhibition of DNA synthesis in response to TPA. These drugs are cell cycle specific and apparently exert their regulatory action distal to activation of protein kinase C by TPA. The expression of four genes necessary for DNA synthesis in response to tumor promoters was examined: two nuclear proto-oncogenes (c-myc and c-fos), a transcription factor (c-jun/AP-1) and a key enzyme involved in polyamine synthesis (ornithine decarboxylase). c-jun mRNA levels are not modulated during the action of cytoskeletal disrupting drugs on TPA-mediated mitogenesis, whereas c-myc and c-fos mRNA levels are similarly enhanced. Expression of ornithine decarboxylase mRNA and protein is increased by vinblastine sulfate but decreased by cytochalasin B in TPA treated cells. These data indicate that changes in cytoskeletal organization may play a role in regulating the levels of an enzyme critical for DNA synthesis. PMID: 8242794 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 747: Biochem Biophys Res Commun. 1993 May 28;193(1):356-63. Cloning of the mouse interleukin 2 receptor gamma chain: demonstration of functional differences between the mouse and human receptors. Kumaki S, Kondo M, Takeshita T, Asao H, Nakamura M, Sugamura K. Department of Microbiology, Tohoku University School of Medicine, Sendai, Japan. We isolated a cDNA clone for the gamma chain of the mouse interleukin 2 receptor. Introduction of the mouse gamma chain cDNA clone into a mouse fibroblast cell line, L929, expressing the mouse alpha beta heterodimer IL-2 receptor converted pseudo-high affinity of the IL-2 receptor into functional high, resulting in internalization of IL-2 and induction of the c-myc, c-fos and c-jun genes. The mouse beta gamma heterodimer, however, failed to bind IL-2 unlike the human beta gamma heterodimer intermediate-affinity receptor. These results indicate that the mouse functional IL-2 receptor is a complex comprising three distinct subunits, alpha, beta and gamma chains, but the beta gamma heterodimer is not functional and different from the human heterodimer. PMID: 8503926 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 748: Blood. 1993 May 15;81(10):2539-46. Mechanism of differential inhibition of factor-dependent cell proliferation by transforming growth factor-beta 1: selective uncoupling of FMS from MYC. Chen AR, Rohrschneider LR. Department of Cell Biology, Fred Hutchinson Cancer Research Center, Seattle, WA 98104. Transforming growth factor-beta 1 (TGF-beta 1) selectively modulates hematopoietic cell proliferation. The proliferation of FDC-P1 clone MAC-11, a factor-dependent murine myeloid progenitor cell line, was inhibited differentially by TGF-beta 1: strongly in macrophage colony-stimulating factor (M-CSF), mildly in interleukin-3, and not at all in granulocyte-macrophage-CSF (GM-CSF). Flow cytometry and Western blots showed an unexpected increase in expression of FMS, the receptor for M-CSF, in response to TGF-beta 1. Metabolic labeling with 35S-methionine showed that synthesis of FMS protein accelerated in response to TGF-beta 1, whereas its degradation was unaffected. Northern analyses showed a rapid increase in c-fms RNA after the addition of TGF-beta 1. TGF-beta 1 did not affect kinase activity, cellular phosphotyrosine response, or internalization of FMS. However, TGF-beta 1 inhibited the induction by M-CSF of c-myc RNA analyzed on Northern blots and protein detected by radioimmuno-precipitation. TGF-beta 1 did not affect induction of c-myc expression by GM-CSF or induction of c-fos or c-jun by M-CSF. Therefore, FMS and the GM-CSF receptor induce c-myc via signal transduction pathways that differ in that only the former is inhibited by TGF-beta 1. This inhibition may account for the selective growth regulation by TGF-beta 1. PMID: 8490168 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 749: Int J Cancer. 1993 May 8;54(2):348-54. FK506 and cyclosporin a regulate proliferation and proto-oncogene expression in HTLV-1-associated myelopathy/tropical-spastic-paraparesis-derived T cells. Natazuka T, Umemiya-Okada T, Matsui T, Saida T, Nakao Y. Department of Medicine, Kobe University School of Medicine, Japan. Human T-cell-leukemia-virus-type-1 (HTLV-1) infection is associated with adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy (HAM)/tropical spastic paraparesis (TSP). The T-cell-targeting immunosuppressants, FK506 and cyclosporin A (CsA), suppressed proliferation of the HAM/TSP-derived T-cell lines, H89-59, H89-79 and H109. FK506 and CsA also reduced expression of the proto-oncogenes, c-myc and c-fos, but not c-jun and interleukin-2-receptor-alpha (IL-2R alpha) gene in H109 cells. The growth-inhibitory effects of FK506 and CsA were not abrogated by interleukin 2 (IL-2). These results suggest that the inhibitory effects of FK506 and CsA are independent of IL-2, and are associated with the reduction of c-myc and c-fos gene expression. PMID: 7683633 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 750: Mol Pharmacol. 1993 May;43(5):709-14. Tamoxifen stimulates expression of the c-fos proto-oncogene in rodent uterus. Kirkland JL, Murthy L, Stancel GM. Department of Pediatrics, Baylor College of Medicine, Houston, Texas 77030. Estrogens regulate the in vivo expression of the c-fos proto-oncogene in rat uterus, and this regulation appears to occur at the transcriptional level. This system thus provides the ability to study the in vivo effects of antiestrogens on specific gene expression in normal estrogen target tissue. Immature rats were treated with estradiol, tamoxifen, or other nonsteroidal antiestrogens, total uterine RNA was isolated, and c-fos transcript levels were monitored by blot analysis. Tamoxifen increases the 2.2-kilobase c-fos transcript approximately 20-fold in 6 hr. This effect is comparable in magnitude to that produced by estradiol, but the maximum response to the hormone occurs in 3 hr. c-fos induction is observed at doses of 0.1-10 mg/kg tamoxifen. The nonsteroidal antiestrogens nafoxidine, Cl-628, and 4-hydroxytamoxifen also induce c-fos expression. The induction of c-fos by both estradiol and tamoxifen is blocked by the progestin medroxyprogesterone acetate. In addition to effects on c-fos mRNA, tamoxifen also increases uterine levels of c-jun, jun-B, and c-myc mRNAs. These results indicate that tamoxifen acts in vivo as an estrogen agonist for activating expression of cellular oncogenes in normal uterine tissue. PMID: 8502228 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 751: J Clin Invest. 1993 May;91(5):2268-74. Multiple autocrine growth factors modulate vascular smooth muscle cell growth response to angiotensin II. Itoh H, Mukoyama M, Pratt RE, Gibbons GH, Dzau VJ. Falk Cardiovascular Research Center, Stanford University School of Medicine, California 94305-5246. Angiotensin (Ang) II stimulates hypertrophic growth of vascular smooth muscle cells (VSMC). Accompanying this growth is the induction of the expression of growth-related protooncogenes (c-fos, c-jun, and c-myc), as well as the synthesis of the autocrine growth factors, such as PDGF-A and TGF-beta 1. In this study, we demonstrate further that Ang II also induces the synthesis of basic fibroblast growth factor (bFGF), a potent mitogen for VSMC. To examine how these factors interact to modulate the growth response of VSMC to Ang II, we used antisense oligomers to determine the relative contribution of these three factors. Treatment of confluent, quiescent smooth muscle cells with specific antisense oligomers complementary to bFGF, PDGF-A, and TGF-beta 1 efficiently inhibited the syntheses of these factors. Our results demonstrate that in these VSMC, TGF-beta 1 affects a key antiproliferative action, modulating the mitogenic properties of bFGF. Autocrine PDGF exerts only a minimal effect on DNA synthesis. An imbalance in these signals activated by Ang II may result in abnormal VSMC growth leading to the development of vascular disease. PMID: 8486785 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 752: Proc Natl Acad Sci U S A. 1993 May 1;90(9):4127-31. Reconstitution of functional interleukin 2 receptor complexes on fibroblastoid cells: involvement of the cytoplasmic domain of the gamma chain in two distinct signaling pathways. Asao H, Takeshita T, Ishii N, Kumaki S, Nakamura M, Sugamura K. Department of Microbiology, Tohoku University School of Medicine, Sendai, Japan. We have previously shown that the interleukin 2 (IL-2) receptor gamma chain is a member of the cytokine receptor superfamily and is indispensable for the formation of receptor complexes with high and intermediate affinities for IL-2. The present study demonstrates that the alpha beta gamma heterotrimer and beta gamma heterodimer complexes of IL-2 receptor reconstituted on murine fibroblast L929 cells can transduce IL-2-mediated signals for activation of tyrosine kinase and for induction of c-myc, c-fos, and c-jun expression. A mutant of the gamma chain lacking the C-terminal 68 amino acids in its cytoplasmic region showed a loss of such signal-transducing ability when incorporated into the IL-2 receptor complexes but brought no effect on IL-2 binding and IL-2 internalization. Another mutant, with a C-terminal deletion of 30 amino acids, retained the ability to activate a tyrosine kinase and to induce c-myc expression but lost the ability to induce c-fos and c-jun expression. These results suggest that at least two distinct signals, one for c-myc induction, which parallels tyrosine kinase activation, and the other for c-fos and c-jun induction, can be transduced from the IL-2 receptor complexes reconstituted on fibroblastoid cells. PMID: 7683423 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 753: J Immunol. 1993 Apr 15;150(8 Pt 1):3109-18. Effect of transforming growth factor-beta on early and late activation events in human T cells. Ahuja SS, Paliogianni F, Yamada H, Balow JE, Boumpas DT. Kidney Disease Section, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892. Transforming growth factors-beta (TGF-beta) modulate immune responses by inhibiting the proliferation of normal T lymphocytes. To examine the mechanism(s) of this inhibition, we studied the effect of TGF-beta 1 on selected events associated with the initiation and progression of the T lymphocyte cell cycle. Human peripheral blood T cells were stimulated with anti-CD3 mAb, PHA, PMA, or ionomycin, alone or in combination. TGF-beta 1 (0.5 to 10 ng/ml) partially inhibited the tyrosine phosphorylation of a 100-kDa protein, but not the calcium influx when cells were stimulated via TCR. Nuclear transcription of early activation genes (c-fos, c-jun, and c-myc) as determined by nuclear run-off assays, and steady state mRNA levels and/or protein products of intermediate activation genes (IL-2, IL-2R alpha, IL-2R beta, and transferrin receptor) were not affected by TGF-beta 1. Total cellular RNA synthesis and cell size after T cell stimulation were also not affected by TGF-beta 1. However, TGF-beta 1 inhibited the IL-2-dependent proliferation of Con A lymphoblasts by -50%. This inhibition was associated with the down-regulation of IL-2-mediated tyrosine phosphorylation of proteins of 120, 100, 85, 75, and 50 kDa. TGF-beta 1 also inhibited the IL-2-dependent phosphorylation of the retinoblastoma susceptibility gene product, which plays an important role in cell cycle progression. These results suggest that TGF-beta 1 inhibits T cell proliferation by down-regulating predominantly IL-2-mediated proliferative signals. PMID: 8468460 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 754: J Cell Biochem. 1993 Apr;51(4):381-6. Effects of electromagnetic field exposure on gene transcription. Phillips JL. Pettis Memorial Veterans Administration Medical Center, Loma Linda, California 92357. Exposure of whole animals, isolated tissues, and cells to electromagnetic fields of various characteristics has resulted in a substantial literature detailing a wide range of effects at the morphological, physiological, biochemical, and molecular levels. In recent years, considerable effort has been devoted to defining a mechanism by which electromagnetic fields can couple to biological systems and generate this plethora of effects. As a consequence, there has been a growing interest in electromagnetic field-induced alterations in gene expression. Key studies are discussed which indicate that exposure of several cell types to electromagnetic fields that differ in waveform, amplitude, and frequency induced general changes in gene transcription. Moreover, exposure of T-lymphoblastoid cells to a 60 Hz sinusoidal magnetic field altered the transcription of genes encoding c-fos, c-jun, c-myc, and protein kinase C. Future studies in this area should focus on independent replication of key studies and identification of which events in the signal transduction pathways leading to gene transcription are altered by electromagnetic field exposure. Publication Types: Review Review, Tutorial PMID: 8496241 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 755: Hum Genet. 1993 Apr;91(3):217-22. Chromosomal localization of four human zinc finger cDNAs. Huebner K, Druck T, LaForgia S, Lasota J, Croce CM, Lanfrancone L, Donti E, Pengue G, La Mantia G, Pelicci PG, et al. Jefferson Cancer Institute, Thomas Jefferson Medical College, Philadelphia, PA 19107. cDNA clones encoding zinc finger motifs were isolated by screening human placenta and T-cell (Peer) cDNA libraries with zinc finger (ZNF) consensus sequences. Unique cDNA clones were mapped in the human genome by rodent-human somatic cell hybrid analysis and in some cases in situ chromosomal hybridization. ZNF80 mapped to 3p12-3qter, ZNF7 was previously mapped to 8q24 and is here shown by in situ hybridization and use of appropriate hybrids to map telomeric to the MYC locus. ZNF79 mapped to 9q34 centromeric to the ABL gene and between a constitutional chromosomal translocation on the centromeric side and the CML specific ABL translocation on the telomeric side. ZNF77 mapped to 19p while ZNF78L1 (pT3) mapped to 19q. Chromosome 19 carries many ZNF loci and other genes with zinc finger encoding motifs; the pT3 clone additionally detected a locus designated ZNF78L2, which mapped to chromosome region 1p, most likely in the region 1p32 where the MYCL and JUN loci map. PMID: 8478004 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 756: Br J Cancer. 1993 Apr;67(4):674-9. Uncoupling of growth inhibition and differentiation in dexamethasone-treated human rhabdomyosarcoma cells. De Giovanni C, Lollini PL, Dolcetti R, Landuzzi L, Nicoletti G, D'Andrea E, Scotland K, Nanni P. Istituto di Cancerologia, University of Bologna, Italy. The effects of dexamethasone, a synthetic glucocorticoid, and of N,N-dimethylformamide on in vitro growth and differentiation and on proto-oncogene expression of human rhabdomyosarcoma cells were studied. RD/18 clone cells (derived from the embryonal rhabdomyosarcoma cell line RD) treated with 100 nM dexamethasone showed an almost complete block of differentiation: about 5% myosin-positive cells were observed after 2 weeks of culture in dexamethasone-supplemented differentiation medium, compared to 20% of untreated cultures. Dexamethasone also induced a 20-30% growth inhibition and a more flattened morphology. The treatment with N,N-dimethylformamide induced a significantly increased proportion of myosin-positive cells (reaching about 30%) and a 40% growth inhibition. Induction of differentiation inversely correlated with the levels of c-myc proto-oncogene expression: after a 2 week culture dexamethasone-treated cells showed the highest c-myc expression and N,N-dimethylformamide-treated cells the lowest. Culture conditions per se down-modulated c-erbB1 and up-regulated c-jun expression, with no relationship to the differentiation pattern. Other proto-oncogenes were not expressed (c-sis, N-myc, c-mos, c-myb) or were not modulated (c-fos, c-raf). Therefore dexamethasone and N,N-dimethylformamide, both causing a decreased growth rate, showed opposing actions on myogenic differentiation and on c-myc proto-oncogene expression of human rhabdomyosarcoma cells. PMID: 8471424 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 757: Endocrinology. 1993 Apr;132(4):1729-34. Mitogenic signaling by prostaglandins in chemically transformed mouse fibroblasts: comparison with phorbol esters and insulin. Fagot D, Buquet-Fagot C, Mester J. INSERM U.55, Paris, France. In the quiescent mouse BP-A31 fibroblasts, prostaglandin F2 alpha (PGF2 alpha) induces the expression of cell cycle-related genes c-fos, c-jun, and c-myc, and after a delay of approximately 12 h the entry into the phase of DNA replication. A weaker mitogenic effect was produced by certain other PGs (F1 alpha > D2), whereas the effects of PGs E and I were marginal or absent. The mitogenic effects of PGF2 alpha as well as of 12-O-tetradecanoyl phorbol 13-acetate (TPA; activator of protein kinase C) but not those of insulin (acting via the insulin-like growth factor 1 receptor) were abolished by a low concentration (7.5 nM) of staurosporine (inhibitor of protein kinase C). Moreover, long-time (24 h) preincubation with phorbol dibutyrate reduced the mitogenic effects of a subsequent exposure either TPA or PGF2 alpha. These observations favor the involvement of protein kinase C in the PGF2 alpha-dependent intracellular signal transduction. However, simultaneous stimulation of the quiescent cells with saturating concentrations of PGF2 alpha and TPA had a greater mitogenic effect than either drug alone, both in cells with and without down-regulation of protein kinase C, indicating that the protein kinase C-dependent signaling does not entirely account for the mitogenic activity of PGF2 alpha. PMID: 8462473 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 758: Leukemia. 1993 Apr;7(4):563-8. Taxol induces internucleosomal DNA fragmentation associated with programmed cell death in human myeloid leukemia cells. Bhalla K, Ibrado AM, Tourkina E, Tang C, Mahoney ME, Huang Y. Department of Medicine, Medical University of South Carolina, Charleston 29425. The present results demonstrate that the exposure of human myeloid leukemia HL-60 and KG-1 cells to clinically achievable concentrations of taxol produced internucleosomal DNA fragmentation of approximately 200 base-pair multiples, and the morphologic changes characteristic of cells undergoing programmed cell death (PCD) or apoptosis. Taxol-induced PCD was associated with a marked inhibition of suspension culture growth and clonogenic survival of HL-60 cells. In addition, taxol treatment decreased BCL-2 oncogene expression, which is known to block PCD. The exposure to taxol moderately decreased c-myc expression, but did not induce c-jun expression--which has been previously noted for a variety of DNA interactive, antileukemic drugs. These findings indicate that taxol may induce leukemic cell death partly by the alternative but gene-directed and active mechanism of PCD. PMID: 8096557 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 759: Liver. 1993 Apr;13(2):102-9. Activation of nuclear protooncogenes and alpha-fetoprotein gene in rat liver during the acute inflammatory reaction. Bernuau D, Moreau A, Tournier I, Legres L, Feldmann G. Laboratoire de Biologie Cellulaire, INSERM U 327, Faculte de Medecine Xavier-Bichat, Paris, France. Nuclear protooncogene and alpha-fetoprotein gene expression is stimulated in hepatocytes during liver regeneration and by various growth factors in vitro. Metabolic adaptation of hepatocytes has been implicated in such gene reprogrammation. We examine here whether induction of an acute inflammation, a physiological situation of important metabolic adjustments, also triggers activation of nuclear oncogenes and of the AFP gene in rat liver. C-fos, c-jun and c-myc mRNA accumulated on Northern blots between 4-12 h of inflammation and the steady-state level of two small alpha-fetoprotein transcripts characteristic of the adult liver increased at 4 h and 24 h of inflammation. In situ hybridization showed accumulation of the mRNA of the four genes studied in all hepatocytes, without any zonal lobular heterogeneity. 3H-histoautoradiography and mitotic counts indicated an inhibition of DNA synthesis and mitosis, prolonged for at least 48 h after inflammation. Thus acute inflammation triggers the activation of nuclear protooncogenes and alpha-feto-protein gene in hepatocytes, but this activation is not followed by passage into the replicative cycle. PMID: 7685462 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 760: Cardioscience. 1993 Mar;4(1):15-20. Increased nuclear proto-oncogene expression in hypertrophic cardiomyopathy. Hengstenberg C, Maisch B. Cardiology Department, Philipps-University of Marburg, Germany. Hypertrophic cardiomyopathy is characterized by an unexplained hypertrophy of the left ventricle, particularly the interventricular septum. Although point mutations in the beta-myosin chain gene have been found in several US families in familiar hypertrophic cardiomyopathy, the pathogenetic pathways leading to myocyte hypertrophy, the most important feature, are still not clear. To examine whether activation (expression) of nuclear proto-oncogenes may play a role in hypertrophic cardiomyopathy, endomyocardial biopsies from 13 patients with hypertrophic cardiomyopathy were examined using monoclonal antibodies against c-myc, c-fos and c-jun. The nuclear proto-oncogenes c-fos, c-jun and c-myc were expressed in 53, 60, and 50%, respectively, of patients with hypertrophic cardiomyopathy. In control biopsies, c-myc was detectable in only 10% of the patients, while c-fos and c-jun were always undetectable. These results show that nuclear proto-oncogenes are induced in patients with hypertrophic cardiomyopathy, although the triggering mechanisms remain unknown. PMID: 8471737 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 761: Am J Physiol. 1993 Mar;264(3 Pt 2):H760-9. ANG II receptors, c-myc, and c-jun in myocytes after myocardial infarction and ventricular failure. Reiss K, Capasso JM, Huang HE, Meggs LG, Li P, Anversa P. Department of Medicine, New York Medical College, Valhalla 10595. To determine the relationship between reactive cardiac hypertrophy and the expression of angiotensin II (ANG II) receptors in surviving myocytes after infarction, large infarcts were produced in rats that were killed 2-3 days later. Measurements of global ventricular dynamics indicated that left ventricular failure and right ventricular dysfunction occurred in experimental animals. These alterations in ventricular pump function were associated with increases in ventricular weight-to-body weight ratio, indicative of developing cardiac hypertrophy. Such a response was coupled with a 6.6-fold increase in ANG II receptor mRNA in myocytes from the left ventricle. A 2.3-fold increase in the expression of ANG II receptor in myocytes from the right ventricle was also found. Radioligand binding assay documented a 44% increase in the density of ANG II receptors on left ventricular myocytes of infarcted hearts. To establish whether the induction of genes commonly associated with myocyte hypertrophy was present, the message for c-myc and c-jun was biventricularly assessed. Myocardial infarction was accompanied by overexpressions of c-myc and c-jun that were more prominent in left than in right ventricular myocytes. In conclusion, the enhanced expression of ANG II receptor and its receptor protein and c-myc and c-jun in myocytes may participate in the reactive growth processes of these cells after infarction. PMID: 8456979 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 762: Mol Cell Biol. 1993 Mar;13(3):1440-8. Reconstituted human granulocyte-macrophage colony-stimulating factor receptor transduces growth-promoting signals in mouse NIH 3T3 cells: comparison with signalling in BA/F3 pro-B cells. Watanabe S, Mui AL, Muto A, Chen JX, Hayashida K, Yokota T, Miyajima A, Arai K. Department of Molecular and Developmental Biology, University of Tokyo, Japan. Granulocyte-macrophage colony-stimulating factor (GM-CSF) plays a critical role in growth and differentiation of myeloid cells. We previously reconstituted high-affinity human GM-CSF receptor (hGM-CSFR) in a pro-B cell line, BA/F3, by cotransfecting alpha- and beta-chain cDNA clones and showed that the reconstituted receptor could transduce growth-promoting signals. The high-affinity hGM-CSFR was also reconstituted in mouse NIH 3T3 cells, but its ability to transduce signals in fibroblasts remained undetermined. In the present study, we further characterized signal transduction by the reconstituted hGM-CSFR in both NIH 3T3 cells and BA/F3 cells. We found that the reconstituted hGM-CSFR transduces signals in NIH 3T3 fibroblasts and BA/F3 cells in response to hGM-CSF to activate transcription of the c-fos, c-jun, and c-myc proto-oncogenes. hGM-CSF also induces protein tyrosine phosphorylation and DNA synthesis in both cell types. These results indicated that hGM-CSFR is functional in fibroblasts, that signal transduction via hGM-CSFR in fibroblasts involves tyrosine kinase(s), and that association of hGM-CSFR with a factor(s) specific to hematopoietic cell lineage is not essential to transduce growth-promoting signals. PMID: 8441389 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 763: Br J Cancer. 1993 Mar;67(3):514-21. Temporal sequence and cellular origin of interleukin-2 stimulated cytokine gene expression. Saraya KA, Balkwill FR. Biological Therapy Laboratory, Imperial Cancer Research Fund, Lincoln's Inn Fields, UK. A study of activation of the cytokine network by interleukin 2, IL-2, may provide a rationale for devising cytokine combination and cytokine antagonist treatments with increased anti-tumour efficacy and decreased toxicity. We have investigated the expression of mRNA for 13 cytokines and three transcription factors during in vitro culture of peripheral blood mononuclear cells, PBMC, with IL-2. A consistent pattern of induction was seen in nine individuals, with early (2-24 h) induction of IL-1 beta, IL-6, tumour necrosis factor, TNF, lymphotoxin, LT, and gro. TNF and LT mRNA was expressed continually throughout culture, but levels of mRNA for IL-1 beta, IL-6, and gro declined by 24-48 h. After 48 h, PBMC began to express mRNA for IFN-gamma, IL-5, GM-CSF, and M-CSF. At 15 min to 1 h post IL-2 mRNA for c-fos, c-jun, and c-myc, and TNF was induced in three individuals studied. IL-4, IFN-alpha, and IL-1 alpha mRNA was not detected. Only a minority of cells expressed mRNA for TNF, IL-1 beta, IL-6 and IFN-gamma, and monocytes were the main source. Levels of cytokine protein in culture supernatants mirrored the pattern of mRNA induction. This in vitro model shows clear parallels with the reported in vivo production of cytokines during IL-2 therapy, and may prove useful in designing new therapeutic strategies. PMID: 8439502 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 764: Curr Probl Cancer. 1993 Mar-Apr;17(2):69-141. Small cell lung cancer: etiology, biology, clinical features, staging, and treatment. Cook RM, Miller YE, Bunn PA Jr. Department of Medicine, University of Colorado Health Sciences Center, Denver. Lung cancer is the leading cause of cancer death in the United States. Small cell lung cancer (SCLC) accounts for 20% to 25% of all bronchogenic carcinoma and is associated with the poorest 5-year survival of all histologic types. SCLC differs in its etiologic, pathologic, biologic, and clinical features from non-SCLC, and these differences have translated to distinct approaches to its prevention and treatment. Compared with other histologic types of lung cancer, exposures to tobacco smoke, ionizing radiation, and chloromethyl ethers pose a substantially greater risk for development of SCLC. The histologic classification of SCLC has been revised to include three categories: (1) small cell carcinoma, (2) mixed small cell/large cell, and (3) combined small cell carcinoma. Ultrastructurally, SCLC displays a number of neuroendocrine features in common with pulmonary neuroendocrine cells, including dense core vesicles or neurosecretory granules. These dense core vesicles are associated with a variety of secretory products, cell surface antigens, and enzymes. The biology of SCLC is complex. The activation of a number of dominant proto-oncogenes and the inactivation of tumor suppressor genes in SCLC have been described. Dominant proto-oncogenes that have been found to be amplified or overexpressed in SCLC include the myc family, c-myb, c-kit, c-jun, and c-src. Altered expression of two tumor suppressor genes in SCLC, p53 and the retinoblastoma gene product, has been demonstrated. Cytogenetic and molecular evidence for chromosomal loss of 3p, 5q, 9p, 11p, 13q, and 17p in SCLC has intensified the search for other tumor suppressor genes with potential import in this malignancy. Bombesin/gastrin-releasing peptide, insulin-like growth factor I, and transferrin have been identified as autocrine growth factors in SCLC, with a number of other peptides under active investigation. Several mechanisms of drug resistance in SCLC have been described, including gene amplification, the recently described overexpression of multi-drug resistance-related protein (MRP), and the expression of P-glycoprotein. The classic SCLC staging system has been supplanted by a revised TNM staging system where limited disease and extensive disease are equivalent to the TNM stages I through III and stage IV, respectively. Therapeutically, recent strategies have attained small improvements in survival but significant reductions in the toxicities of chemotherapeutic regimens. Presently, the overall 5-year survival for SCLC is 5% to 10%, with limited disease associated with a significantly higher survival rate.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 8395998 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 765: Anticancer Res. 1993 Mar-Apr;13(2):375-8. P-glycoprotein associated expression of c-fos and c-jun products in human lung carcinomas. Volm M. German Cancer Research Center, Heidelberg. Surgical specimens of non-small cell lung carcinomas of 167 previously untreated patients were analyzed for expression of c-fos, c-jun, c-myc and c-neu products and for resistance to drugs. Because most of the patients were treated only by surgery, an in vitro test was used to determine the resistance. For the detection of the oncoproteins the streptavidin-biotin-peroxidase-complex method was used. An association between the resistance and c-fos and c-jun proteins was found (c-fos p = 0.01, c-jun p = 0.09), whereas a correlation between resistance and expression of c-neu and c-myc products was not observed. P-glycoprotein 170 was detected immunohistochemically in 91 tumors using the monoclonal antibody JSB-1. There was a significant correlation between the resistance measured by the in vitro test and P-glycoprotein 170 expression (p < 0.001). Also a significant correlation between the c-fos and c-jun proteins and the expression of P-glycoprotein was found (c-fos p = 0.017, c-jun p = 0.036). In contrast, no significant relationship was found between the expression of the c-neu or c-myc products and the expression of P-glycoprotein 170. Thus, there exists a significant relationship between resistance, P-glycoprotein 170, and c-fos and c-jun products in human non-small cell lung carcinomas. P-glycoprotein 170 may be regulated by the c-fos/c-jun protein complex, which binds specifically to AP-1. PMID: 8100127 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 766: J Neurochem. 1993 Mar;60(3):877-85. Induction of primary response genes by excitatory amino acid receptor agonists in primary astroglial cultures. Condorelli DF, Dell'Albani P, Amico C, Kaczmarek L, Nicoletti F, Lukasiuk K, Stella AM. Institute of Biochemistry, Faculty of Medicine, University of Catania, Italy. We have characterized the genomic response of astroglial cells to excitatory amino acids by using selective agonists and antagonists for the various receptor subtypes and by analyzing different primary response genes, such as members of the Fos (c-fos and fosB) and Jun (c-jun, junB, and junD) families, zif/268, and c-myc. A rapid and transient elevation of mRNA levels for c-fos, fosB, c-jun, junB, and zif/268 was observed after addition of glutamate to cultured astrocytes, whereas junD and c-myc expression was not affected. The level of AP-1 DNA binding activity, as measured by the electrophoretic mobility shift assay, also increased after addition of glutamate to cultured astrocytes. Glutamate-induced c-fos expression was not affected by the N-methyl-D-aspartate receptor antagonists MK-801 and D-2-amino-5-phosphonopentanoate, by the kainate/alpha-amino-3-hydroxy-5- methylisoxazole-4-propionate (AMPA) receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX), or by the broad-spectrum antagonist kynurenate. Kainate and AMPA were also effective in inducing primary response gene expression, and their actions were antagonized by kynurenate and DNQX but not by MK-801. 1S,3R-1-Aminocyclopentane-1,3-dicarboxylic acid, a selective agonist for the metabotropic glutamate receptor, induced primary response gene expression, but its action was not antagonized by different glutamate antagonists, including L-2-amino-3-phosphonopropionate. In conclusion, our data suggest that cultured astrocytes express both kainate/AMPA ionotropic receptors and metabotropic receptors coupled to the rapid and coordinated activation of different classes of transcriptional factor genes. PMID: 8094745 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 767: Cell Mol Biol (Noisy-le-grand). 1993 Feb;39(1):35-43. A peptide containing the leucine zipper domain specifically inhibits CREB binding and transcription. Dash PK, Moore AN. Department of Neurobiology and Anatomy, University of Texas Health Science Center, Houston 77225. Formation of dimers via leucine zippers is an absolute requirement for several transcription factors to express their activity. Using circular dichroism spectroscopy, it was found that synthetic peptides which mimic the leucine zippers from CREB, Jun and Myc are alpha-helices in solution. The CREB leucine zipper peptide specifically inhibited the binding of wild type CREB protein and transcription from the CRE containing c-fos promoter in vitro. This inhibition is most likely caused by competition of the peptide to form complex with CREB monomers through leucine zippers. PMID: 8467239 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 768: Hypertension. 1993 Feb;21(2):142-8. Myotrophin induces early response genes and enhances cardiac gene expression. Mukherjee DP, McTiernan CF, Sen S. Department of Heart and Hypertension Research, Cleveland Clinic Foundation, OH 44195-5071. We have identified and partially sequenced a soluble factor, myotrophin, from spontaneously hypertensive rat hearts and hypertrophic human hearts that enhances myocyte protein synthesis and stimulates myocardial cell growth. Our studies suggest that myotrophin may be a biochemical link between hemodynamic stress and myocardial cellular hypertrophy. When rat neonatal cardiac myocytes maintained in culture were incubated with myotrophin for 30 minutes, they showed a marked increase in c-myc, c-fos, and c-jun messenger RNA levels. Cardiac myocytes treated for 24 hours with myotrophin showed a fourfold increase in connexin 43 (gap junction protein), a sixfold increase in atrial natriuretic factor, a threefold increase in skeletal alpha-actin, and a threefold increase in total myosin transcript levels. Studies on myosin isoforms showed a selective increase in the beta-myosin heavy chain transcript levels but no reciprocal decrease in alpha-myosin heavy chain transcript levels. Our data suggested that myotrophin appears to be a primary modulator for myocardial cell growth and differentiation and may play an important role in the pathogenesis of cardiac hypertrophy. Myotrophin may be involved in the upregulation of myofibrillar protein and the activation of cardiac gene transcription during growth and hypertrophy of the myocardium, and the induction of early response gene expression may be linked to this response. PMID: 8428777 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 769: Oncogene. 1993 Feb;8(2):361-70. Raf revertant cells resist transformation by non-nuclear oncogenes and are deficient in the induction of early response genes by TPA and serum. Kolch W, Heidecker G, Troppmair J, Yanagihara K, Bassin RH, Rapp UR. Laboratory of Viral Carcinogenesis, National Cancer Institute, National Institutes of Health, Frederick, Maryland 21702. A revertant cell line was generated from v-raf transformed NIH/3T3 fibroblasts. These cells, termed CHP25, express a functional v-raf oncogene. However, they are non-tumorigenic, do not form colonies in soft agar and possess a flat morphology. CHP25 cells are resistant to re-transformation by sis, ras, tyrosine kinase- as well as serine/threonine kinase-encoding oncogenes suggesting that Raf functions downstream of most membrane associated signal transducers. In contrast to v-raf transformed cells, in which the endogenous Raf-1 protein kinase is constitutively activated, v-Raf in CHP25 cells does not activate endogenous Raf-1 kinase. Since mitogen regulation of Raf-1 kinase in CHP25 cells is intact, we conclude that CHP25 cells are blocked at the level of Raf-1 substrate phosphorylation. Consistent with this interpretation CHP25 cells show specific alterations of early gene induction. The serum induction of c-fos and junD as well as the serum and TPA (12-O-tetradecanoylphorbol-13-acetate) induction of junB and egr-1 are almost completely abolished. Only v-fos can transform CHP25, whereas c-fos, v-myc, c-jun and junB are ineffective. These data suggest that the lesion responsible for the revertant phenotype of CHP25 cells is the inability to activate the AP-1 complex. We conclude that Raf-1 signaling is essential for transformation of NIH/3T3 cells by peripheral oncogenes and for regulation of a subset of early response genes by TPA and serum growth factors. PMID: 8426742 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 770: J Cell Physiol. 1993 Feb;154(2):294-300. Timing of protooncogene expression varies in toxin-induced liver regeneration. Schmiedeberg P, Biempica L, Czaja MJ. Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461. Hepatic expression of the protooncogenes c-fos and c-myc occurs within 2 h after partial hepatectomy, and these immediate early genes are thought to prime the hepatocytes for subsequent proliferation. To examine whether such gene activation occurred in the setting of hepatocyte proliferation after toxic liver injury, protooncogene expression was examined during the regenerative response following liver injury from carbon tetrachloride (CCl4) or galactosamine (GalN). The pattern of protooncogene expression after CCl4 mirrored that seen after partial hepatectomy, with rises in c-fos and c-myc mRNA content within 2 h, and then a rapid return to baseline levels. In contrast, early c-fos and c-myc expression did not occur after GalN injury. Instead GalN-induced regeneration led to a delayed, and prolonged c-fos and c-myc activation which peaked 24-48 h after injury. Increases in c-jun, jun-B, and jun-D mRNA levels also occurred in both models at times similar to the rises of c-fos and c-myc expression. Although the timing of DNA synthesis was identical after GalN or CCl4 treatment, the proliferative response after GalN injury was significantly less than that of CCl4, and marked by the histologic appearance of oval cells. The coadministration of 2-acetylaminofluorene, an inhibitor of differentiated hepatocyte proliferation, together with CCl4 altered the usual pattern of post-CCl4 protooncogene expression to one resembling that seen after GalN injury. Thus, the timing of protooncogene expression during liver regeneration may vary considerably. These variations may influence the nature of the proliferative response in terms of which cell type(s) proliferates, and the amount of regeneration that ensues. PMID: 8425910 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 771: Endocrinology. 1993 Feb;132(2):598-603. Endothelin-1 promotes steroidogenesis and stimulates protooncogene expression in transformed murine Leydig cells. Ergul A, Glassberg MK, Majercik MH, Puett D. Department of Biochemistry, University of Miami School of Medicine, Florida 33101. The effects of endothelin-1 (ET-1), a potent vasoconstrictor and mitogen to various cell types, on proliferation and differentiated functions of the murine Leydig tumor cell line MA-10 were investigated. Kinetic binding experiments at room temperature showed that [125I] ET-1 bound to MA-10 cells and reached equilibrium in 2 h. The data from competitive binding experiments yielded an apparent single class of high affinity binding sites characterized by a Kd and maximum binding capacity of 1 nM and 59 fmol/10(6) cells, respectively. For steroidogenic assays, cells were incubated with ET-1 (1 pM to 1 microM) and with epidermal growth factor (10 ng/ml) for 4 h at 37 C, and the progesterone levels in the medium were measured by RIA. Like epidermal growth factor, ET-1 caused about a 6-fold increase in progesterone production. ET-1 also enhanced the transient expression of the protooncogenes c-jun and c-myc by 3- and 2-fold, respectively. For proliferation studies, ET-1 (1 pM to 1 microM) was added to quiescent MA-10 cells for 24 h, and cell counts were performed; no increase in cell number was observed. The results of this study demonstrate that MA-10 cells possess high affinity binding sites for ET-1 and that ET-1 stimulates progesterone production and protooncogene expression, but not mitosis in this cell line. PMID: 8425480 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 772: Virology. 1993 Feb;192(2):578-86. v-erbA cooperates with bFGF in neuroretina cell transformation. Garrido C, Li RP, Samarut J, Gospodarowicz D, Saule S. Cancer Research Institute, University of California, San Francisco 94143-0128. We have studied the transforming ability of the oncogenes erbA and/or erbB in chicken neuroretina (CNR) cells and the effect of basic fibroblast growth factor (bFGF) on the transformed phenotype. The erbA oncogene alone was only transforming in the presence of bFGF. In contrast, cells expressing erbB as well as erbA + erbB were transformed in a bFGF-independent manner and were unresponsive to the growth factor. We studied whether other oncogenes could also block the cooperation between erbA and bFGF. Cytoplasmic or membrane-bound oncogenes (src, ras, or mill raf) increased the transforming potential of erbA but rendered the cells unresponsive to bFGF. Conversely, the nuclear oncogenes tested (fos and myb-ets + myc) also cooperated with erbA in CNR cell transformation but the cells remained responsive to the growth factor. A likely explanation is that CNR cells carrying the cytoplasmic but not the nuclear oncogenes have already activated the bFGF signal transduction pathway. PMID: 7678474 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 773: Eur J Biochem. 1993 Jan 15;211(1-2):7-18. Erratum in: Eur J Biochem 1993 Aug 1;215(3):907. The Ets family of transcription factors. Wasylyk B, Hahn SL, Giovane A. CNRS-LGME/INSERM-U. 184, Institut de Chimie Biologique, Faculte de Medecine, Strasbourg, France. Interest in the Ets proteins has grown enormously over the last decade. The v-ets oncogene was originally discovered as part of a fusion protein expressed by a transforming retrovirus (avian E26), and later shown to be transduced from a cellular gene. About 30 related proteins have now been found in species ranging from flies to humans, that resemble the vEts protein in the so-called 'ets domain'. The ets domain has been shown to be a DNA-binding domain, that specifically interacts with sequences containing the common core trinucleotide GGA. Furthermore, it is involved in protein-protein interactions with co-factors that help determine its biological activity. Many of the Ets-related proteins have been shown to be transcription activators, like other nuclear oncoproteins and anti-oncoproteins (Jun, Fos, Myb, Myc, Rel, p53, etc.). However, Ets-like proteins may have other functions, such as in DNA replication and a general role in transcription activation. Ets proteins have been implicated in regulation of gene expression during a variety of biological processes, including growth control, transformation, T-cell activation, and developmental programs in many organisms. Signals regulating cell growth are transmitted from outside the cell to the nucleus by growth factors and their receptors. G-proteins, kinases and transcription factors. We will discuss how several Ets-related proteins fit into this scheme, and how their activity is regulated both post- and pre-translationally. Loss of normal control is often associated with conversion to an oncoprotein. vEts has been shown to have different properties from its progenitor, which might explain how it has become oncogenic. Oncogene-related products have been implicated in the control of various developmental processes. Evidence is accumulating for a role for Ets family members in Drosophila development, Xenopus oocyte maturation, lymphocyte differentiation, and viral infectious cycles. An ultimate hope in studying transformation by oncoproteins is to understand how cells become cancerous in humans, which would lead to more effective treatments. vEts induces erythroblastosis in chicken. Cellular Ets-family proteins can be activated by proviral insertion in mice and, most interestingly, by chromosome translocation in humans. We are at the beginning of understanding the multiple facets of regulation of Ets activity. Future work on the Ets family promises to provide important insights into both normal control of growth and differentiation, and deregulation in illness. Publication Types: Review PMID: 8425553 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 774: Cancer. 1993 Jan 15;71(2):419-29. Gene structure and expression analysis of the epidermal growth factor receptor, transforming growth factor-alpha, myc, jun, and metallothionein in human ovarian carcinomas. Classification of malignant phenotypes. Bauknecht T, Angel P, Kohler M, Kommoss F, Birmelin G, Pfleiderer A, Wagner E. Universitats-Frauenklinik, Freiburg, Germany. This study reports the structure and expression rates of genes of the transforming growth factor-alpha (TGF-alpha) signal transduction pathway (TGF-alpha, epidermal growth factor receptor [EGF-R], jun, myc, and metallothionein [MT]) in 47 specimens of ovarian cancer and 21 nonmalignant tissues. The objective was to establish a direct correlation between the genetic activities and the malignant phenotype of the ovarian cancer. The Southern blot technique identified four samples with myc amplification and two with rearranged EGF-R genes. By using the S1 nuclease assay, the analysis of myc transcription showed a similar use of both promotors. Although the size of the investigated transcripts was unaltered, significant differences in the transcription rates were noticed in malignant tissue probes (using northern blot analysis and RNAase protection assay). The following results of messenger RNA analysis in ovarian cancer were observed: EGF-R, negative in 25%, low in 65%, and strongly positive in 10%; TGF-alpha, negative in 34%, low in 36%, and strongly positive in 30%; myc, negative in 8%, low in 64%, and strongly positive in 28%; jun, negative in 4%, low in 58%, and strong in 38%; and MT, low in 80% and strongly positive in 20%. In most nonmalignant tissues studied, no or only a low expression of TGF-alpha, EGF-R, and myc. was found. A comparison of these messenger RNA results with the clinical data from tumors showed four different subgroups of ovarian carcinomas. The results of chemotherapy were known in 32 cases. Tumors with negative or low expression rates of all investigated genes did not respond to chemotherapy; 13 of 18 tumors with high expression rates did respond. Additional signal transduction chains distinct from the TGF-alpha pathway, however, are likely to influence both the expression and activity of transcription factors and MT. PMID: 8422634 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 775: Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):477-81. Induction of protein-tyrosine-phosphatase activity by interleukin 6 in M1 myeloblastic cells and analysis of possible counteractions by the BCR-ABL oncogene. Zafriri D, Argaman M, Canaani E, Kimchi A. Department of Molecular Genetics and Virology, Weizmann Institute of Science, Rehovot, Israel. Interleukin 6 (IL-6) induces in M1 myeloblastic cells growth arrest and terminal differentiation toward monocytes. It is reported here that IL-6 reduced by 5- to 20-fold the tyrosine phosphorylation of cellular proteins in these cells. The same-fold reduction was also observed in M1 cells that were transfected with the BCR-ABL deregulated protein kinase. In these stable clones, the levels of tyrosine phosphorylation of cellular proteins were 30- to 100-fold higher than in the parental cells. IL-6 did not reduce the expression levels or the inherent tyrosine kinase activity of BCR-ABL p210. By measuring the protein-tyrosine-phosphatase (PTPase; protein-tyrosine-phosphate phosphohydrolase, EC 3.1.3.48) activity in crude cell lysates, we found that protein dephosphorylation resulted, at least partially, from induction of PTPase activity by IL-6. The induction of PTPase in the BCR-ABL-transfected clones was not sufficient to confer the minimal protein phosphorylation levels characteristic of IL-6-treated cells. Yet, the transfected M1 clones showed normal growth and differentiation responses to IL-6. None of the gene responses to IL-6 including suppression in the levels of c-myc, c-myb, and cyclin A mRNA; junB and c-jun mRNA induction; and dephosphorylation of retinoblastoma protein were rescued by the BCR-ABL oncogene. The functional relevance of PTPase induction by IL-6 is discussed. PMID: 8421678 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 776: Cancer Res. 1993 Jan 15;53(2):291-7. Induction of jun gene family members by transforming growth factor alpha but not 17 beta-estradiol in human breast cancer cells. Davidson NE, Prestigiacomo LJ, Hahm HA. Johns Hopkins Oncology Center, Baltimore, Maryland 21231. To investigate whether estrogen treatment of hormone-responsive human breast cancer cells was associated with activation of members of the jun family of immediate early response genes, the expression of these oncogenes in human breast cancer cells was examined. 17 beta-Estradiol had little effect on expression of c-jun, jun B, jun D, or c-fos mRNA by MCF-7 cells over 12 h, although it stimulated c-myc expression 4-fold within 30 min. In contrast, several peptide growth factors, including transforming growth factor-alpha (TGF-alpha), rapidly and transiently induced expression of c-jun, jun B, and c-fos mRNA 4- to 10-fold over control. A similar pattern of expression was seen in two other estrogen-responsive human breast cancer cell lines, ZR-75-1 and T47D. Inhibition of protein synthesis by cycloheximide did not abrogate induction of c-jun or jun B mRNA by TGF-alpha in MCF-7 cells, suggesting that new protein synthesis was not required. In addition, nuclear runoff transcription analysis demonstrated that increased expression of c-jun and jun B mRNA after TGF-alpha treatment of MCF-7 cells was regulated at least in part at the transcriptional level. Chronic exposure of MCF-7 cells to 17 beta-estradiol for 24-48 h was associated with decreased expression of jun B mRNA only, while similar treatment with TGF-alpha did not change mRNA expression of any jun family member. Thus, expression of jun family members is induced by peptide growth factors like TGF-alpha but not 17 beta-estradiol in human breast cancer cells. These results suggest that these nuclear protooncogenes play different roles in modulating gene expression by MCF-7 cells after exposure to TGF-alpha or 17 beta-estradiol. PMID: 8417822 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 777: J Biol Chem. 1993 Jan 15;268(2):968-73. Involvement of immediate-early gene expression in the synergistic effects of steel factor in combination with granulocyte-macrophage colony-stimulating factor or interleukin-3 on proliferation of a human factor-dependent cell line. Horie M, Broxmeyer HE. Department of Medicine (Hematology/Oncology), Indiana University School of Medicine, Indianapolis 46202-5121. Steel factor (SLF) synergizes with granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-3 (IL-3) to stimulate proliferation of human factor-dependent cell line, MO7e. To elucidate molecular mechanisms underlying this synergism, induction of immediate-early genes was studied. Treatment of MO7e cells with SLF, GM-CSF, and IL-3 induced/enhanced expression of c-fos, junB, egr-1, and c-myc genes. SLF treatment of MO7e cells led to higher expression of c-fos, junB, and egr-1 genes than did treatment with GM-CSF or IL-3. However, GM-CSF and IL-3 had more prolonged effects on enhancement of the c-myc gene than SLF. Using optimal dosages for cell proliferation, induction of c-fos and junB was greater than additive with SLF plus GM-CSF or IL-3, as compared with each factor alone. Using suboptimal amounts of SLF with optimal GM-CSF or IL-3, induction of c-fos, junB, egr-1, and c-myc genes was greater than additive. De novo protein synthesis was not required for greater induction of these immediate-early genes by the combination of SLF plus GM-CSF. Based on nuclear run-on and actinomycin D experiments, the data suggest that the synergistic effects of SLF plus GM-CSF on the induction of immediate-early genes may be mediated in part at the level of transcription and mRNA stabilization for c-fos, at the level of mRNA stabilization for junB, and at the level of transcription for egr-1. PMID: 7678261 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 778: Annu Rev Immunol. 1993;11:245-68. The IL-2 receptor complex: its structure, function, and target genes. Minami Y, Kono T, Miyazaki T, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. Proliferation of T lymphocytes is triggered by the interaction of IL-2 with its specific receptor following T lymphocyte activation. The receptor for IL-2 consists of at least three distinct subunits, the alpha chain (IL-2R alpha), the beta chain (IL-2R beta), and the gamma chain (IL-2R gamma). Although the role of IL-2R gamma in IL-2 signalling remains unclear, IL-2R beta is the subunit critical for receptor-mediated signalling. Because IL-2R beta lacks any apparent catalytic motifs, IL-2R beta may be physically or functionally coupled to other signalling molecules. Structure-function studies of IL-2R beta have revealed that at least two distinct cytoplasmic regions of IL-2R beta are involved in IL-2-induced cellular signalling. The "serine-rich" region of IL-2R beta was identified as a region critical for IL-2-induced mitotic signalling from experiments in which IL-2R beta mutant cDNAs lacking a particular cytoplasmic region or regions were expressed in an IL-3-dependent mouse pro-B cell line (BAF-B03). Meanwhile, another cytoplasmic region of IL-2R beta, the "acidic" region, is responsible for its physical association with an src-family protein tyrosine kinase (PTK), p56lck and is critical for activating the p56lck PTK following IL-2 stimulation. It is now evident that IL-2R beta is linked to at least two intracellular signalling pathways that mediate nuclear proto-oncogene induction. One pathway is linked to tyrosine phosphorylation events, mediated by a src-family protein tyrosine kinase (PTK), and that pathway leads to the induction of the c-fos, c-jun, and other genes of this family. Another pathway leads to c-myc gene induction by an as yet unknown mechanism. We discuss the complex signalling machinery that links the cell surface receptor to the nuclear events. Publication Types: Review PMID: 8476561 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 779: Differentiation. 1993 Jan;52(2):161-8. Endogenous activation of c-myc expression and DNA synthesis in serum-starved neonatal rat smooth muscle cells. Hultgardh-Nilsson A, Krondahl U, Jiang WQ, Nilsson J, Ringertz NR. Department of Medical Cell Genetics, Medical Nobel Institute, Karolinska Institutet, Stockholm, Sweden. Earlier studies have shown that smooth muscle cells (SMC) from arteries of neonatal and adult rats differ markedly in their in vitro growth characteristics. Since some of these differences may be relevant to the proliferation of SMC in atherosclerotic plaques we examined the expression of three proto-oncogenes (c-fos, c-jun, and c-myc) and an SMC-specific differentiation marker (alpha-actin) in cultured SMC. In presence of serum cultured adult SMC contained lower levels of alpha-actin mRNA than neonatal cells. In neonatal cells serum-starvation resulted in a distinct increase in both c-myc and alpha-actin mRNA levels, whereas the expression of these genes appeared to be unaffected in adult cells. Transfer of adult SMC proliferating in the presence of fetal calf serum to serum-free medium for 48 h almost completely inhibited DNA synthesis, whereas transfer of neonatal SMC to serum-free medium reduced DNA synthesis only to about 50%. Serum-starved adult and neonatal SMC did not contain c-fos or c-jun transcripts, but in both cell types serum-stimulation resulted in a comparable increase in the expression of both genes. The present results demonstrate clear differences in the mechanisms regulating gene expression in adult and neonatal SMC. PMID: 8472886 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 780: Ren Fail. 1993;15(2):163-71. Induction of immediate early and stress genes in rat proximal tubule epithelium following injury: the significance of cytosolic ionized calcium. Yamamoto N, Maki A, Swann JD, Berezesky IK, Trump BF. Department of Pathology, University of Maryland, School of Medicine, Baltimore. This study was designed to investigate the influence of intracellular ionized calcium ([Ca2+]i) on the induction of c-fos, c-jun, c-myc, and hsp70 genes after oxidant stress induced by xanthine/xanthine oxidase (X/XOD) treatment or after heat shock using primary cultures of rat proximal tubule epithelium (PTE). X/XOD (500 microM/25 mU/mL) induced all of these genes; ionomycin also resulted in similar kinetics of induction of all genes. The expression of both c-fos following X/XOD treatment and hsp70 following heat shock was markedly decreased through chelation of [Ca2+]i by Quin 2/AM. The c-fos expression following X/XOD treatment was partly reduced by a protein kinase C inhibitor, staurosporine (ST), and markedly inhibited by another protein kinase inhibitor, 2-aminopurine (2AP), while both ST and 2AP markedly reduced hsp70 expression. The ADP-ribosylation transferase inhibitor 3-aminobenzamide had no effect on either c-fos or hsp70 expression. These results suggest that cell injuries leading to increased [Ca2+]i in PTE result in induction of c-fos, c-jun, c-myc, and hsp70; and that the activation of c-fos and hsp70 genes may be regulated by [Ca2+]i and [Ca2+]i-dependent protein kinases. PMID: 8469783 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 781: Cancer Invest. 1993;11(2):198-211. Erratum in: Cancer Invest 1998;16(3):212. Cancer Invest 1998;16(6):429. Modulation of cytotoxicity and differentiation-inducing potential of arabinofuranosylcytosine in myeloid leukemia cells by hematopoietic cytokines. Brach MA, Mertelsmann RH, Herrmann F. Department of Internal Medicine 1, University of Freiburg Medical Center, Federal Republic of Germany. Hematopoietic growth factors may be useful in improving the clinical effectiveness of arabinofuranosylcytosine (ara-C). In vitro studies have indicated that interleukin 3(IL-3) and, to a lesser extent, granulocyte-macrophage colony-stimulating factor (GM-CSF), but not G-CSF or M-CSF, may be capable of specifically augmenting the ability of ara-C to kill leukemic myeloid cells by pharmacological and cytokinetic mechanisms including increase of intracellular ara-CTP/dCTP pool ratios and enhanced ara-C DNA incorporation in leukemic blast cells, decrease of IC 90 of ara-C for leukemic colony-forming cells (CFC) as compared with normal CFC growth, and recruitment of quiescent leukemic cells into the cell cycle. In contrast, the combination of ara-C with M-CSF or with the leukemia inhibitory factor (LIF) appears to be useful in overcoming the block in differentiation of leukemic blast, while the effects of GM-CSF and IL-3 on ara-C-induced differentiation appear limited. The combined treatment of human myeloid leukemia cells by ara-C and LIF is associated with down-regulation of c-myc gene expression, transcriptional activation of jun/fos gene expression, and features of functional differentiation (e.g., the capability to reduce nitroblue tetrazolium, to express lysozyme, or to display differentiation-related surface receptors including C3bi and the c-fms protein). On the basis of these in vitro studies first clinical trials are underway that are examining the efficacy of ara-C combinations with these molecules for the treatment of myeloid disorders. Publication Types: Review Review, Tutorial PMID: 8462021 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 782: Am J Pathol. 1993 Jan;142(1):95-105. Coordinate synthesis of stromelysin, interleukin-1, and oncogene proteins in experimental osteoarthritis. An immunohistochemical study. Pelletier JP, Faure MP, DiBattista JA, Wilhelm S, Visco D, Martel-Pelletier J. University of Montreal, Rheumatic Disease Unit, Quebec, Canada. Metalloproteases appear to play an important role in the pathophysiology of osteoarthritis (OA) and their expression is believed to be regulated by cytokines such as interleukin-1 (IL-1). Nuclear oncogene products are suggested as mediators through which IL-1 induces metalloprotease gene expression. Little data are available on the in vivo involvement of these agents in the pathophysiology of OA. This study examined by immunohistochemistry, using specific antibodies, the distribution of stromelysin, IL-1 alpha, IL-1 beta, and oncogene products (c-FOS, c-JUN, and c-MYC) in synovium and cartilage from normal and experimental canine models of OA. In the OA synovium, stromelysin and IL-1 were localized in the cytoplasm of superficial synovial lining cells, infiltrating mononuclear cells, and endothelial and smooth muscle cells of the blood vessels, whereas oncoproteins were detected predominantly in the synovial lining cells. Normal synovial membranes demonstrated low levels of specific staining in synovial lining cells with occasional staining of blood vessel cells for IL-1 alpha, IL-1 beta, and stromelysin. In OA cartilage, chondrocytes at the superficial and middle layers as well as in fibrillated areas were found to be involved in the synthesis of stromelysin, IL-1, and oncoproteins. Diffuse staining of stromelysin and IL-1 beta in OA cartilage matrix was also identified. In normal cartilage, only a few chondrocytes at the superficial layer showed a low level of antigens. These results demonstrate the in vivo concomitant cellular and/or matrical presence of stromelysin, IL-1, and oncogene proteins in tissues from experimentally induced OA with the most intense staining at the sites of cartilage erosion and synovial proliferation. These findings suggest that they may be involved in the pathophysiology of OA, and that the regulatory mechanisms involved in the expression of these proteins may be associated. PMID: 8424468 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 783: J Cell Physiol. 1993 Jan;154(1):143-51. Pulsatile and steady flow induces c-fos expression in human endothelial cells. Hsieh HJ, Li NQ, Frangos JA. Department of Chemical Engineering, Pennsylvania State University, University Park 16802. The effects of pulsatile and steady fluid flow on the mRNA levels of proto-oncogenes c-fos, c-jun, and c-myc in cultured human umbilical vein endothelial cells (HUVEC) were investigated. c-fos mRNA levels in stationary cultures were very low. A 1 Hz pulsatile flow with an average shear stress of 16 dynes/cm2 induced a dramatic increase of c-fos mRNA levels in HUVEC 0.5 h after the onset of flow, which declined rapidly to basal levels within 1 h. Steady flow with a similar shear stress also induced a transient increase of c-fos mRNA levels, but to a lesser extent. In addition, increased c-fos mRNA levels were observed when low shear (2-6 dynes/cm2) was replaced by high shear (16-33 dynes/cm2). Pulsatile and steady flow caused a slight increase of c-jun and c-myc mRNA levels. The role of pulsatility was also investigated in platelet-derived growth factor (PDGF) expression. Pulsatile flow induced a transient increase of PDGF A- and B-chain mRNA levels with peaks at 1.5-2 h. Pulsatile flow, which was more stimulatory in mediating c-fos expression, however, was less stimulatory than steady flow in mediating PDGF expression. By using various inhibitors, protein kinase C was found to be an important mediator in flow-induced c-fos expression, with the involvement of G proteins, phospholipase C, and intracellular calcium. Protein kinase C was previously shown as a possible major mediator in flow-induced PDGF expression which, at least partly, appeared to follow the induction mechanism of c-fos, suggesting a possible connection between c-fos and PDGF induction. However, the c-fos antisense treatment, which significantly inhibited c-fos transcription, failed to block the flow-induced PDGF expression, suggesting that flow-induced c-fos expression may not play an important role in the mechanism of flow-induced PDGF expression. The difference in the induction of c-fos and PDGF expression under pulsatile as compared to steady flow indicates that a complex, flow-mediated regulatory mechanism of gene expression exists in HUVEC. The increased expression of these proto-oncogenes mediated by flow may be important in regulating long-term cellular responses. PMID: 8419400 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 784: Crit Rev Oncog. 1993;4(4):361-88. Estrogen regulation of proto-oncogenes coding for nuclear proteins. Weisz A, Bresciani F. Istituto di Patologia Generale e Oncologia, Facolta di Medicina e Chirurgia, Seconda Universita degli Studi di Napoli, Italy. Estrogen hormones are known to exert a complex influence on development and function of the female reproductive organs of vertebrates by regulating cell growth and differentiation, as well as to be implicated in oncogenesis and maintenance of tumor growth. Estrogen acts on cells via interaction with an intracellular receptor, which, like all receptors for steroid hormones, is a trans-acting transcription enhancer factor activated by the cognate ligand and capable of binding to specific, cis-acting enhancer elements usually located within the 5'-flanking regions of target genes. Additionally, estrogen regulates gene expression by influencing mRNA stability or via interaction of the estrogen receptor with transcription regulatory factors. This article reviews data indicating that estrogen directly activates (primary activation) expression of proto-oncogenes codifying for nuclear proteins that, in turn, are responsible for indirect (secondary) activation of other genes. This cascade mechanism of gene activation is likely to progress for several more steps and allows us to envisage how estrogen can direct a complex task such as cell reproduction. Among proto-oncogenes codifying for nuclear proteins, we focus on fos, jun, myc, and related genes. The mechanisms of regulation of these genes by estrogen, including regulation of transcription, messenger RNA stabilization, and protein-protein interaction, are reviewed. Publication Types: Review PMID: 8353138 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 785: NIDA Res Monogr. 1993;125:3-24. Regulation of immediate early gene expression. Cochran BH. Center for Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge 02139. From this brief overview of the regulation of the c-fos promoter, it can be seen that the regulation of early-response genes is a complex affair. Therefore, it is not easy to predict from the upstream sequence of a given early-response gene exactly which elements are responsible for responding to what signals in a given cell type. However, from studies of other early-response genes, it is clear that several of the elements found upstream of c-fos appear frequently and are important in the regulation of other early-response genes. For instance, the zif/268 gene has four separate CArG boxes that are similar to fos. These CArG boxes can function in the serum and TPA response, but interestingly, they are not imbedded in a region of dyad symmetry as in c-fos (Christy and Nathans 1989). Upstream of the c-jun oncogene is an AP-1 site that can modulate the expression and induction of this gene and is responsive to TPA (Angel et al. 1988). Moreover, other studies suggest that cAMP-mediated signals can repress induction of c-jun through this element (de Groot et al. 1991; Mechta et al. 1989). This would explain why in certain circumstances, such as depolarization of PC12 cells or in the striatum in response to cocaine, there is an uncoupling of the induction of c-jun and jun-B. Depolarization induces c-fos and jun-B but not c-jun; however, growth factors such as NGF can induce all three genes in the same cell (Bartel et al. 1989). Upstream of the jun-B gene there does not appear to be an SRE, but there is a new element that can be responsive to both cAMP and phorbol esters (de Groot et al. 1991). Genes such as c-myc, JE, and KC have no consensus SREs upstream, and the regulatory elements responsible for the induction of these genes have not been clearly identified (Rollins et al. 1988). However, there is some evidence from the c-myc gene that the E2F binding sites are important for its regulation by serum (Mudryj et al. 1990; Sacca and Cochran 1990). In addition, there are two SIF sites upstream of the c-myc proto-oncogene (B.H. Cochran and T.E. Hayes, unpublished results). Upstream of the nur-77 gene there are no SREs, but there are four potential calcium/CRE-like elements (Watson and Milbrandt 1989).(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 8341367 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 786: Crit Rev Eukaryot Gene Expr. 1993;3(2):117-35. Erratum in: Crit Rev Eukaryot Gene Expr 1993;3(4):316-7. Control of liver growth. Fausto N, Webber EM. Department of Pathology and Laboratory Medicine, Brown University, Providence, Rhode Island 02912. The liver is an excellent tissue for the study of growth regulation because of its ability to regenerate by a process of compensatory growth following surgical resection or toxic injury. Much of the investigation on the mechanisms of hepatic growth has been done in partially hepatectomized animals in vivo and in hepatocytes in primary culture. Almost immediately after partial hepatectomy there are major changes in the binding capacity of transcription activators and in the expression of a relatively large number of genes. Many of the immediate early response gene products are themselves transcription activators and thus can multiply and propagate the initial gene activation process. An important issue in the regulation of liver growth is to identify growth factors that may play a role in hepatocyte replication in vivo. In addition to substances that are adjuvants in the mitogenic response at least three growth factors, EGF, TGF alpha, and HGF, are complete mitogens for hepatocytes in culture and appear to play important roles as stimulators of liver growth. We discuss data that indicate that none of these growth factors seems capable of causing a significant increase in DNA synthesis in quiescent hepatocytes in vivo. In contrast, EGF, TGF alpha, and HGF (both the monomer and the heterodimer) increase DNA synthesis in vivo in hepatocytes that have become "competent" to proliferate. We suggest that the competence process involves the activation of transcription factors and protooncogenes, and that during liver regeneration at least some of these changes precede rather than follow growth factor/ligand signaling. The available data suggest that the three growth factors by themselves may not trigger regeneration in strictly quiescent hepatocytes. There is now extensive evidence from work on TGF alpha in developing and regenerating liver as well as studies with transgenic animals to indicate that the factor may act as a cell cycle progression agent. We suggest that "priming" of hepatocytes after partial hepatectomy might have similarities to the widespread activation of genes observed in response to stimuli such as heat shock, ionic imbalances, and changes in redox potentials, and that TGF alpha and also probably the other growth factors act to make primed cells progress through the cycle and undergo DNA synthesis. TGF beta 1 is a potent antagonist to the mitogenic effects of these growth factors on cultured hepatocytes and could be an important factor for terminating the proliferative response of hepatocytes after partial hepatectomy. Although we have moved closer to finding "on" and "off" switches for regeneration, their precise identities and mechanisms remain elusive. Publication Types: Review PMID: 8324292 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 787: Nucleic Acids Symp Ser. 1993;(29):109-10. Molecular mechanism of cell death induced dNTP pool imbalance. Wataya Y, Hwang H, Nakazawa T, Takahashi K, Otani M, Igaki T. Faculty of Pharmaceutical Sciences, Okayama University, Japan. We have previously reported that treatment of the FM3A cells with 5-fluorodeoxyuridine (FdUrd) induced DNA double strand breaks and cell death. We proposed the hypothesis of dNTP pool imbalance death in order to understand these phenomena: intracellular dNTP pool imbalance induced by FdUrd would be a trigger for activation or induction of endonuclease which would attack DNA to cause double strand breaks and subsequent cell death. To observe the mechanism of dNTP imbalance death we have purified and characterized the endonuclease that was detected in FdUrd treated FM3A cells but not untreated cells. As the result, we suggested that purified enzyme is a new endonuclease and responsible for DNA degradation component of the apoptotic process. Furthermore, we study that mRNA levels of nuclear protooncogenes, c-fos, c-jun and c-myc, in FdUrd-treated cells. These results suggested that the endonuclease might be induced by the protooncogene related pathway and generates DNA fragments accompanied by cell death. PMID: 8280290 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 788: Postepy Hig Med Dosw. 1993;47(6):475-85. [Renin-angiotensin system and atherosclerosis] [Article in Polish] Stajszczyk M, Gminski J. Katedra i Zaklad Biochemii i Chemii Slaskiej AM, SKatowiaicach. Based upon literature the renin-angiotensin system involvement in the pathogenesis of atherosclerosis has been discussed. Angiotensin II leads to the increased production of growth factors such as PDGF, TGF-beta, FGF and extracellular matrix proteins. There are evidences that angiotensin II stimulates expression of egr-1, c-jun, c-fos and c-myc oncogenes in vascular smooth muscle cells. Proliferation of aortic smooth muscle cells in response to the injury can be reduced by inhibitors of renin-angiotensin system what supports the hypothesis that angiotensin II can contribute to the pathogenesis of atherosclerosis. Publication Types: Review PMID: 8208630 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 789: J Cancer Res Clin Oncol. 1993;119(9):507-10. Prognostic significance of the expression of c-fos, c-jun and c-erbB-1 oncogene products in human squamous cell lung carcinomas. Volm M, Drings P, Wodrich W. German Cancer Research Center, Heidelberg. The expression of the oncogenes c-fos, c-jun, c-myc, c-erbB-1 and c-erbB-2 at the protein level was analyzed in squamous cell lung carcinomas of 121 patients by means of immunohistochemistry. Patients with overexpression of proteins encoded by the oncogenes c-fos, c-jun and c-erbB-1 had significantly shorter survival times than these without overexpression of these oncogene products (c-fos: p = 0.009; c-jun: p = 0.029; c-erbB-1: p = 0.018). No significant correlations were found between the expression of c-myc and c-erbB-2 products and the survival of the patients. In addition to the univariate analyses (Kaplan-Meier-estimates) multivariate analyses (Cox-regression-model) revealed that protein expression of the oncogenes c-fos, c-jun and c-erbB-1 are significant prognostic factors in addition to staging. PMID: 8100821 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 790: Oncol Res. 1993;5(10-11):397-408. DNA damage, gene expression, growth arrest and cell death. Gewirtz DA. Department of Medicine, Medical College of Virginia, Richmond 23298. The sequence of biochemical and molecular events that mediate growth arrest and cell death in tumor cells exposed to agents that induce DNA damage is poorly defined. This commentary exploits the recent explosion of information regarding oncogenes, tumor suppressor genes, and cell-cycle regulatory genes to develop a model for growth arrest/cell death. The model focuses on changes in the expression of these genes, in the level and phosphorylation of their protein products, and in the interaction(s) between these proteins. It is recognized that such a model is, of necessity, incomplete, since new gene functions associated with the cellular response to DNA damage will continuously be uncovered; in addition, the proposed sequence of events will likely require modification as the relationships between the functions of the discrete gene products are clarified. Nevertheless, it is hoped that this commentary will provide a conceptual framework within which to fit currently available information as well as future findings relating to the expression and function of DNA-damage-responsive genes, and that the sections of the model that are incomplete will provide a springboard for the development of research approaches designed to answer specific questions regarding the nature of the cellular response to DNA damage. Publication Types: Review PMID: 8054700 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 791: Cytotechnology. 1993;11 Suppl 1:S18-20. The in utero initiation with DMN alters the complement of cytosolic glutathione S-transferases and the phenobarbital-induced expression of c-jun and c-myc oncogenes in primary neonatal rat hepatocytes. Armato U, Wu J, Menegazzi M, Menapace L, Ribecco M, Testolin L, Carcereri De Prati A, Suzuki H. Institute of Anatomy, School of Medicine, University of Verona, Italy. As revealed by a novel in utero-in vitro hepatocarcinogenesis model, a single exposure to dimethylnitrosamine (DMN) elicited in postnatal (and fetal) primary rat hepatocytes (i) immunocytochemically detectable, widespread increases in their complement of the alpha, mu and especially pi classes of cytosolic glutathione S-transferases (GSTs); and (ii) changed patterns (with respect to controls) of the phenobarbital (PB)-evoked increases in steady state levels of c-jun and c-myc mRNA's. These results indicate that DMN causes both transient cytotoxic effects and a broad, permanent initiation in fetal proliferating hepatocytes. PMID: 7763748 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 792: Tumour Biol. 1993;14(4):201-12. Nuclear oncogenes and alpha-fetoprotein gene expression in hepatoma cell lines. Tournier-Thurneyssen I, Feldmann G, Bernuau D. INSERM U-327, Laboratoire de Biologie Cellulaire, Faculte de Medecine Xavier-Bichat, Paris, France. We have previously demonstrated a correlation between the kinetics of activation of the nuclear oncogenes c-fos, c-jun and c-myc and the alpha-fetoprotein (AFP) gene in adult rat hepatocytes proliferating in culture, which led us to raise the hypothesis of a possible regulation of the AFP gene by nuclear oncogenes. Whether the mechanisms of AFP gene expression in normal and transformed hepatocytes are similar is unknown. In this study we have searched for a possible correlation between the basal level of AFP gene expression, by several hepatoma cell lines that express the AFP gene differently, and the basal level of expression of the nuclear oncogenes c-fos, c-jun and c-myc. The analysis has been performed at the mRNA and protein levels. Our results demonstrate that cell lines that strongly express the AFP gene do not express higher levels of nuclear oncogenes than cell lines with weak expression of the AFP gene. These results, therefore, do not support a direct involvement of nuclear oncogenes on the basal level of AFP gene expression in hepatoma cell lines. PMID: 7692587 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 793: Leuk Lymphoma. 1993;10 Suppl:123-31. Effect of hemopoietic growth factors G-CSF and pIXY 321 on the activity of high dose Ara-C in human myeloid leukemia cells. Bhalla K, Tourkina E, Huang Y, Tang C, Mahoney ME, Ibrado AM. Department of Medicine, Medical University of South Carolina, Charleston 29425-2225. Recently, high dose Ara-C (HIDAC) has been shown to induce leukemic cell death in vitro by the alternative process of programmed cell death (PCD) or apoptosis which correlates with the inhibition of their clonogenic survival. Since co-treatment with hemopoietic growth facts (HGFs) GM-CSF and IL-3 have been demonstrated to enhance the metabolism and cytotoxic effects of HIDAC against leukemic progenitor cells, we examined the effect of HGFs pIXY 321 (a GM-CSF/IL3 fusion protein) and G-CSF on HIDAC induced PCD and related gene expressions as well as HIDAC mediated colony growth inhibition of human myeloid leukemia cells. Treatment with G-CSF or pIXY 321 alone for up to 24 hours neither suppressed nor induced PCD in HL-60 or KG-1 cells. However, exposure to either of the HGFs for 20 hours followed by a combined treatment for 4 hours with HIDAC plus either of the HGFs versus HIDAC alone significantly enhanced the intracellular Ara-CTP accumulation and the oligonucleosomal DNA fragmentation characteristic of PCD. This was temporally associated with a marked induction of C-jun expression but a significant repression in BCL-2 and c-myc expressions. In addition, the treatment with either of the HGFs plus HIDAC versus HIDAC alone produced a significantly greater inhibition of the clonogenic survival of the myeloid leukemia cells. These findings underscore an additional mechanism of leukemic cell death induced by HIDAC which can be modulated by the HGFs to improve the antileukemic activity of HIDAC. PMID: 7683227 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 794: J Cell Biochem. 1992 Dec;50(4):411-24. Glucocorticoid regulation of alkaline phosphatase, osteocalcin, and proto-oncogenes in normal human osteoblast-like cells. Subramaniam M, Colvard D, Keeting PE, Rasmussen K, Riggs BL, Spelsberg TC. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905. In humans, glucocorticoids are known to have marked effects on bone metabolism and function, including the significant regulation of osteoblast cells. To aid in the understanding of the mechanism of glucocorticoid action on normal human osteoblasts (hOB), confluent cells were analyzed for the presence of glucocorticoid receptors (GR) as well as for the effects of the glucocorticoid dexamethasone (Dex) on the expression of both the rapid responding nuclear proto-oncogenes and the late responding structural genes for bone matrix proteins. The interactions between Dex and 1,25 dihydroxy vitamin D3 (1,25 D3) on the gene expression in these cells were also examined. Using a functional receptor assay, a mean of 11,600 functional nuclear bound glucocorticoid receptors (range 6,000-22,000) was measured in fifteen separate cell strains. Northern blot analysis with a cDNA probe to the human GR was used to demonstrate the presence of a 7Kb transcript which is a candidate mRNA for GR in these cells. In agreement with previous studies, treatment of the hOB cells with Dex increased the steady state mRNA levels for alkaline phosphatase (AP) but displayed little or no effect on the mRNA levels for osteocalcin (OC) and glyceraldehyde phosphate dehydrogenase (GAPDH). Interestingly, the 1,25 D3 inductions of mRNA levels for OC were blocked by Dex but enhanced for AP. The above effects of Dex on AP and OC gene expression, including the interaction with 1,25 D3, were also shown to occur at the level of protein. The effect of Dex on the mRNA levels of the nuclear proto-oncogenes c-myc, c-fos, and c-jun was also investigated, since the oncoproteins (Fos/Jun) appear to play a role in the delayed glucocorticoid regulation of structural genes. Interestingly, Dex increased the steady state levels of c-myc, c-fos, and c-jun mRNAs in nonproliferating (confluent) hOB cells by 3.5-, 10-, and 2.0-fold, respectively, over control (untreated cells) values within one h of steroid treatment. The Dex-induced mRNA levels were transient and returned to basal values within 24 h of the steroid treatment. A reduced but qualitatively similar pattern of response was found in proliferating hOB cells. The pattern of response of these genes to glucocorticoids in hOB cells mimics the response in avian liver cells but not in reproductive cells. These results support the theory that hOB cells are target cells for glucocorticoids, and that as a primary event glucocorticoids rapidly regulate the expression of the nuclear oncoproteins Fos/Jun in these cells. PMID: 1469072 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 795: Oncogene. 1992 Dec;7(12):2455-60. c-myc and p53 gene expression in the differentiation of temperature-sensitive mutants of teratocarcinoma F9 cells. Kosaka M, Iwai SA, Nishina Y, Sumi T, Nishimune Y. Research Institute for Microbial Diseases, Dental Faculty, Osaka University, Japan. We have previously reported the isolation of several temperature-sensitive (ts) mutants of F9 cells. Further investigations showed that some mutants were induced to differentiate at non-permissive temperature of cell growth, accompanied by changes in the expression of various genes, whereas others were not. During the differentiation induced by shifting up to the non-permissive temperature, a rapid and transient decrease in both c-myc and p53 mRNA levels and rapid induction of c-jun mRNA were observed. These changes were specific in differentiation-inducible mutants and were not observed in a non-inducible mutant. In both types of mutants, the level of c-myc mRNA decreased in association with growth retardation at the non-permissive temperature. The p53 mRNA, however, showed specific increase in the differentiation-inducible ts mutants. These observations suggest distinct roles for p53 and c-myc from proliferation to differentiation in teratocarcinoma stem cells. PMID: 1461650 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 796: J Dent Res. 1992 Dec;71(12):1885-90. Effects of sialagogues on ornithine decarboxylase induction and proto-oncogene expression in murine parotid gland. Kawano M, Ueno A, Ashida Y, Matsumoto N, Inoue H. Department of Biochemistry, School of Dentistry, University of Tokushima, Japan. The mechanism of a sialagogue-induced increase in ornithine decarboxylase (ODC) activity and the expressions of proto-oncogenes in murine parotid gland were investigated by use of isoproterenol (IPR), carbachol (CC), and methoxamine (MTX). The results were as follows: (1) The three sialagogues had similar effects on the parotid in vivo (mouse parotid after a single injection of IPR) and/or in vitro (rat parotid explants cultured on siliconized lens paper floating on 199 medium containing IPR, CC, or MTX), the order of their effectiveness being IPR > CC > MTX. (2) Northern/dot and Western blot analyses revealed that the sialagogues elevated the steady-state levels of ODC mRNA and ODC protein to maxima at two h and six h, respectively, after stimulation. The increases were roughly proportional to those in ODC activity, suggesting that sialagogue-dependent enzyme induction is regulated at the transcriptional level. (3) The mRNAs of four of nine proto-oncogenes examined showed sialagogue-dependent increases to maxima at 30 min (c-fos) or 60 min (c-jun, c-myc, and c-src) after the beginning of stimulation. These increases were all transient, with the levels returning to the control values (without sialagogue) within 60 min. (4) The IPR-dependent elevations of ODC activity and the mRNAs of ODC, c-fos, and c-jun were inhibited by monensin, but not by polymyxin B. On the other hand, the CC-dependent increases in these parameters were inhibited by polymyxin B but not by monensin.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1452888 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 797: Blood. 1992 Dec 1;80(11):2883-90. Granulocyte-macrophage colony-stimulating factor/interleukin-3 fusion protein (pIXY 321) enhances high-dose Ara-C-induced programmed cell death or apoptosis in human myeloid leukemia cells. Bhalla K, Tang C, Ibrado AM, Grant S, Tourkina E, Holladay C, Hughes M, Mahoney ME, Huang Y. Division of Hematology/Oncology, Medical University of South Carolina, Charleston 29425. High dose Ara-C (HIDAC) induces programmed cell death (PCD) or apoptosis in vitro in human myeloid leukemia cells, which correlates with the inhibition of their clonogenic survival. Hematopoietic growth factors (HGFs) granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3) have been demonstrated to enhance the metabolism and cytotoxic effects of HIDAC against leukemic progenitor cells. We examined the effect of pIXY 321 (a GM-CSF/IL-3 fusion protein) on HIDAC-induced PCD and related gene expressions as well as HIDAC-mediated colony growth inhibition of human myeloid leukemia cells. Unlike the previously described effects of HGFs on normal bone marrow progenitor cells, exposure to pIXY 321 alone for up to 24 hours did not suppress PCD in HL-60 or KG-1 cells. However, exposure to pIXY 321 for 20 hours followed by a combined treatment with Ara-C plus pIXY 321 for 4 or 24 hours versus treatment with Ara-C alone significantly enhanced the oligonucleosomal DNA fragmentation characteristic of PCD. This was temporally associated with a marked induction of c-jun expression and a significant decrease in BCL-2. In addition, the treatment with pIXY 321 plus HIDAC versus HIDAC alone produced a significantly greater inhibition of HL-60 colony growth. These findings highlight an additional mechanism of HIDAC-induced leukemic cell death that is augmented by cotreatment with pIXY 321 and may contribute toward an improved antileukemic activity of HIDAC. PMID: 1450413 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 798: Mol Cell Biol. 1992 Dec;12(12):5355-62. Mitogenic signaling by colony-stimulating factor 1 and ras is suppressed by the ets-2 DNA-binding domain and restored by myc overexpression. Langer SJ, Bortner DM, Roussel MF, Sherr CJ, Ostrowski MC. Department of Microbiology and Immunology, Duke University Medical Center, Durham, North Carolina 27710. The activity of p21ras is required for the proliferative response to colony-stimulating factor 1 (CSF-1), and signals transduced by both the CSF-1 receptor (CSF-1R) and p21ras stimulate transcription from promoter elements containing overlapping binding sites for Fos/Jun- and Ets-related proteins. A sequence encoding the DNA-binding domain and nuclear localization signal of human c-ets-2, which lacked portions of the c-ets-2 gene product necessary for trans activation, was fused to the bacterial lacZ gene and expressed from an actin promoter in NIH 3T3 cells expressing either the v-ras oncogene or human CSF-1R. Nuclear expression of the Ets-LacZ protein, confirmed by histochemical staining of beta-galactosidase, inhibited the activity of ras-responsive enhancer elements and suppressed morphologic transformation by v-ras as well as CSF-1R-dependent colony formation in semisolid medium. When CSF-1R-bearing cells expressing the Ets-LacZ protein were stimulated by CSF-1, induction of c-ets-2, c-jun, and c-fos ensued, but the c-myc response was impaired. Enforced expression of the c-myc gene overrode the suppressive effect of ets-lacZ and restored the ability of these cells to form colonies in response to CSF-1. NIH 3T3 cells engineered to express a CSF-1R (Phe-809) mutant similarly cannot form CSF-1-dependent colonies in semisolid medium and exhibit an impaired c-myc response, but expression of an exogenous myc gene resensitizes these cells to CSF-1 [M. F. Roussel, J. L. Cleveland, S. A. Shurtleff, and C. J. Sherr, Nature (London) 353:361-363, 1991]. The ability of these cells to respond to CSF-1 was also rescued by enforced expression of an endogenous c-ets-2 gene. The ets family of transcription factors therefore plays a central role in integrating both CSF-1R and ras-induced mitogenic signals and in modulating the myc response to CSF-1 stimulation. PMID: 1448070 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 799: Circ Res. 1992 Dec;71(6):1351-60. Proto-oncogene expression in porcine myocardium subjected to ischemia and reperfusion. Brand T, Sharma HS, Fleischmann KE, Duncker DJ, McFalls EO, Verdouw PD, Schaper W. Department of Experimental Cardiology, Max Planck Institute of Physiological and Clinical Research, Bad Neuheim, FRG. The molecular basis of myocardial adaptation to ischemia and reperfusion is poorly understood. It is thought that nuclear proto-oncogenes act as third messengers, converting cytoplasmic signal transduction into long-term changes of gene expression. We studied the expression of six nuclear proto-oncogenes (Egr-1, c-fos, fosB, c-jun, junB, and c-myc) in myocardium subjected to ischemia and reperfusion in anesthetized pigs. Stunning was achieved by two 10-minute left anterior descending coronary artery occlusions separated by 30 minutes of reperfusion. Hearts were excised after the first occlusion, after the first reperfusion, and at 30, 120, 150, and 210 minutes of reperfusion after the second occlusion. Total RNA was prepared from stunned as well as normally perfused myocardial tissue and subjected to Northern blotting. The response of the six nuclear proto-oncogenes varied.fosB gene expression was never detected. The c-myc gene was expressed, but its level was unchanged by ischemia. c-jun expression was slightly increased by ischemia (3.1 +/- 0.6-fold). The c-fos, Egr-1, and junB genes were highly induced, being fivefold to sevenfold higher in experimental than in control tissue. In three animals pretreated with the beta 1-antagonist metoprolol and then subjected to the above experimental protocol, the induction of proto-oncogenes was similar to that in nonblocked controls. Our results show that the myocardial adaptive response to ischemic stress includes the induction of at least four transcription factors that may be further operative in repair processes and angiogenesis. PMID: 1385005 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 800: Nippon Rinsho. 1992 Dec;50(12):3056-63. [Proto-oncogene, proliferating cell nuclear antigen, perforin and growth factor gene expression in peripheral blood mononuclear cells from patients with IgA nephropathy] [Article in Japanese] Nakamura T, Ebihara I, Koide H. Department of Medicine, Juntendo University School of Medicine. IgA nephropathy, which is widely recognized as one of the most common primary glomerulonephritides, is characterized by the constant presence of IgA in the glomerular mesangium. We investigated proto-oncogenes, proliferating cell nuclear antigen (PCNA), perforin, platelet-derived growth factor (PDGF) and insulin-like growth factor (IGF) mRNA expression in peripheral blood mononuclear cells (PBMC) obtained from patients with IgA nephropathy, patients with other types of glomerulonephritis and healthy age-matched controls. The majority of patients with IgA nephropathy showed elevated c-myc, c-fos, c-jun, c-raf, PCNA, perforin, PDGF-B chain, and IGF-I and -II mRNA expression in PBMC, while no these mRNA expression was detected in PBMC obtained from patients with other types of glomerulonephritis or normal controls. A positive correlation was noted between mRNA levels and urinary protein excretion. mRNA expression also correlated with the histopathological findings in the renal tissue of patients with IgA nephropathy. These studies suggest that abnormal regulation of proto-oncogenes, PCNA, perforin, PDGF and IGF gene expression in PBMC may be associated with the progression of IgA nephropathy and may be useful as an indicator of disease activity. Publication Types: Review PMID: 1362782 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 801: Nihon Kyobu Shikkan Gakkai Zasshi. 1992 Dec;30 Suppl:27-32. [Studies on the regulation of normal bronchial epithelial cell proliferation and proto-oncogene expression] [Article in Japanese] Takizawa H. Department of Medicine and Physical Therapy, Tokyo University School of Medicine, Japan. The regulatory mechanisms of proliferation and differentiation of normal airway epithelial cells seem important for the better understanding of lung carcinogenesis. We isolated bovine and human bronchial epithelial cells by enzymatic digestion of bronchi, and cultured them in serum-free growth factor-supplemented medium. To determine the role of growth factors in the proliferation of bovine bronchial epithelial cells, the effect of each of these additives was evaluated. Of these, fetal calf serum (FCS), bovine pituitary extract (BPE), insulin and insulin-like growth factor-1 (IGF-1) showed a significant stimulatory effect on cell growth. These four additives induced transient increases in c-fos, c-jun and c-myc proto-oncogene mRNA levels. Transforming growth factor beta (TGF beta) inhibited growth factor-induced cell proliferation, and also showed selective inhibition of c-myc induction. We also studied the effect of interleukin 6 (IL-6) on human bronchial epithelial cell proliferation. IL-6 showed a significant inhibitory effect on cell growth. These cells were capable of expressing and releasing IL-6 and had specific receptors for IL-6, suggesting an autocrine mechanism. These results suggest that TGF beta and IL-6 may play roles in the regulation of airway epithelial cell growth as inhibitory growth factors. PMID: 1306233 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 802: Pathol Res Pract. 1992 Dec;188(8):1104-21. What's new in the role of cytokines on osteoblast proliferation and differentiation? Zheng MH, Wood DJ, Papadimitriou JM. Department of Orthopaedics, University of Western Australia, Queen Elizabeth II Medical Centre, Nedlands. This review assesses recent data concerning the role of cytokines produced by a variety of cells in bone on osteoblast function. The following themes are presumed: (1) osteoblasts are mesenchymal cells which act as either the major cellular agents of bone formation or as modulators of bone resorption by osteoclasts. The regulation of osteoblast proliferation and differentiation may involve a negative feedback process resulting in phenotype suppression; (2) cytokines including platelet-derived growth factors (PDGF), parathyroid hormone-related proteins (PTHrP), bone morphogenic proteins (BMP), transforming growth factor beta (TGF beta), fibroblast growth factors (FGF), insulin-like growth factors (IGF), epidermal growth factors (EGF), interleukin-1 and 6, tumour necrosis factors (TNF), interferon and haematopoietic growth factors have effects on osteoblast differentiation and proliferation but their effectiveness may not be identical in vitro and in vivo; (3) finally, therapeutic strategies for cytokine use in clinical practice are considered. Publication Types: Review Review, Tutorial PMID: 1300606 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 803: Ann N Y Acad Sci. 1992 Nov 21;663:202-14. Proliferation, differentiation arrest, and survival in leukemic blast cells. Ferrari S, Manfredini R, Grande A, Torelli G, Torelli U. Experimental Hematology Center, University of Modena, Italy. Publication Types: Review Review, Tutorial PMID: 1482054 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 804: Ann N Y Acad Sci. 1992 Nov 21;663:158-66. Oxidant carcinogenesis and antioxidant defense. Cerutti P, Shah G, Peskin A, Amstad P. Department of Carcinogenesis, Swiss Institute for Experimental Cancer Research, Epalinges/Lausanne. Growth promotion by oxidants is observed with cultured human and mouse fibroblasts as well as epidermal cells. It is expected to play a role in inflammation, fibrosis, and tumorigenesis. Indeed, oxidants trigger (patho)physiological reactions that resemble those induced by growth and differentiation factors. For example, active oxygen activates protein kinases, causes DNA breakage, and induces the growth competence-related protooncogenes c-fos and c-myc. The cellular antioxidant defenses affect the consequences of oxidant exposure. Transfectants of mouse epidermal cells that overproduce Cu,Zn-superoxide dismutase (SOD) were sensitized to the toxic effects of an extracellular burst of O2-. plus H2O2, whereas overproducers of catalase (CAT) were protected. Transfection of SOD overproducers with CAT corrected their hypersensitivity. Inducibility of the protooncogene c-fos by oxidants was diminished in SOD and CAT overproducers, albeit probably for different reasons. It is concluded that a fine balance of the multiple components of the antioxidant defense determines the growth response of cells to oxidative stress. In studies of the mechanism of the transcriptional induction of c-fos by oxidants, we identified the joint DSE-AP1 elements (dyad symmetry element, DSE) as major enhancer motifs in the 5'-upstream regulatory sequences of c-fos. Oxidants also increased the de novo synthesis of protein factors that bind to the fos-AP1 enhancer motif. Protein kinase and ADPR transferase inhibitors suppressed the transcriptional induction of c-fos as well as the increase in factor binding to fos-AP1. We conclude that protein phosphorylation and protein polyADP-ribosylation are required for the transcriptional induction of c-fos and the synthesis of protein factors that bind to fos-AP1. It is likely that the FOS and JUN proteins are among these factors and that they participate in the regulation of c-fos expression by oxidants. PMID: 1482049 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 805: Biochem Biophys Res Commun. 1992 Nov 16;188(3):1261-6. Studies of the relationship between cell proliferation and cell death. II. Early gene expression during concanavalin A-induced proliferation or dexamethasone-induced apoptosis of rat thymocytes. Grassilli E, Carcereri de Prati A, Monti D, Troiano L, Menegazzi M, Barbieri D, Franceschi C, Suzuki H. Istituto di Patologia Generale, Universita di Modena, Italy. Several data in the literature suggest that an intriguing relationship exists between cell proliferation and cell death. Accordingly, we studied the early expression of different genes in the same cells, i.e. rat thymocytes, undergoing cell proliferation upon stimulation with Concanavalin A or cell death following dexamethasone treatment. We showed that an early accumulation of c-fos, c-jun and c-myc mRNA occurred in both phenomena but with different kinetics. It can be speculated that the early induction of nuclear oncogenes is necessary to allow the later induction of other genes probably regulated at the transcriptional level by the AP-1 complex and/or by Myc protein. The accumulation of the transcript for another gene, i.e. poly(ADP-ribose)polymerase, an enzyme responsible for posttranslational modifications of several nuclear proteins, could instead be related to chromatin modifications occurring in both processes. PMID: 1445359 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 806: Biochem Biophys Res Commun. 1992 Nov 16;188(3):1067-76. c-fos, c-jun and c-myc expressions are not growth rate limiting for the human MCF-7 breast cancer cells. Wosikowski K, Eppenberger U, Kung W, Nagamine Y, Mueller H. Department of Research, University Clinics, Kantonsspital Basel, Switzerland. Insulin-like growth factor I (IGF-I) was 3 times more potent in pagating MCF-7 cell proliferation than epidermal growth factor (EGF). IGF-I stimulated c-fos mRNA expression about 5 times less than EGF. Both growth factors were equipotent in inducing c-jun and c-myc mRNA expressions. The protein level of c-Myc correlated with the mRNA level. IGF-I and EGF stimulated the transcriptional activity dependent on the phorbol 12-myristate 13-acetate-responsive element (TRE) to the same extent, when measured by the chloramphenicol acetyl transferase activity of a transiently transfected multiple TRE construct. These results strongly indicate that the expression levels of the measured proto-oncogenes do not correlate with the increase of growth stimulation by IGF-I and EGF and are not growth rate limiting for the human MCF-7 breast cancer cells. PMID: 1445344 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 807: Nature. 1992 Nov 12;360(6400):177-9. Erratum in: Nature 1992 Dec 3;360(6403):491. Abrogation by c-myc of G1 phase arrest induced by RB protein but not by p53. Goodrich DW, Lee WH. Center for Molecular Medicine, University of Texas Health Science Center, San Antonio 78245. Inactivating mutations of the retinoblastoma gene (RB) are found in a wide variety of tumour cells. Replacement of wild-type RB can suppress the tumorigenicity of some of these cells, suggesting that the RB protein (Rb) may negatively regulate cell growth. As activation of c-myc expression promotes cell proliferation and blocks differentiation, it may positively regulate cell growth. The c-myc protein is localized in the nucleus and can physically associate with RB protein in vitro, hence c-myc may functionally antagonize RB function. Microinjection of Rb in G1 phase reversibly arrests cell-cycle progression. Here we co-inject RB protein with c-myc, EJ-ras, c-fos or c-jun protein. Co-injection of c-myc, but not EJ-ras, c-fos or c-jun, inhibits the ability of Rb to arrest the cell cycle. The c-myc does not inhibit the activity of another tumour supressor, p53 (ref. 12). Thus, c-myc and RB specifically antagonize one another in the cell. PMID: 1436095 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 808: Blood. 1992 Nov 1;80(9):2298-305. Retinoic acid inhibits interleukin-6-induced macrophage differentiation and apoptosis in a murine hematopoietic cell line, Y6. Oritani K, Kaisho T, Nakajima K, Hirano T. Division of Molecular Oncology, Osaka University Medical School, Japan. We established a radiation-induced murine hematopoietic cell line, Y6, that could be induced to differentiate into macrophages by interleukin-6 (IL-6). IL-6 also induced growth inhibition and apoptosis in Y6 cells. Retinoic acid (RA) inhibited such effects of IL-6 on Y6 cells. The inhibitory effect of RA on the effects of IL-6 was not caused by the downregulation of the IL-6 receptor, because RA neither affected the expression of IL-6 receptor mRNA nor the expression of IL-6 receptor molecule on the cell surface. Furthermore, RA did not inhibit the IL-6-induced expression of junB mRNA, indicating that the expression of functionally active IL-6 receptor and the signal transduction pathway activating the junB gene are not inhibited by RA. IL-6-induced macrophage differentiation of Y6 cells was preceded by the downregulation of the c-myc gene, which was also prevented by RA. Because the inhibitory effect of RA on Y6 cells was reversible and seemed not to require de novo protein synthesis, the RA receptor by itself might be directly involved in the inhibition of the IL-6 signal transduction pathway. The results indicated that the IL-6 signal transduction pathways leading to the induction of macrophage differentiation and junB gene expression can be dissected by RA. PMID: 1384800 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 809: Biokhimiia. 1992 Oct;57(10):1467-71. [An unusual leucine motif in amino acid sequences of subunit 9 of ATP-synthase from plant mitochondria, modified by mRNA editing] [Article in Russian] Konstantinov IuM, Derenko MV. The amino acid sequences of subunit 9 of plant mitochondrial ATP-synthase contain a leucine motif which differs from the leucine repeat earlier detected within the composition of proteinaceous products of Myc, Fos, and Jun proto-oncogens. The structural organization of this repeat in proteolipid sequences can be modified via transformation of serine triplets into leucine ones as a result of editing of an appropriate mRNA. PMID: 1457594 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 810: Virus Res. 1992 Oct;26(1):71-90. Alterations in the levels of expression of specific cellular genes in adenovirus-infected and -transformed cells. Rosahl T, Doerfler W. Institute for Genetics, University of Cologne, Germany. There is increasing evidence that changes in the transcriptional program of cellular genes in virus-transformed cells can contribute to virus transformation. It is, therefore, important to study altered expression patterns of cellular genes in adenovirus-infected and -transformed cells. We have used 40 different cellular genes or gene segments as hybridization probes to analyze the cytoplasmic RNA from adenovirus type 2 (Ad2)-infected KB cells, from Ad5-transformed human cells (293) or from several Ad2- or adenovirus type 12 (Ad12)-transformed hamster cell lines. Many of the genes probed were not expressed in human or hamster cells. Transcription of the ADPRT and the heat shock protein 70 genes was increased in Ad2-infected KB cells and in 293 cells. In Ad2-infected KB cells, c-myc gene transcription was decreased. In 293 cells and in three adenovirus-transformed hamster cell lines (T637, BHK21-Ad2E1A-E1B, and BHK21-Ad2 HindIII-G), the transcription of the c-jun gene was increased, whereas c-myc transcription was decreased in the latter two cell lines. The data presented here demonstrate that, among 40 different mammalian gene probes, alterations in steady state levels of RNA were detected for five of these genes. These results suggest major alterations in transcription patterns in adenovirus-infected and -transformed cells. PMID: 1441738 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 811: Biochem J. 1992 Oct 1;287 ( Pt 1):1-15. Nuclear protein phosphorylation and growth control. Meek DW, Street AJ. Department of Biochemistry, University of Dundee, Scotland, U.K. Publication Types: Review PMID: 1417761 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 812: Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):9210-4. Growth arrest induced by wild-type p53 protein blocks cells prior to or near the restriction point in late G1 phase. Lin D, Shields MT, Ullrich SJ, Appella E, Mercer WE. Department of Microbiology and Immunology, Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, PA 19107. Conditional expression of wild-type (wt) p53 protein in a glioblastoma tumor cell line has been shown to be growth inhibitory. We have now more precisely localized the position in the cell cycle where growth arrest occurs. We show that growth arrest occurs prior to or near the restriction point in late G1 phase of the cell cycle. The effect of wt p53 protein on the expression of four immediate-early genes (c-FOS, c-JUN, JUN-B, and c-MYC), one delayed-early gene (ornithine decarboxylase), and two late-G1/S-phase genes (B-MYB and DNA polymerase alpha) was also examined. Of this subset of growth response genes, only the expression of B-MYB and DNA polymerase alpha was significantly repressed. The possibility that decreased expression of B-MYB may be an important component of growth arrest mediated by wt p53 protein is discussed. PMID: 1409626 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 813: Mol Cell Biol. 1992 Oct;12(10):4612-21. Downregulation of JE and KC genes by glucocorticoids does not prevent the G0----G1 transition in BALB/3T3 cells. Rameh LE, Armelin MC. Instituto de Quimica, Universidade de Sao Paulo, Brazil. The effects of glucocorticoid hormones on the expression of the growth factor-inducible genes JE, KC, and c-myc were analyzed in parental BALB/3T3 and polyomavirus middle-T antigen-transfected cell lines. Northern (RNA) blot hybridization and run-on transcription analysis showed that (i) glucocorticoid hormones selectively inhibit JE and KC expression at the transcriptional level and (ii) the downregulatory effect of glucocorticoids on JE and KC expression is partial for serum-stimulated and middle T antigen-transformed cells and total for quiescent and exponentially growing cells. Gel mobility assays using AP-1 oligonucleotides showed a positive correlation between glucocorticoid downregulating effect and presence of the AP-1 complex. JE and KC downregulation by means of the AP-1 complex may play a role in the actions of glucocorticoids as anti-inflammatory and antitumor agents. The ability of glucocorticoids to downregulate JE and KC was used to investigate the relevance of these genes to the mitogenic response to serum growth factors. Hydrocortisone did not alter the basal DNA synthesis level displayed by quiescent 3T3 cells, but it potentiated both the mitogenic effect of platelet-derived growth factor and c-myc induction by serum growth factors. Upon serum restimulation, untreated and dexamethasone-treated quiescent 3T3 cultures entered the S phase after an identical time lag (G1). These results suggest that (i) JE and KC are not necessary for the G0----G1----S transition and (ii) c-myc overexpression is likely to be the basis for the potentiating effect of glucocorticoids on serum growth factors. PMID: 1406651 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 814: Biochim Biophys Acta. 1992 Sep 24;1132(2):140-4. Magnetic field-induced changes in specific gene transcription. Phillips JL, Haggren W, Thomas WJ, Ishida-Jones T, Adey WR. Jerry L. Pettis Memorial VA Hospital, Research Service, Loma Linda 92357. Magnetic fields are physical, environmental agents that have been shown to produce a variety of responses in cellular and animal studies, including general changes in gene transcription. In this study, the nuclear run-off assay has been employed to assess alterations in specific gene transcription in CEM-CM3 T-lymphoblastoid cells exposed for 15-120 min to a 1 gauss sinusoidal magnetic field at 60 Hz. Time-dependent and cell density-dependent changes in the transcription of c-fos, c-jun, c-myc and protein kinase C (beta-form) have been observed and quantitated. Additionally, changes in transcript levels, assessed by slot-blot analysis, have been found to parallel the changes in gene transcription. These data suggest an important role for magnetic field exposure in altering cellular processes. PMID: 1390886 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 815: Mol Cell Biochem. 1992 Sep 22;115(1):85-96. Vanadate-induced gene expression in mouse C127 cells: roles of oxygen derived active species. Yin X, Davison AJ, Tsang SS. Bioenergetics Research Laboratory, Faculty of Applied Sciences, Simon Fraser University, Burnaby, British Columbia, Canada. An underinvestigated aspect of the mitogenic and cell regulatory actions of vanadium is the regulation of gene expression. Among the fifteen cellular genes studied in cultured mouse C127 cells, vanadium (as 10 microM sodium vanadate) increased levels of mRNA of the actin and c-Ha-ras to four times control values. These increases represented de novo synthesis of mRNA, since they were inhibited by actinomycin D. Vanadate did not increase mRNA corresponding to c-src, c-mos, c-myc, p53, HSP70, pODC or RB genes, and expression of c-erb A, c-erb B, c-sis and c-fes genes was undetectable whether vanadium was present or not. Expression of a third gene affected by vanadium, c-jun, was augmented by addition of a reductant or oxidant together with the vanadate. Addition of NADH (marginally effective on its own) or H2O2 (effective alone) dramatically enhanced the effect of vanadate on c-jun gene expression. Catalase inhibited the effect of NADH partly. The vanadate-stimulated expression of actin and c-Ha-ras mRNA were unaffected by oxidants, reductants, metal chelators, or anti-oxidant enzymes. Evidently vanadate acts by two separate mechanisms on these two categories of genes. The alternate hypothesis that the actions of vanadate on actin and c-Ha-ras were mediated by a protein kinase cascade was inconsistent with the following observations. Neither insulin nor epidermal growth factor increased mRNA levels of c-Ha-ras or actin gene. Neither genistein (a tyrosine kinase inhibitor) nor pretreatment with 12-O-tetradecanoylphorbol-13-acetate blocked the actions of vanadate on these genes. Clearly the biological actions of vanadium depend in part on altered expression of genes. Since two of the genes are proto-oncogenes, this mechanism is potentially relevant to the mitogenic responses of cells to vanadium. PMID: 1435769 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 816: J Biol Chem. 1992 Sep 5;267(25):17498-501. Analysis of the oligomerization of myogenin and E2A products in vivo using a two-hybrid assay system. Chakraborty T, Martin JF, Olson EN. Department of Biochemistry and Molecular Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030. Members of the helix-loop-helix (HLH) family of proteins bind DNA and activate transcription as homo- and heterodimers. Myogenin is a muscle-specific HLH protein that binds DNA in vitro as a heterodimer with several widely expressed HLH proteins, such as the E2A gene products E12 and E47. We describe a method for detection of protein-protein interactions among HLH proteins in vivo in which dimerization through the HLH motif reconstructs a hybrid transcription factor containing the DNA-binding domain of yeast GAL4 linked to one HLH motif and the activation domain of VP-16 linked to another. We have used this assay to investiagate whether myogenin forms homomeric or heteromeric complexes in vivo and to determine whether growth factors and oncogenes that inhibit myogenesis influence myogenin's ability to dimerize. The results show that myogenin heterodimerizes with E12 and E47 in vivo, but it does not homodimerize to a measurable extent. Peptide growth factors, as well as the immediate early gene products c-Jun, v-Fos, and c-Myc, inhibit the activity of myogenin through a mechanism independent of its association with E2A products. PMID: 1325437 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 817: Exp Cell Res. 1992 Sep;202(1):161-6. An altered repertoire of fos/jun (AP-1) at the onset of replicative senescence. Irving J, Feng J, Wistrom C, Pikaart M, Villeponteau B. Department of Biological Chemistry, University of Michigan, Ann Arbor 48105-2007. With multiple divisions in culture, normal diploid cells suffer a loss of growth potential that leads to replicative senescence and a finite replicative capacity. Using quantitative RT-PCR, we have monitored mRNA expression levels of c-fos, c-jun, JunB, c-myc, p53, H-ras, and histone H4 during the replicative senescence of human fibroblasts. The earliest and the largest changes in gene expression occurred in c-fos and junB at mid-senescence prior to the first slowing in cell growth rates. The basal level of c-fos mRNA decreased to one-ninth that of the early-passage levels, while junB declined to one-third and c-jun expression remained constant. The decline in the basal c-fos mRNA level in mid-senescence should lead to an increase in Jun/Jun AP-1 homodimers at the expense of Fos/Jun heterodimers and may trigger a cascade of further changes in c-myc, p53, and H-ras expression in late-passage senescent fibroblasts. PMID: 1511730 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 818: Biol Reprod. 1992 Sep;47(3):428-35. Regulation of proto-oncogene expression and deoxyribonucleic acid synthesis in granulosa cells of perifused immature rat ovaries. Delidow BC, Lynch JP, White BA, Peluso JJ. Department of Obstetrics and Gynecology, University of Connecticut Health Center, Farmington 06032. The present series of experiments examined the effects of follicle-stimulating hormone (FSH) and insulin (IN) on granulosa cell (GC) proto-oncogene expression and DNA synthesis. In the first study, GCs were harvested from immature rat ovaries after 15, 30, or 60 min of perifusion and DNA synthesis (3H-thymidine incorporation) and proto-oncogene mRNA levels were determined. The presence of c-myc and c-fos proteins was localized within GCs immunocytochemically. GCs of control ovaries exhibited modest levels of DNA synthesis and proto-oncogene expression. FSH/IN not only stimulated DNA synthesis but also increased c-myc, c-fos, and c-jun mRNA levels and the percentage of cells staining for c-fos and c-myc proteins. The protein kinase inhibitor, 2-aminopurine (2-AP), inhibited the FSH/IN-induced increases in c-myc and c-fos mRNA levels, the percentage of cells staining for Myc and Fos protein, and DNA and protein synthesis. The effects of 48 h of perifusion with FSH in the presence or absence of IN were also examined. These treatments were selected because after 48 h of continuous exposure to FSH alone, estradiol-17 beta (E2) secretion is enhanced and 3H-thymidine incorporation is inhibited. Conversely, FSH/IN maintains 3H-thymidine incorporation for up to 48 h of perifusion culture without stimulating E2 (Peluso et al., Endocrinology 1991; 128:191-196). After 48 h of perifusion, both FSH and FSH/IN stimulated c-fos mRNA and protein levels. However, high levels of c-jun mRNA and protein were detected only within GCs of FSH/IN-treated ovaries.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1511096 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 819: Mol Cell Biol. 1992 Sep;12(9):3890-902. Overexpression of Mos, Ras, Src, and Fos inhibits mouse mammary epithelial cell differentiation. Jehn B, Costello E, Marti A, Keon N, Deane R, Li F, Friis RR, Burri PH, Martin F, Jaggi R. Laboratory for Clinical and Experimental Research, University of Bern, Switzerland. Mammary epithelial cells terminally differentiate in response to lactogenic hormones. We present evidence that oncoprotein overexpression is incompatible with this hormone-inducible differentiation and results in striking cellular morphological changes. In mammary epithelial cells in culture, lactogenic hormones (glucocorticoid and prolactin) activated a transfected beta-casein promoter and endogenous beta-casein gene expression. This response to lactogenic hormone treatment was paralleled by a decrease in cellular AP-1 DNA-binding activity. Expression of the mos, ras, or src (but not myc) oncogene blocked the activation of the beta-casein promoter induced by the lactogenic hormones and was associated with the maintenance of high levels of AP-1. Mos expression also increased c-fos and c-jun mRNA levels. Overexpression of Fos and Jun from transiently transfected constructs resulted in a functional inhibition of the glucocorticoid receptor in these mouse mammary epithelial cells. This finding clearly suggests that glucocorticoid receptor inhibition arising from oncogene expression will contribute to the block in hormonally induced mammary epithelial cell differentiation. Expression of Src resulted in the loss of the normal organization and morphological phenotype of mammary epithelial cells in the epithelial/fibroblastic line IM-2. Activation of a conditional c-fos/estrogen receptor gene encoding an estrogen-dependent Fos/estrogen receptor fusion protein also morphologically transformed mammary epithelial cells and inhibited initiation of mammary epithelial differentiation-associated expression of the beta-casein and WDNM 1 genes. In response to estrogen treatment, the cells displayed a high level of AP-1 DNA-binding activity. Our results demonstrate that high cellular AP-1 levels contribute to blocking the ability of mammary epithelial cells in culture to respond to lactogenic hormones. This and other studies indicate that the oncogene products Mos, Ras, and Src exert their effects, at least in part, by stimulating cellular Fos and probably cellular Jun activity. PMID: 1508191 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 820: J Environ Pathol Toxicol Oncol. 1992 Sep-Oct;11(5-6):345. The effects of radiation on oncogenes. Li YJ, Wang DW. Institute of Radiation Medicine, Academy of Military Medical Science, Beijing, P. R. China. PMID: 1464820 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 821: Am J Physiol. 1992 Sep;263(3 Pt 1):G327-32. CCK, bombesin, and carbachol stimulate c-fos, c-jun, and c-myc oncogene expression in rat pancreatic acini. Lu L, Logsdon CD. Department of Physiology, University of Michigan, Ann Arbor 48109-0622. To identify possible nuclear signals mediating long-term regulation of the pancreas by gastrointestinal hormones, the expression of c-fos, c-jun, and c-myc was investigated in rat pancreatic acini. Stimulation of the acini with cholecystokinin octapeptide (CCK-8, 100 pM), bombesin (10 nM), or carbachol (10 microM), but not gastrin (100 nM), secretin (100 nM), or vasoactive intestinal peptide (10 nM) induced an increase in oncogene mRNA expression. The percent increases of c-fos, c-jun, and c-myc mRNA were 207 +/- 40, 171 +/- 26, and 46 +/- 19 (n = 5) for CCK-8; 223 +/- 71, 159 +/- 31, and 43 +/- 21 (n = 5) for bombesin; and 125 +/- 51, 123 +/- 58, and 67 +/- 19 (n = 5) for carbachol, respectively. CCK-induced increases in oncogene mRNA were rapid and transient. c-fos and c-jun mRNA levels were increased after 30 min stimulation, peaked at 1 h, and returned to basal level in 2 h. Activation of c-myc was more prolonged with levels remaining elevated for at least 3 h. The effects of CCK-8 were concentration dependent. Detectable stimulation was seen at 10 pM; maximal stimulation occurred at 10 nM and was not affected by further increase in the concentration of CCK-8. JMV-180, a high-affinity site CCK receptor agonist and low-affinity site antagonist, alone did not stimulate c-fos mRNA expression but inhibited c-fos mRNA expression induced by CCK-8. These results suggest that the interaction between CCK and the low-affinity state of the CCK receptor is responsible for oncogene activation. PMID: 1415544 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 822: Oncogene. 1992 Sep;7(9):1837-45. Retinoic acid receptor alpha suppresses polyomavirus transformation and c-fos expression in rat fibroblasts. Talmage DA, Lackey RS. Institute of Human Nutrition, Columbia University, New York, New York 10032. To explore the molecular mechanisms by which retinoic acid inhibits oncogenic transformation, we have examined the effects of retinoic acid on the polyomavirus-induced transformation of rat fibroblasts. Treatment of rat F111 fibroblasts with high concentrations of retinoic acid (10(-6) M) partially inhibited the ability of polyomavirus to induce dense focus formation (50-70%). This effect was not secondary to a retinoic acid-dependent block of cellular proliferation. To address the role of the retinoic acid receptor (RAR-alpha) in mediating the transformation-inhibitory effect of retinoic acid, we have overexpressed either RAR-alpha or cellular retinoic acid-binding protein I (CRABP) cDNAs in F111 cells. Introduction of a CRABP I expression vector did not alter the responsiveness of F111 cells to retinoic acid in any detectable fashion. In contrast, overexpression of RAR-alpha increased the sensitivity of F111 cells to the transformation-inhibitory action of retinoic acid by 10- to 100-fold. At high concentrations, retinoic acid inhibited transformation of F111-RAR cells by polyomavirus by about 90%. At near physiological concentrations, retinoic acid inhibited transformation by 25-50% in F111-RAR cells but not in control cells. Retinoic acid did not inhibit either the synthesis of polyoma middle T (mT) or pp60c-src, the cellular target for mT action, or the formation of mT:pp60c-src:PI-3 kinase (phosphatidylinositol-3 kinase) complexes. Therefore, RAR-alpha was not acting as a negative regulator of expression of either the polyomavirus middle T oncogene or the cellular proto-oncogene, c-src. It seems likely that RAR-alpha regulates the expression of cellular genes whose products interact in some way with mT-regulated signaling pathways, leading to a ligand-dependent suppression of polyoma transformation. In addition, RAR-alpha overexpression selectively inhibits the serum-stimulated expression of the c-fos gene, but does not affect the expression of a number of other serum- and polyomavirus-inducible genes including c-jun, junB, c-myc and actin. PMID: 1380150 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 823: J Biochem (Tokyo). 1992 Sep;112(3):321-9. SV40 large T inhibits myogenic differentiation partially through inducing c-jun. Endo T. Department of Biology, Chiba University. Terminal differentiation of skeletal muscle cells is obligatorily accompanied by the expression of a battery of muscle-specific genes and cell fusion to form multinucleated myotubes. The transcription of some of the muscle-specific genes is activated by the products of myoD gene family including MyoD and myogenin. The mouse skeletal muscle cell line C2SVTts11, which is a clone of C2 cells transfected with SV40 T antigen genes (temperature-sensitive large T and wild-type small t) fused to metallothionein gene promoter, is prevented from differentiation when the large T is induced. If the large T is induced in the myotubes, which are preformed in the absence of large T expression, the terminally differentiated cells reenter the cell cycle. In good accordance with the induction of large T, endogenous c-jun but not c-fos or c-myc mRNA was induced, whereas the expression of myoD and myogenin was suppressed. Treatment of quiescent C2 cells with a tumor promoter, 12-O-tetradecanoylphorbol 13-acetate, transiently induced c-jun and c-fos mRNAs, and temporarily deinduced myoD and myogenin mRNAs just after the expression of the protooncogenes. To ascertain whether c-jun induced by large T is sufficient to inhibit myogenic differentiation, c-jun cDNA was transfected into C2 cells. As the levels of exogenous c-jun expression were higher in the transfected clones, the cells expressed lower levels of myoD gene family and they formed fewer myotubes. Even the cells expressing the highest levels of exogenous c-jun, however, still formed small myotubes containing a few nuclei under differentiation conditions. These results suggest that large T inhibits myogenic differentiation by suppressing the expression of the members of myoD gene family, partly through inducing c-jun. In addition to this, other mechanisms seem to be required to achieve complete inhibition. PMID: 1331038 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 824: J Cell Physiol. 1992 Aug;152(2):232-9. Modulation of mRNA levels during human keratinocyte differentiation. Younus J, Gilchrest BA. Department of Pathology, Boston University School of Medicine, Massachusetts 02118. Cultures of human keratinocytes provide an excellent model system in which to study differentiation. Using the phorbol ester 12-O-tetradecanoyl-phorbol 13-acetate (TPA) and calcium, two agents known to induce keratinocyte differentiation in vitro, we examined the expression of the genes encoding c-fos, c-myc, and c-jun; involucrin, a protein precursor of the keratinocyte cornified envelope; and L-7, a ribosomal protein. Overall, at the doses studied, TPA induced a more rapid and profound differentiation than did calcium, as evaluated by culture morphology and northern blot analysis. Our studies showed a constant low level of c-fos and c-jun expression in unstimulated cells with no significant change after addition of either TPA or calcium except when transcript breakdown was inhibited by cycloheximide. The c-myc proto-oncogene, known to have a high constitutive expression in actively proliferating cells, was strongly downregulated by TPA, but calcium had no effect over a 32 hour period, consistent with the greater growth inhibition of TPA in this system. Involucrin was induced about ninefold by both TPA and calcium as early as 8 hours after stimulation, suggesting transcriptional regulation of this gene during differentiation. L-7, recently demonstrated to be downregulated in late passage human fibroblasts in an in vitro model of senescence, was also strongly downregulated by either TPA or calcium, consistent with an interrelationship between the basic cellular processes of aging and differentiation. These finding expand our knowledge of the differentiation process in human keratinocytes. PMID: 1639858 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 825: Am J Physiol. 1992 Aug;263(2 Pt 1):C420-8. TGF-beta 1 potentiates growth factor-stimulated proliferation of vascular smooth muscle cells in genetic hypertension. Saltis J, Agrotis A, Bobik A. Baker Medical Research Institute, Alfred Hospital, Prahran, Victoria, Australia. We have examined the interactions between transforming growth factor-beta 1 (TGF-beta 1) and epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), or platelet-derived growth factor (PDGF) isoforms PDGF-AB and PDGF-BB on the proliferation of vascular smooth muscle cells isolated from the spontaneously hypertensive rat. TGF-beta 1 alone stimulated [3H]thymidine incorporation approximately twofold without a corresponding increase in cell number. In combination, TGF-beta 1 action was synergistic in further stimulating both DNA synthesis and cell proliferation 100-300% above the responses elicited by each growth factor. To gain further insight into the mechanism responsible for this potentiation, we examined the interaction between TGF-beta 1 and EGF. The synergistic interaction between TGF-beta 1 and EGF on DNA synthesis was independent of initial cell density. This effect of TGF-beta 1 was initiated early in the G1 phase of the cell cycle and did not appear to be mediated through the mobilization of Ca2+ or alterations in c-jun mRNA expression. However, in the presence of both TGF-beta 1 and EGF, there was a sustained elevation of c-myc mRNA levels over a 24-h period. These results suggest that TGF-beta 1 may interact with other growth factors in vivo to enhance their proliferative action on vascular smooth muscle of spontaneously hypertensive rats via mechanisms dependent on c-myc mRNA expression. PMID: 1514589 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 826: Gut. 1992 Aug;33(8):1033-8. Oncogenes and onco-suppressor gene in adenocarcinoma of the oesophagus. Jankowski J, Coghill G, Hopwood D, Wormsley KG. Department of Medicine, University of Dundee. While the activation of the proto-oncogenes has been implicated in the development and progression of cancer of many tissues, the role of oncogenes in the development of oesophageal adenocarcinoma has not been defined. Fifteen patients who had undergone resection for oesophageal adenocarcinoma and 15 who had undergone oesophagectomy or biopsy for Barrett's oesophagus were studied. The latter patients also had adjacent normal gastric mucosa biopsied for comparison with the metaplastic oesophageal mucosa. The mucosal samples were snap frozen and subsequently stained with monoclonal antibodies to the following oncogene associated proteins; c-erbB2 (neu and CE-1) (external domain), c-erbB2 (NCL-CB11) (internal domain), c-src, c-ras, c-myc, c-fos, c-jun, and the onco-suppressor gene--p53. All tumours were well or moderately differentiated adenocarcinomas arising from the lower third of the oesophagus. Eleven specimens showed strong membraneous staining with both c-erbB2 (neu) and c-erbB2 (CBL-CB11). Seven specimens showed strong nuclear staining with p53 onco-suppressor gene. Three specimens were positive for c-ras and c-src, and two were positive for c-jun. In Barrett's epithelium, nine specimens were positive for c-erbB2 (neu and CB11), three were positive for c-src, two were positive for c-ras and c-jun, and one was positive for c-fos. Two of the gastric mucosal biopsy specimens expressed c-erbB2 weakly but no other oncogenes were found. The frequency of positive staining for c-erbB2 is very high, compared with the expression of these genes in other tumours. It is also concluded that errors in the onco-suppressor gene p53, and especially in the external and internal domains of c-erbB2, which is also often expressed in Barrett's mucosa, may be implicated in the development of adenocarcinoma of the oesophagus. PMID: 1398227 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 827: Leukemia. 1992 Aug;6(8):796-800. BCR/ABL confers growth factor independence upon a murine myeloid cell line. Mandanas RA, Boswell HS, Lu L, Leibowitz D. Department of Medicine, Indiana University School of Medicine, Indianpolis. The BCR/ABL oncogene in chronic myelogenous leukemia produces an activated tyrosine kinase fusion protein (p210). Like other tyrosine kinase oncogenes, BCR/ABL can abrogate the interleukin-3 (IL-3) dependence of lymphoid cell lines. To investigate the ability of BCR/ABL to generate growth factor independence in myeloid cells, the IL-3 dependent myeloid cell line NFS/N1.H7 (H7) was transfected with the p210BCR/ABL-containing plasmid, pGD210. Stable clones A54 and A74 were capable of IL-3 independent growth and tumor formation in syngeneic mice. Relief of growth factor dependence was not mediated by autocrine release of IL-3. The baseline proliferation rate of the BCR/ABL transformed cells was greater than that of the parental H7 cells maximally stimulated by IL-3. Abundant constitutive expression of c-myc, c-jun, and c-fos was observed in the p210BCR/ABL transfectants even in low serum conditions. In contrast, c-myc expression in H7 cells was dependent upon IL-3 stimulation, and neither c-jun nor c-fos was highly expressed following IL-3 stimulation in H7 cells. Thus, BCR/ABL transformation and relief of IL-3 dependence involve not only pathways that can substitute for IL-3 induced growth via tyrosine kinase mediated signals, but also pathways that recruit constitutive c-jun and c-fos expression. PMID: 1379313 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 828: Cell. 1992 Jul 10;70(1):57-67. IL-2 and EGF receptors stimulate the hematopoietic cell cycle via different signaling pathways: demonstration of a novel role for c-myc. Shibuya H, Yoneyama M, Ninomiya-Tsuji J, Matsumoto K, Taniguchi T. Institute for Molecular and Cellular Biology, Osaka University, Japan. Stimulation via cytokine receptors such as IL-2 and IL-3 receptors, but not by the EGF receptor (EGFR), induces cells of the BAF-B03 hematopoietic cell line to transit the cell cycle. We demonstrate that the IL-2 receptor beta chain (IL-2R beta) is linked to at least two intracellular signaling pathways. One pathway may involve a protein tyrosine kinase of the src family, which leads to the induction of the c-jun and c-fos genes, among others. A second pathway, involving an as yet unknown mechanism, leads to c-myc gene induction. Stimulation of the EGFR, expressed following transfection of an appropriate recombinant construct, can activate the former, but not the latter, pathway in this cell line and cause the cells to enter S phase but not progress further. This deficiency can be rescued by ectopic expression of the c-myc gene, indicating a novel role for this proto-oncogene in the S to G2/M transition of the cell cycle. PMID: 1535827 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 829: Jpn J Cancer Res. 1992 Jul;83(7):723-8. A serine protease-inhibitory benzamidine derivative inhibits the growth of human colon carcinoma cells. Nishimura Y, Yasui W, Yoshida K, Matsuyama T, Dohi K, Tahara E. First Department of Pathology, Hiroshima University School of Medicine. The inhibitory effect of a serine protease-inhibiting tetra-benzamidine derivative, TAPP-Br, on the cell growth of 8 human colon carcinoma cell lines was examined and the mechanism of the inhibition was analyzed. TAPP-Br inhibited the cell growth of all the colon carcinoma cell lines, and this effect was irreversible. The expression of mRNAs for nuclear oncogenes such as MYC, FOS and JUN was decreased by TAPP-Br after treatment for 3 h and the effect continued for 48 h. mRNA expression of epidermal growth factor receptor, transforming growth factor-beta and type IV collagenase was suppressed at 48 h after the initiation of TAPP-Br treatment, suggesting an indirect action of TAPP-Br. TAPP-Br decreased protein kinase C activity in the particulate fraction, whereas it increased the enzyme activity in the soluble fraction. These findings overall suggest that the serine protease inhibitor, TAPP-Br, might inhibit the cell growth of colon carcinoma cell lines through suppressing the expression of genes whose promoter contains a 12-O-tetradecanoylphorbol-13-acetate-responsive element or serum-responsive element. PMID: 1517149 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 830: Cell Growth Differ. 1992 Jul;3(7):429-34. Stable induction of c-jun mRNA expression in normal human keratinocytes by agents that induce predifferentiation growth arrest. Blatti SP, Scott RE. Department of Pathology, University of Tennessee Medical School, Memphis 38163. A variety of agents can induce predifferentiation growth arrest (PGA) in human keratinocytes; these include transforming growth factor beta 1 (TGF-beta 1) and razoxane. We evaluated the ability of these and other agents to induce the expression of a variety of transcription factor genes including c-fos, c-myc, junB, and c-jun. The results show that both TGF-beta 1 and razoxane induce maximal c-jun mRNA expression 4 days after initiation of treatment concurrent with the development of PGA. In contrast, no detectable induction of c-fos, c-myc, or junB was observed. Keratinocytes maintained in the presence of TGF-beta 1 for an additional 3 days continued to show high levels of c-jun mRNA, indicating stable induction. Razoxane treatment also induces PGA and high c-jun mRNA levels for 4 days, but thereafter a decay of c-jun expression occurs. Run-off transcription experiments comparing rapidly growing cells with cells treated with TGF-beta 1 for 4 days demonstrated a significant increase in transcriptional activity of the c-jun gene. This result indicates that the increase in c-jun gene expression is due in part to a change in transcriptional regulation of c-jun. The stable induction of c-jun mRNA in keratinocytes at the PGA state is unique because the induction of this gene is usually transient. The finding that c-fos is not coinduced suggests that c-Jun homodimers or other AP-1 heterodimers may be formed at the PGA state to facilitate the stable induction of c-jun mRNA. This experimental system should therefore serve as a model system to study the molecular mechanisms for the stable control of c-jun gene expression and the control of AP-1-dependent gene expression during the process of keratinocyte differentiation. PMID: 1419906 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 831: Cell Growth Differ. 1992 Jul;3(7):391-9. Regulation of c-jun expression during induction of monocytic differentiation by okadaic acid. Kharbanda S, Datta R, Rubin E, Nakamura T, Hass R, Kufe D. Laboratory of Clinical Pharmacology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115. The present work has examined the effects of okadaic acid, an inhibitor of type 1 and 2A protein phosphatases, on the regulation of c-jun expression during monocytic differentiation of U-937 leukemia cells. The results demonstrate that okadaic acid treatment is associated with induction of a differentiated monocyte phenotype characterized by: (a) growth arrest; (b) increases in Mac-1 cell surface antigen expression; (c) down-regulation of c-myc transcripts; and (d) induction of tumor necrosis factor gene expression. This induction of monocytic differentiation was associated with transient increases in c-jun mRNA levels, which were maximal at 6 h. Similar effects were obtained for the c-fos gene. Run-on analysis demonstrated detectable levels of c-jun transcription in U-937 cells and that this rate is increased approximately 40-fold following okadaic acid exposure. c-jun mRNA levels were superinduced in cells treated with both okadaic acid and cycloheximide, whereas inhibition of protein synthesis had little, if any, effect on okadaic acid-induced c-jun transcription. The half-life of c-jun mRNA was similar (45-50 min) in both untreated and okadaic acid-induced cells. In contrast, treatment with both okadaic acid and cycloheximide was associated with stabilization (t 1/2 = 90 min) of c-jun transcripts. Taken together, these findings indicate that the induction of c-jun transcription by okadaic acid is controlled primarily by a transcriptional mechanism. Since previous studies have demonstrated that the c-jun gene is autoinduced by Jun/AP-1, we also studied transcription of c-jun promoter (positions -132/+170)-reporter gene constructs with and without a mutated AP-1 element.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1419903 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 832: Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5437-41. Brief visual experience induces immediate early gene expression in the cat visual cortex. Rosen KM, McCormack MA, Villa-Komaroff L, Mower GD. Massachusetts General Hospital, Department of Neurobiology, Harvard Medical School, Boston 02129. Brief visual experience causes rapid physiological changes in the visual cortex during early postnatal development. A possible mediator of these effects is the immediate early genes whose protein products are involved in the rapid response of neurons to transsynaptic stimulation. Here we report evidence that the levels of immediate early gene mRNAs in the visual cortex can be altered by manipulating the visual environment. Specifically, we find that brief (1 h) visual experience in dark-reared cats causes dramatic transient inductions of egr1, c-fos, and junB mRNAs in the visual cortex but not in the frontal cortex. Levels of c-jun and c-myc mRNAs are unaffected. These results suggest that select combinatorial interactions of immediate early gene proteins are an important step in the cascade of events through which visually elicited activity controls visual cortical development. PMID: 1376920 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 833: Endocrinology. 1992 Jun;130(6):3223-30. Progesterone inhibits the estrogen-induced expression of c-fos messenger ribonucleic acid in the uterus. Kirkland JL, Murthy L, Stancel GM. Department of Pediatrics, Baylor College of Medicine, Houston, Texas 77030. Estradiol produces a large increase in the uterine level of c-fos mRNA, which is maximum in 3 h. The administration of progesterone antagonizes this estrogen-induced increase in protooncogene transcript levels in both the rat and mouse. The inhibitory effect of progesterone is observed within 1 h after hormone treatment and persists for 9-18 h. In the rat, this effect can be observed at a dose of 0.25 mg progesterone and is maximum at a dose of 2.5 mg. A similar inhibition of fos mRNA levels after estrogen administration is produced by the glucocorticoid dexamethasone, but not by androgens or mineralocorticoids. Progesterone does not block the induction of c-jun or c-myc mRNA by estradiol. Uterine levels of c-fos mRNA observed after treatment with the phorbol ester phorbol 12-myristate 13-acetate are not decreased by a 3-h pretreatment with progesterone. Under the conditions of our experiments, progesterone does not decrease occupied levels of nuclear estrogen receptors in the uterus after estradiol administration. These findings are consistent with a mechanism in which progesterone inhibits transcriptional activation by the estrogen receptor at the level of the c-fos gene. PMID: 1375896 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 834: J Biol Chem. 1992 May 25;267(15):10551-60. Molecular characterization of the stretch-induced adaptation of cultured cardiac cells. An in vitro model of load-induced cardiac hypertrophy. Sadoshima J, Jahn L, Takahashi T, Kulik TJ, Izumo S. Molecular Medicine Unit, Beth Israel Hospital, Boston, Massachusetts 02215. Although it is a well-known fact that hemodynamic load is a major determinant of cardiac muscle mass and its phenotype, little is known as to how mechanical load is converted into intracellular signals of gene regulation. To address this question, we characterized the stretch-induced adaptation of cultured neonatal cardiocytes grown on a stretchable substrate in a serum-free medium. Static stretch (20%) of the cells was applied without cell injury. Stretch caused hypertrophy in myocytes and hyperplasia in non-myocytes. Stretch caused an induction of immediate-early genes such as c-fos, c-jun, c-myc, JE, and Egr-1, but not Hsp70. Immunostaining showed that the stretch-induced Fos protein localized in the nucleus of both myocytes and non-myocytes. Nuclear extracts from stretched myocytes contained DNA binding activity to the AP-1 and Egr-1 consensus sequences. In myocytes, the induction of immediate-early genes was followed by expression of "fetal" genes such as skeletal alpha-actin, atrial natriuretic factor, and beta-myosin heavy chain. DNA transfection experiments showed that the "stretch-response element" of the c-fos gene promoter is present within 356 base pairs of the 5'-flanking region, whereas that of the atrial natriuretic factor and the beta-myosin heavy chain genes is probably located outside of 3412 and 628 base pairs of the 5'-flanking region, respectively. These results demonstrate that the phenotype of stretched cardiocytes in this in vitro model closely mimics that of hemodynamic load-induced hypertrophy in vivo. This model seems to be a suitable system with which to dissect the molecular mechanisms of load-induced hypertrophy of cardiac muscle. PMID: 1534087 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 835: Int J Cancer. 1992 May 8;51(2):310-4. Harderian gland hyperplasia in c-mos transgenic mice. Heath LA, Rosenberg MP, Thorogood P, Speight P, Propst F. Ludwig Institute for Cancer Research, St. Mary's Hospital Medical School, London, UK. Transgenic mice carrying the mouse mos proto-oncogene linked to a retroviral LTR develop hyperplasia of the Harderian glands. Enlargement of the glands is evident as early as 18 weeks after birth, with glands reaching up to 10 times their normal weight. Approximately 65% of the cases of hyperplasia occur bilaterally, and the majority of mice affected are male (66%). Elevated levels of mos expression are found in all Harderian glands of mice from the affected transgenic line, but not in glands of normal mice or a non-affected transgenic line, indicating that hyperplasia is dependent on mos expression. Histological examination of the tissue reveals a general involvement of the entire gland epithelium in hyperplastic growth, with no evidence of focal or malignant tumours. These observations show that in addition to neu, myc, ras and ret transgenes, mos, a member of the protein-serine/threonine kinase family of oncogenes, can induce Harderian gland hyperplasia, thus revealing an unusual response by this organ to various classes of oncogenes. Analysis of fos, jun, myc and ets oncogene RNA in mos-induced hyperplastic Harderian glands shows that there are no consistent changes in the level of expression of these oncogenes, suggesting that mos acts via a mechanism other than by increasing the expression of these genes. PMID: 1568797 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 836: Cell Growth Differ. 1992 May;3(5):307-13. Regulation of EGR-1, c-jun, and c-myc gene expression by oncostatin M. Liu J, Clegg CH, Shoyab M. Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, Washington 98121. Oncostatin M (OM) is a cytokine that shares a structural and functional relationship with interleukin 6, leukemia-inhibitory factor, and granulocyte colony-stimulating factor. In this report, we tested for correlations between immediate-early gene expression and some of the cellular responses elicited by OM. We determined that OM stimulated a rapid and transient elevation of EGR-1, c-jun, and c-myc mRNA in human fibroblasts prior to their proliferation. OM also stimulated a transient induction of these genes in M1 leukemic cells that differentiated into nonreplicating, macrophage-like cells. The expression of c-myc, however, decreased significantly as the cells stopped dividing. Interestingly, OM had no detectable effect on the expression of EGR-1, c-jun, and c-myc during the cell cycle arrest of human A375 melanoma cells. Our results indicate that an early nuclear event associated with OM action is the regulation of immediate-early gene expression. We suggest that the transcription factors encoded by the EGR-1, c-jun, and c-myc genes are utilized in both cell proliferation and differentiation but are not part of the mechanism by which OM inhibits A375 cell growth. PMID: 1633113 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 837: J Rheumatol. 1992 May;19(5):693-9. c-myc mRNA expression in minor salivary glands of patients with Sjogren's syndrome. Skopouli FN, Kousvelari EE, Mertz P, Jaffe ES, Fox PC, Moutsopoulos HM. Department of Internal Medicine, School of Medicine, University of Ioannina, Greece. c-myc protooncogene is implicated in the pathogenesis of B cell lymphoid malignancies and high levels of c-myc mRNA expression are observed in activated blood mononuclear cells. Sjogren's syndrome (SS) is characterized by lymphocytic infiltrates of exocrine glands, remarkable B cell hyperreactivity and a strong predisposition to B cell neoplasia. In this study, c-myc protooncogene mRNA expression in 29 labial minor salivary gland biopsies from patients with primary SS and 15 controls was examined using in situ hybridization histochemistry. Two 40mer oligonucleotides from the 1st and the 2nd exon of the c-myc gene, labeled with 35S, were used as probes. To detect the origin of the cell hybridized with a c-myc probe, a combined immunochemistry in situ hybridization histochemistry technique was used. High c-myc mRNA expression was detected on acinar epithelial cells. c-myc did not correlate with c-fos and c-jun protein expression. Stronger c-myc mRNA expression was detected in labial salivary glands of patients with longer disease duration (p less than or equal to 0.002) and more intense T lymphocyte infiltrates (p less than 0.05) although these patients revealed no hypergammaglobulinemia. No correlation was observed between c-myc mRNA and B lymphocyte monoclonicity or lymphoma. In conclusion, strong c-myc mRNA expression was observed on epithelial cells of labial salivary glands from patients with primary SS. Our findings may indicate the presence of a reactivated virus hosted in these cells. PMID: 1613697 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 838: Blood. 1992 May 1;79(9):2404-14. Dissociation of nuclear events on p21 RAS transformation of FDC-P1 myeloid cells: c-jun/AP-1 expression versus c-myc transcription. Litz-Jackson S, Miller AH, Burgess GS, Boswell HS. Division of Hematology/Oncology, Indiana University School of Medicine, Indianapolis. We have previously reported transformation to growth factor-independent proliferation in the interleukin-3 (IL-3)-dependent cell line FDC-P1 by high-level expression of the valine 12 Harvey RAS oncogene, following from a nonautocrine mechanism. The present study was undertaken to examine nuclear tertiary messenger, transcriptional response gene expression to deduce the intracellular signaling pathways responsible for this autonomous proliferation. We confirmed other reports that transformed p21RAS-expressing cells constitutively express the transcription factor complex jun/AP-1, in this case resulting from the ongoing expression of the c-jun and c-fos genes in the absence of IL-3. However, the ongoing growth factor independent expression of c-myc by a transcriptional mechanism in FDC-P1 cells expressing p21 RAS cannot be explained by intracellular signaling in the jun/AP-1 (protein kinase C) pathway. This conclusion derives from the observation that c-jun expression mediated via protein kinase C activation with phorbol ester (12-0-tetra decanoylphorbol-13-acetate, TPA) treatment does not lead to c-myc expression in parent FDC-P1 cells. On the contrary, FDC-P1 cells stably transfected with a c-myc gene controlled under the influence of a metallothionein IIA promoter (containing the TPA-responsive element [TRE]) express the transfected MTIIA-c-myc and downregulate the endogenous c-myc in response to protein kinase C activation with TPA. Further, nuclear proteins derived from cells expressing p21 RAS, which bind specifically to the purified c-myc P2 promoter, are not competed in their binding to the motif-rich P2 element by AP-1 oligonucleotide. Therefore, expression of the Harvey RAS oncogene in FDC-P1 myeloid cells leads to at least two pathways of cytoplasmic signaling. One pathway involves protein kinase C and c-jun/AP-1, but another pathway that is protein kinase C-independent appears to mediate c-myc transcription. PMID: 1571552 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 839: J Cell Physiol. 1992 May;151(2):415-26. Induction of differentiation and c-jun expression in human leukemic cells by okadaic acid, an inhibitor of protein phosphatases. Adunyah SE, Unlap TM, Franklin CC, Kraft AS. Division of Hematology/Oncology, University of Alabama, Birmingham 35294. Okadaic acid, a protein phosphatase inhibitor, is a strong tumor promoter which activates protein phosphorylation. Because another activator of protein phosphorylation, phorbol esters, stimulates hematopoietic differentiation, we sought to determine whether okadaic acid could also induce the differentiation of the human leukemic cell line U937. Differentiation was assessed by measuring changes in the following: mRNA levels, cell growth, morphology, cell surface markers, and the ability to induce superoxide. We found that okadaic acid treatment of U937 cells induces immediate increases in total cellular levels of both c-jun and c-fos mRNAs. Nuclear run-on experiments demonstrate that initial increases are secondary to increases in transcription, whereas latter changes may be secondary to mRNA stabilization. Like phorbol esters, okadaic acid treatment also activates AP-1 enhancer activity and induces the phosphorylation of c-Jun protein. Approximately 6-12 hours after treatment with okadaic acid, mRNA levels of c-myc, p34cdc2, and p58GTA, two cell cycle regulated protein kinases, decrease. Okadaic acid inhibits the growth of U937 cells, induces changes in nuclear morphology, stimulates increases in Mac-1 and Leu 11 surface antigens, and induces these cells to produce superoxide. These changes, taken together, suggest that U937 cells have been induced by okadaic acid to differentiate towards a more mature cell type. PMID: 1315324 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 840: J Biol Chem. 1992 Apr 25;267(12):8222-9. Stable expression of HB24, a diverged human homeobox gene, in T lymphocytes induces genes involved in T cell activation and growth. Deguchi Y, Thevenin C, Kehrl JH. Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892. A diverged homeobox gene, HB24, which is known to be induced following lymphocyte activation, was introduced into Jurkat T cells under the control of a constitutive promoter. Stable transfectants of HB24 were established that expressed high levels of HB24 mRNA and possessed an altered phenotype suggestive of activated T cells. A number of genes known to be induced following T cell activation and associated with cell growth were increased in the transfectants, including c-fos, c-myc, c-myb, HLA-DR, lck, NF-kappa B, interleukin-2 and interleukin-2 receptor alpha (IL-2R alpha). Analysis of IL-2R alpha expression by transient transfection of IL-2R alpha promoter constructs into the HB24 transfectants revealed constitutive expression (about 60% of phytohemagglutinin- and phorbol ester-activated Jurkat cells) that was dependent on the kappa B site in the IL-2R alpha promoter. Furthermore, as a consequence of the increased HB24 mRNA levels, the Jurkat HB24 transfectants proliferated more rapidly than control cell lines. Thus, stable expression of HB24 confers an activation phenotype on a human T cell line, implicating this gene as an important transcriptional factor during T cell activation and growth. PMID: 1349016 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 841: Blood. 1992 Apr 15;79(8):2107-15. Inhibition of c-jun causes reversible proliferative arrest and withdrawal from the cell cycle. Smith MJ, Prochownik EV. Division of Hematology/Oncology, University of Michigan School of Medicine, Ann Arbor 48109. We studied the effect of c-jun depletion in Friend murine erythroleukemia (F-MEL) cells stably transfected with a plasmid that allowed for the glucocorticoid-mediated conditional expression of c-jun antisense sequences. The c-jun cDNA used for the construction of the vector was modified so as to prevent the nonspecific targeting of junB and junD transcripts. High level and rapid induction of c-jun antisense transcripts was achieved with as little as 10(-8) mol/L dexamethasone (DEX) and resulted in a 80% to 90% reduction in c-jun protein levels. The continuous exposure of the transfected cells to DEX inhibited growth by greater than 85% over a 5-day period, whereas DEX had no effect on the growth rate of control F-MEL cells. This proliferative block was associated with a reversible accumulation of cells with a 2n DNA content. When these cells were recultured in the absence of DEX, c-jun protein rapidly reappeared and the immediate early response genes egr-1, junB, and c-myc were transiently expressed. Thus, inhibition of c-jun protein causes logarithmically growing cells to leave the cell cycle and to enter a state closely resembling, if not identical to, G0. These results underscore the importance of c-jun in maintaining cellular proliferation and provide additional evidence for the participation of proto-oncogenes in cell cycle control. PMID: 1562737 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 842: Rinsho Ketsueki. 1992 Apr;33(4):467-72. [A case report of AML M0:CD7, 33 (+) AML M0 case initially presented with cervical lymphadenopathy] [Article in Japanese] Horiguchi J, Yamamura S, Nemoto T, Fujikawa T, Inaba S, Yamazaki Y, Yamada H. Department of Internal Medicine, Aoto Hospital, Jikei University, School of Medicine. A 59-year-old man was admitted because of generalized lymphadenopathy with fever and vomiting. His peripheral blood showed leukocytosis with a WBC of 93,500/microliters, and the bone marrow picture revealed a predominance of blast cells. The blasts were negative for peroxidase, alpha-naphthyl butyrate esterase and PAS, and had the phenotype of CD 7, 13 and 33 positive. A diagnosis of AML M0 was made, based on the criteria of the NCI-sponsored workshop in 1988. His initial status had been compromised by acute renal failure which necessitated hemodialysis. He responded partially to chemotherapy consisting of daunorubicin, cytarabine and prednisolone. However leukemia recurred and the patient suffered from various episodes of infection and died six months after admission. The Southern blotting showed the germ line configuration for TCR-beta chain and immunoglobulin heavy chain genes. No messenger RNA was detected for myeloperoxidase, c-myc and c-jun, while c-fms, c-fos and c-myb were expressed on Northern blotting. It is intriguing to detect c-fms and c-fos expression in these poorly differentiated leukemic cells. Publication Types: Case Reports PMID: 1602610 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 843: Carcinogenesis. 1992 Apr;13(4):601-4. Elevation of protooncogene messenger RNAs in estrogen-induced kidney tumors in the hamster. Liehr JG, Chiappetta C, Roy D, Stancel GM. Department of Pharmacology, University of Texas Medical Branch, Galveston 77550. Cellular oncogenes such as c-fos, c-jun and c-myc are expressed prior to estrogen-induced growth of normal target tissues such as rodent uterus. Transient increases in the levels of these genes are induced by the administration of estradiol and are followed by DNA replication. In this study, we examined the expression of these three oncogenes in estradiol-induced kidney tumors in Syrian hamsters in order to understand mechanistic aspects of hormonal carcinogenesis. Kidney tumors were induced in all male Syrian hamsters treated chronically with estradiol for 7 or 9 months, whereas neoplasms were not detected in animals treated for 5 months. mRNA levels of fos, myc and jun were elevated 15-, 4- and 6-fold respectively in kidney tumors of estradiol-treated hamsters (9 months) compared with age-matched untreated control kidneys. The expression of all three protooncogenes was also increased in the kidney tissue surrounding tumors, though there was no consistent pattern in the ratios of transcripts in the tumor and kidney tissues. After 7 months of estrogen treatment, kidney tumors also contained elevated amounts of c-fos, c-jun and c-myc transcripts at levels comparable with older tumors. In abdominal metastases of hamster kidney tumors, mRNA levels of fos, myc and jun were elevated 9-, 12- and 3-fold respectively over control levels. In kidneys of hamsters treated with estradiol for 5 months, in which tumors were not yet detected, the expression of protooncogenes was slightly increased. Ratios of c-fos, c-myc and c-jun in estrogen-treated (5 months) over control tissue were 1.4, 1.1 and 1.3 respectively. Overexpression of cellular oncogenes such as c-fos, c-jun and c-myc may have played a role in the induction and growth of kidney tumors by estradiol in hamsters. PMID: 1576713 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 844: J Biol Chem. 1992 Mar 25;267(9):6240-8. Interleukin-1-inducible tumor growth arrest is characterized by activation of cell type-specific "early" gene expression programs. Rangnekar VV, Waheed S, Rangnekar VM. Department of Medicine, University of Chicago, Illinois 60637. Human melanoma cells, A375-C6, were "committed" to growth arrest within a few hours of exposure to interleukin-1 (IL-1). Co-treatment with actinomycin D rescued the cells from the "commitment," suggesting that "early" gene activation events may be crucial for growth arrest. To understand the mechanism of IL-1 action, we are studying early genes whose expression is induced by the cytokine. Five early genes associated with IL-1 action in the melanoma cells were isolated by differential screening of a cDNA library, which was enriched for sequences representing IL-1 responsive genes (IRGs). Nucleotide sequencing identified four of the genes as gro-alpha, gro-beta, c-jun and nur77/NGF1-B/NAK1, respectively, while the fifth was judged as novel by GenBank search and designated IRG-9. None of the early genes was uniquely associated with the antiproliferative action of IL-1: other growth-inhibitory as well as growth-stimulatory signals induced these genes in diverse cell types. However, analysis of the induction patterns of the IRGs and other well known early genes revealed that IL-1 action in the melanoma cells is characterized by activation of a unique primary gene expression program. This program was defined by the magnitude and temporal pattern of induction of the five IRGs, feeble induction of c-fos, and lack of induction of Egr-1 and c-myc. We present evidence that this program is growth arrest-specific in the melanoma cells and that distinct cell type-specific programs are associated with IL-1 growth-regulatory actions in other tumor cells. Based on these data, we propose that early genes may play multifunctional roles in tumor growth control, but specificity for the growth arrest action of IL-1 is determined by the composite early gene induction program. PMID: 1372901 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 845: J Biol Chem. 1992 Mar 25;267(9):5921-6. Use of a multiple S1 nuclease protection assay to monitor changes in RNA levels for type 1 phosphatase and several proto-oncogenes in response to insulin. Thompson DB, Sommercorn J. Clinical Diabetes and Nutrition Section, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Phoenix, Arizona 85016. Changes in insulin-regulated gene expression occur in a time- and tissue-dependent fashion. To monitor these changes we have adapted the S1 nuclease protection assay to allow simultaneous estimation of multiple RNA species in a single sample by using synthetic oligonucleotides of various lengths as probes for specific RNA species, which can then be resolved by electrophoresis. The multiple S1 nuclease protection assay was used to assess the influence of insulin on the RNA concentrations of 12 different genes in human skeletal muscle. Estimates obtained by this assay were comparable with those obtained by Northern analysis. RNA levels for proto-oncogene c-src displayed a transient 4-fold increase, whereas RNA levels for type 1 protein phosphatase were suppressed by 50% during the same time period. RNAs corresponding to known insulin-responsive genes such as c-fos, c-myc, c-Ha-ras, and c-src displayed rapid and transient 2-4-fold increases between 30 and 60 min as detected by either Northern analysis or the multiple S1 nuclease protection assay. In addition, RNA levels for the insulin receptor, Glut-4, Glut-3, and c-jun were apparently unaffected by exposure of the cells to insulin. PMID: 1372896 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 846: Cell Signal. 1992 Mar;4(2):163-77. Inducible overexpression of human protein kinase C alpha in NIH 3T3 fibroblasts results in growth abnormalities. Finkenzeller G, Marme D, Hug H. University of Freiburg, Institute of Molecular Cell Biology, Germany. We have stably overexpressed the human protein kinase C alpha (hPKC alpha) in NIH 3T3 fibroblasts under the control of the interferon (IFN) type I inducible murine Mx promoter. These cells showed a 10-fold increase in the transcription of hPKC alpha mRNA after induction with interferon alpha. The increase in the amount and activity of protein kinase C (PKC)-protein in these cells was only about 3-fold after induction with interferon alpha. Compared to control cells which were transfected with the vector only, the NIH 3T3 fibroblasts transfected with the hPKC alpha cDNA showed already a slightly increased PKC-activity and amount of PKC-protein in the absence of interferon alpha. The hPKC alpha overexpressing cells had an altered, "transformed-like" morphology, which was reversed by staurosporine, an increased growth rate and a higher saturation density. The growth rate was further increased by treating the cells with interferon alpha. The hPKC alpha overexpressing cells were able to grow in soft agarose after treatment with phorbol ester such as TPA (12-O-tetradecanoylphorbol 13-acetate). After phorbol ester and interferon treatment a stronger expression of the protooncogene c-jun was detectable in the hPKC alpha overexpressing cells, whereas expression of c-fos and c-myc was not affected. Since these cells show a specific response pattern due to induced PKC alpha expression they might be useful as an assay system for the development of PKC isozyme-specific inhibitors and activators. PMID: 1616823 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 847: J Steroid Biochem Mol Biol. 1992 Mar;41(3-8):523-8. Transcriptional activation of jun and actin genes by estrogen during mitogenic stimulation of rat uterine cells. Cicatiello L, Ambrosino C, Coletta B, Scalona M, Sica V, Bresciani F, Weisz A. Istituto di Patologia generale e Oncologia, Prima Facolta di Medicina e Chirurgia, Universita di Napoli, Italy. Estrogens induce transcriptional activation of c-fos and c-myc proto-oncogenes during mitogenic stimulation of human, chicken, mouse and rat cells in vivo and in vitro. In this paper we show that 17 beta-estradiol injected into adult ovariectomized rats increases c-jun, jun-B and jun-D gene transcription in the uterus. Kinetics and amplitude of response are different for each gene, since c-jun is activated first, within 30 min after injection, followed by jun-D and jun-B, 60 and 90 min after injection, respectively. Maximal activation of jun-B marks a drop in transcription of all the jun genes. Furthermore, transcriptional activation by 17 beta-estradiol of the growth-regulated beta- and gamma-cytoskeletal actin genes is prevented by an inhibitor of protein synthesis, indicating that it is a secondary response to the hormone. These data support the hypothesis that during growth stimulation of target cells the estrogen receptor induces transcription of regulatory genes, triggering in this way a cascade of gene regulation events that results in progression through the cell cycle. PMID: 1373300 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 848: Am J Physiol. 1992 Mar;262(3 Pt 2):F389-96. Gene expression of growth-related proteins and ECM constituents in response to unilateral nephrectomy. Nakamura T, Ebihara I, Tomino Y, Koide H, Kikuchi K, Koiso K. Department of Medicine, Juntendo University School of Medicine, Tokyo, Japan. To identify the specific regulatory mechanism associated with the events following unilateral nephrectomy, we measured the levels of mRNA encoding for extracellular matrix (ECM) constituents, for protooncogenes, and for proliferating cell nuclear antigen (PCNA) in renal cortex and glomeruli. One hour after left nephrectomy, c-jun and c-fos mRNA levels in renal cortex increased rapidly and then decreased rapidly to the control level, whereas c-myc and PCNA mRNA levels showed a slower and more sustained increase, with a peak at 6 h after nephrectomy, and then decreased to the control level after 7 days. mRNA levels for basement membrane components including alpha 1-chain of type IV collagen, laminin B1 and B2 chains, and heparan sulfate proteoglycan core protein were significantly increased in renal cortex at 12 h after nephrectomy, whereas those for interstitial collagens including alpha 1-chains of type I and type III collagen were unchanged following nephrectomy. On the other hand, the glomerular expression of all genes examined in this study showed little change during the experimental period. These results suggest that the time course of mRNA expression of ECM constituents is different from that of growth-related proteins in renal cortex and that glomerular mRNA levels for these components may not be associated with renal hypertrophy in the early stages following unilateral nephrectomy. PMID: 1373035 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 849: Proc Natl Acad Sci U S A. 1992 Feb 15;89(4):1291-5. Myb and Ets proteins cooperate in transcriptional activation of the mim-1 promoter. Dudek H, Tantravahi RV, Rao VN, Reddy ES, Reddy EP. Wistar Institute of Anatomy and Biology, Philadelphia, PA 19104. In the generation of the acutely transforming avian retrovirus E26, both myb and ets genes have been transduced, leading to the production of a Gag-Myb-Ets fusion protein. This co-occurrence of v-myb and v-ets oncogenes suggests that the two might have a functional relationship. To look for such a relationship, we tested the transcriptional activation activity of Myb alone or with coexpressed Ets-1 or Ets-2. Using the promoter of the v-Myb-inducible mim-1 gene as a target, we found that full-length c-Myb gene products were poor activators of transcription, while an oncogenic (truncated) form of this protein was a strong trans-activator. However, coexpression of Ets-2 with full-length or truncated forms of Myb greatly increased trans-activation. Coexpression of Ets-1, Fos, Jun, or Myc with Myb did not increase trans-activation of the mim-1 promoter. The ability of Myb and Ets-2 to transactivate was cooperative, since Ets-2 alone gave little or no activation. Bacterially synthesized Ets-2 protein was found to bind specifically to the mim-1 promoter, suggesting that it may be a target for both Myb and Ets proteins. Thus, Myb and Ets proteins can cooperate in transcriptional activation, and their co-occurrence in the E26 virus may reflect a functional relationship between these two oncoproteins. Truncated forms of Myb may have a reduced need for cooperating factors such as Ets-2, and this might constitute an important mechanism associated with oncogenic activation. PMID: 1741383 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 850: FASEB J. 1992 Feb 1;6(3):919-24. Role of [Ca2+]i in induction of c-fos, c-jun, and c-myc mRNA in rat PTE after oxidative stress. Maki A, Berezesky IK, Fargnoli J, Holbrook NJ, Trump BF. Department of Pathology, University of Maryland School of Medicine, Baltimore 21201. Oxidative stress plays an important role in various types of cell injury and tumor promotion. Cells respond to oxidative stress in many ways including changes in membrane organization, ion movements, and altered gene expression, all of which contribute to the subsequent fate of affected cells. In this study, we investigated the expression of the proto-oncogenes c-fos, c-myc, and c-jun, which play a key role in proliferation and differentiation, using primary cultures of rat proximal tubular epithelium exposed to oxidative stress generated by the xanthine/xanthine oxidase system. This system generates superoxide and H2O2 in the extracellular space stimulating the release of active oxygen species from inflammatory cells. c-fos mRNA was expressed within 15 min, peaked at 30 min, and returned to constitutive levels by 3 h. c-jun mRNA began to rise after 30 min, peaked at 120 min, and remained above the constitutive levels up to 180 min. c-myc mRNA expression was less affected by the treatment, with levels increasing gradually over the 180 min period. The expression of c-fos was inhibited by superoxide dismutase but not by catalase and was super-induced by cycloheximide. H2O2 alone did not induce any c-fos mRNA in this system. Chelation of extracellular ionized calcium by EGTA or of intracellular ionized calcium by Quin 2/AM resulted in a marked decrease of c-fos expression. Two protein kinase C inhibitors, H-7 and staurosporine, partly diminished the expression of c-fos, whereas a third, 2-aminopurine, which has a broader spectrum of inhibiting protein kinases, almost completely abolished it. A poly ADP-ribosylation inhibitor, 3-aminobenzamide, had no effect on c-fos expression in this system. Our results show that oxidative stress provokes sequential expression of c-fos, c-jun, and c-myc, mRNA in this order. This c-fos expression appears to be largely controlled by calcium ion movement, which could include protein kinase C activation. Another protein kinase or kinases also appear to play an important role. PMID: 1740241 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 851: J Steroid Biochem Mol Biol. 1992 Feb;41(2):113-23. Estrogen regulates expression of the jun family of protooncogenes in the uterus. Chiappetta C, Kirkland JL, Loose-Mitchell DS, Murthy L, Stancel GM. Department of Pharmacology, University of Texas Medical School, Houston. Treatment of immature female rats with estradiol increases uterine levels of c-jun and jun-B mRNAs approx. 10-fold. This effect is specific for estrogenic steroids. The induction of jun transcripts is blocked by actinomycin D but not puromycin, suggesting that the hormonal effect is due at least in part to transcriptional activation. The hormone effect is rapid and peak levels of jun mRNAs are seen within 3 h after treatment. Inductions of jun and fos transcripts in the uterus by estradiol exhibit similar dose response curves (maximum responses at 4 micrograms/kg). Estradiol also elevates uterine levels of jun-D, and this induction is insensitive to puromycin. In vivo treatment with the phorbol ester TPA rapidly elevates uterine levels of fos, jun, and myc transcripts, indicating that expression of these protooncogenes is under non-estrogenic as well as estrogenic regulation in this target tissue. These results suggest that multiple members of the jun and fos protooncogene families may play a role in amplifying the uterine response to estrogens. PMID: 1543678 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 852: Exp Cell Res. 1992 Feb;198(2):305-14. Mitogenic growth factors regulate differentially early gene mRNA expression: a study on two clones of 3T3 fibroblasts. Janet T, Labourdette G, Sensenbrenner M, Pettmann B. Laboratoire de Neurobiologie Ontogenique, CNRS UPR 417, Strasbourg, France. The relationship between cell proliferation and mRNA levels of the immediate early genes c-fos, c-jun, and jun B has been investigated in two clones of 3T3 fibroblasts (D1-3T3 and N2-3T3) upon treatment with basic fibroblast growth factor (bFGF), thrombin, phorbol 12-myristate 13-acetate (PMA) and dibutyryl cyclic AMP (Bt2cAMP). The 3T3-derived clone D1-3T3 almost stops dividing upon serum deprivation, while the N2-3T3 clone does not. The proliferation of the two clones was stimulated by thrombin and PMA and inhibited by Bt2cAMP. Basic FGF stimulated the growth of D1-3T3 but partly inhibited that of N2-3T3 cells. In spite of variable mitogenic response, immediate early genes, c-fos, c-jun, jun B, and c-myc, were induced by the growth factors and by PMA in both cell clones. In our experimental conditions the early gene mRNAs were expressed independently; i.e., the expression of one protooncogene had no bearing on the expression of the other. The cell growth was not directly related to the expression of a particular protooncogene mRNA. Data are presented showing that early gene mRNA expression induced by bFGF or thrombin was not mediated by protein kinase C activation while thrombin-induced mitosis was. Basic FGF induced a part of c-jun mRNA expression, but not mitosis, through a pertussis toxin-sensitive mechanism. PMID: 1309504 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 853: Eur J Biochem. 1992 Jan 15;203(1-2):305-11. Baculovirus-mediated expression and characterisation of rat glycogen synthase kinase-3 beta, the mammalian homologue of the Drosophila melanogaster zeste-white 3sgg homeotic gene product. Hughes K, Pulverer BJ, Theocharous P, Woodgett JR. Ludwig Institute for Cancer Research, London, England. Molecular cloning of glycogen synthase kinase-3 (GSK-3) has demonstrated the existence of a novel form, termed GSK-3 beta, which is highly related to the well characterised GSK-3 alpha protein but derived from a distinct gene. The cDNA cloning also revealed a striking degree of amino acid identity between the two GSK-3 proteins, particularly the beta-form, and the zeste-white3/shaggy (zw3sgg) homeotic gene of Drosophila melanogaster. Abrogation of zw3sgg causes pleiotropic effects on fruitfly development affecting segmental organisation and cell fate determination. In view of the potential importance of GSK-3 beta in mammalian development and the lack of previous characterisation, we have expressed this protein in insect cells using recombinant baculovirus. A rapid purification scheme has been developed yielding essentially pure GSK-3 beta protein in three chromatographic steps. The protein has autonomous protein kinase activity and similar, but not identical, substrate preferences to GSK-3 alpha. Both GSK-3 proteins activate the MgATP-dependent form of protein phosphatase-1 and thus display 'factor A' activity. Since GSK-3 beta exhibits an identical site specificity to GSK-3 alpha with respect to phosphorylation of the proto-oncogene/transcription factors c-jun and c-myc, it is likely that the Drosophila zw3sgg protein kinase has a similar specificity for such transcription factors which may underlie the pleiotropic phenotypes observed when the Drosophila homologue is mutationally inactivated. PMID: 1346104 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 854: Am J Obstet Gynecol. 1992 Jan;166(1 Pt 1):206-12. Comment in: Am J Obstet Gynecol. 1992 Oct;167(4 Pt 1):1154. Both 17 beta-estradiol and tamoxifen induce c-fos messenger ribonucleic acid expression in human endometrial carcinoma grown in nude mice. Sakakibara K, Kan NC, Satyaswaroop PG. Department of Obstetrics and Gynecology, Milton S. Hershey Medical Center, Pennsylvania State University, Hershey 17033. We previously demonstrated the estrogen-like effects of tamoxifen on the acceleration of growth and increased progesterone receptor concentrations of human endometrial carcinomas grown in the nude mouse experimental model. In our current study the modulation of protooncogene expression by 17 beta-estradiol and tamoxifen in human endometrial carcinomas was investigated. The protooncogenes investigated in this study were c-fos, c-jun, c-myc, N-myc, HER-2/neu, c-erbB, c-fms, and c-Ha-ras. Among those we found that c-fos expression was induced by 17 beta-estradiol in the following 17 beta-estradiol-sensitive tumors: EnCa-101 and EnCa-X. The induction was apparent within 1 hour, reached peak level at 2 hours (16-fold), and remained constant up to 4 hours. The c-fos messenger ribonucleic acid returned to prestimulation level by 12 hours. Tamoxifen also stimulated c-fos expression, the expression pattern being similar to that of 17 beta-estradiol albeit of a lesser degree. The messenger ribonucleic acid transcripts for other protooncogenes tested did not show significant changes during hormonal manipulation. The induction of c-fos expression by tamoxifen is consistent with its estrogen-like effect on endometrial carcinoma growth. PMID: 1733196 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 855: Crit Rev Eukaryot Gene Expr. 1992;2(1):19-63. New concepts in steroid hormone action: transcription factors, proto-oncogenes, and the cascade model for steroid regulation of gene expression. Landers JP, Spelsberg TC. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN 55905. The past 6 years have seen a significant increase in our understanding of steroid receptor-mediated regulation of gene transcription. As a means to understanding receptors as transcriptional activators, several steroid receptor genes have been identified, cloned, and are now known to belong to a receptor superfamily. All steroid receptors possess conserved domains which confer various aspects of receptor function such as those that regulate DNA and steroid binding. In addition to the distinct intrinsic functions of these domains, nonreceptor proteins associated with the unactivated forms of the receptor appear to play a crucial role in receptor function. One such protein, hsp90, is speculated to stabilize the unactivated form of the receptor in the absence of hormone. Posttranslational modification also appears to be important in regulating the transcriptional activity of steroid receptors. Steroid receptors may exist in several phosphorylation states, each intimately linked to steps involved in the conversion of the newly synthesized protein to the steroid-bound form capable of transcriptional activation. The activated steroid-receptor complexes bind to chromatin "acceptor sites", the composition of which is presently under investigation. Steroid response elements, DNA-binding "acceptor" proteins in the nuclear matrix, and transcription factors and their elements appear to play a role in this binding and in the transcriptional control of genes exerted by steroid receptors. Different genes utilize different elements and factors for each particular steroid receptor species, reflecting the steroid- and gene-specific patterns of regulation of gene expression. In this review, a cascade model is used to explain how the receptor interaction with specific sites upstream of "regulatory (early) genes" may regulate a variety of steps in gene expression from transcription and mRNA half-life to protein processing. This model not only accounts for the paradoxical "lag phase" observed between steroid treatment and structural gene transcription, but also shows how steroid-regulated gene expression may occur an posttranscriptional steps. The rapid regulation of the nuclear proto-oncogenes, e.g., c-myc, c-fos, and c-jun, are used as examples of these early regulatory genes in steroid-regulated, receptor-mediated gene transcription. Publication Types: Review PMID: 1543897 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 856: Mol Carcinog. 1992;5(1):52-61. Alterations in mRNA levels for growth-related genes after transplantation into castrated hosts in oncogene-induced clonal mouse prostate carcinoma. Egawa S, Kadmon D, Miller GJ, Scardino PT, Thompson TC. Scott Department of Urology, Baylor College of Medicine, Houston, TX 77030. A clonal mouse prostate carcinoma was established by the introduction of the ras and myc oncogenes via the recombinant retrovirus Zipras/myc 9 using a mouse prostate reconstitution model system. A single-cell suspension derived from an early passage ras+myc-induced carcinoma was inoculated into the flanks of intact or castrated adult male C57BL/6 mice, and tumors were harvested 3 wk postinoculation for northern and Southern blotting. Tumor volume analysis showed that this carcinoma was not dependent on testicular androgens for growth. Southern blot analysis of virus-cell DNA junction fragments revealed that tumor cell populations recovered from both intact and castrated mice were progeny of the same virus-infected cell. Northern blotting showed that mRNA levels for the four growth-related genes transforming growth factor-beta 1 (TGF-beta 1), transforming growth factor-beta 3 (TGF-beta 3), tissue-type plasminogen activator (tPA), and c-myc were significantly elevated in clonal mouse prostate carcinomas grown in castrated hosts. In contrast, androgen receptor mRNA levels were significantly reduced under the same conditions. The response of TGF-beta 1, tPA, and c-myc mRNA levels in the carcinomas grown in castrated hosts was similar to that shown previously in normal rat ventral prostate. However, unlike normal rat ventral prostate after castration, increased numbers of apoptotic cells were not seen in the castrated group relative to the intact group at the time of analysis, indicating that the altered gene expression was not associated with cell death. In addition, testosterone-repressed prostate mRNA number 2 levels, shown previously to be elevated after castration in normal rat ventral prostate, were not increased in the androgen-deprived clonal mouse prostate carcinomas. Therefore, this early passage clonal ras+myc-induced prostate carcinoma demonstrates unique patterns of expression for a set of growth-related genes in an androgen-deprived environment. PMID: 1543541 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 857: Acta Obstet Gynecol Scand Suppl. 1992;155:19-24. Oncogene and growth factor expression in ovarian cancer. Kommoss F, Bauknecht T, Birmelin G, Kohler M, Tesch H, Pfleiderer A. Department of Gynaecology, Albert-Ludwig University, Freiburg, Germany. The varying tumor-biological behavior of ovarian carcinomas probably influences both their operability and response to chemotherapy, which are the most relevant prognostic factors. The phenotype of different ovarian carcinomas is obviously associated with an activation of the EGF/TGF-alpha signal pathway, including c-myc and c-jun expression. Analysis of EGF-R, TGF-alpha, c-myc and c-jun expression in 33 stage III/IV, and 2 stage I/II ovarian carcinomas with biochemical, molecular-chemical and immunohistochemical methods showed a correlation between the mRNA and protein levels of EGF-R and TGF-alpha for tumors with low or high expressing rates. However, the concentration of measurable free EGF-Rs seems to depend on the amount of TGF-alpha expression by the tumors. The EGF-R binding ligand TGF-alpha is produced by epithelial tumor cells; stromal cells are usually TGF-alpha-negative, as shown by immunohistochemistry. High expression rates of EGF-R. TGF-alpha and c-myc were detected in 6, 7, and 10 out of 35 ovarian carcinomas, respectively. C-jun mRNA was detected in 18/19 cases studied. Non-malignant tissues originating from myometrium or ovary expressed no (or only small amounts of) EGF-R or TGF-alpha mRNA, whereas a high c-myc expression was found in 1/7 normal myometria, and in 2/5 normal ovaries. There was no strong correlation between EGF-R/TGF-alpha and c-myc/c-jun expression.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1502888 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 858: Oncol Res. 1992;4(7):291-8. HL-60 cells isolated for resistance to vincristine are defective in 12-O-tetradecanoylphorbol-13-acetate induced differentiation and the formation of a functional AP-1 complex. Ma L, Krishnamachary N, Perbal B, Center MS. Division of Biology, Kansas State University, Manhattan 66506. HL-60 cells isolated for resistance to vincristine are multidrug resistant and defective in the cellular accumulation of drug. Further studies demonstrate that these cells are also highly defective in 12-O-tetradecanoylphorbol-13-acetate (TPA) induced differentiation to macrophages. Analysis of this system demonstrates that certain protooncogenes which may contribute to differentiation are expressed at similar levels in sensitive and resistant cells. Thus, treatment of cells with TPA results in a reduction in the levels of c-myb and c-myc mRNA, while the expression of c-fos, c-jun, and junB is greatly enhanced. Immunoprecipitation experiments also demonstrate a TPA induced increase in the c-jun protein in both sensitive and resistant cells. Gel mobility shift assays show that TPA induces AP-1 formation in sensitive cells, whereas in parallel experiments with the HL-60/Vinc isolate, AP-1 is essentially absent. It has been found, however, that in resistant cells which have reverted to drug sensitivity, the levels of TPA inducible AP-1 is essentially identical to that of sensitive cells. Revertant and sensitive cells differentiate at similar levels in the presence of TPA. These studies therefore demonstrate that HL-60/Vinc cells are defective in the TPA induction of a functional AP-1 complex and that this may account for the inability of these cells to differentiate to macrophages. The molecular basis of the finding that AP-1 is not formed in resistant cells remains to be determined. PMID: 1450490 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 859: Eur J Cancer. 1992;28A(10):1615-7. No correlation between ras, c-myc and c-jun proto-oncogene expression and prognosis in advanced carcinoma of cervix. Symonds RP, Habeshaw T, Paul J, Kerr DJ, Darling A, Burnett RA, Sotsiou F, Linardopoulos S, Spandidos DA. Beatson Oncology Centre, Western Infirmary, Glasgow, U.K. 55 patients suffering from stage III or IV carcinoma of cervix were treated with two pulses of neo-adjuvant chemotherapy prior to radical radiotherapy. 51% (26/51) had a partial response. The initial response to chemotherapy is associated with significantly better long-term survival. The 3-year survival of chemotherapy responders is 62% against 21% for non-responders (P = 0.009 log-rank test). To detect possible differences in oncogene expression in biopsy specimens taken from responding and non-responding patients, paraffin-fixed material was immunocytochemically stained for the expression of the protein products of ras, c-myc and c-jun proto-oncogenes. The frequency of oncogene expression was ras 80.4%, c-myc 45.1% and c-jun 39.2%. There was no statistically significant association between oncogene expression, time to local recurrence or development of metastases or survival. PMID: 1389474 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 860: Biol Cell. 1992;75(1):61-8. Cellular analysis of the kinetics of alpha-fetoprotein and nuclear oncogene activation in primary cultures of adult rat hepatocytes stimulated by epidermal growth factor. Tournier-Thurneyssen I, Feldmann G, Bernuau D. INSERM U327 Laboratoire de Biologie Cellulaire, Faculte de Medecine Xavier, Bichat, France. Alpha-fetoprotein (AFP), a fetal gene normally inactivated in quiescent adult hepatocytes, is re-expressed in hepatocytes during the proliferating response induced by liver regeneration in vivo, or epidermal growth factor (EGF) in vitro. The nuclear oncogenes c-jun, c-fos and c-myc are 'immediate early' genes also activated during the proliferative response of hepatocytes. Whether AFP gene activation is linked to oncogene activation is not known. As a first step in answering this question, we have analysed the cellular kinetics of nuclear oncogene and AFP gene activation in primary cultures of adult rat hepatocytes stimulated by EGF. Gene activation was evaluated at the mRNA level by dot blot and in situ hybridization, and at the protein level by immunoperoxidase. C-jun, c-fos and c-myc mRNA steady state levels in total cellular RNA were increased from 30 min-2 h after EGF stimulation. In situ hybridization analysis showed that transcripts of the three oncogenes increased in all hepatocytes after EGF stimulation. While unstimulated cultures did not immunostain for Fos and Myc proteins. Fos immunostaining was visible in the majority of hepatocyte nuclei 1 and 2 h after EGF addition, and Myc cytoplasmic immunostaining of the majority of hepatocytes was observed at 2 h of stimulation. AFP mRNA increased in total cellular RNA 2 and 4 h after EGF stimulation, with elevated in situ hybridization signal for AFP mRNA in all hepatocytes. No hepatocytes immunostained for AFP in unstimulated cultures, but a cytoplasmic labeling of 20-30% of the hepatocytes was observed 6 h after stimulation.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1381255 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 861: J Cardiovasc Pharmacol. 1992;20 Suppl 1:S37-40. The role of angiotensin II in vascular smooth muscle cell growth. Naftilan AJ. University of Alabama, Birmingham. One of the major consequences of hypertension is an increase in the thickness of the arterial medial smooth muscle cell layer. This has been shown in both large and medium size resistance vessels caused by smooth muscle cell hypertrophy. Both in vivo and in vitro data suggest that the vasoconstrictor peptide angiotensin II (Ang II) may play an important role in the development of the smooth muscle hypertrophy. We have demonstrated that Ang II, when added to quiescence cultures of vascular smooth muscle cells, results in the rapid induction of the early growth response genes c-fos, c-myc, and c-jun. This is due to new transcription as demonstrated by nuclear runoff transcription assay, but is not dependent on new protein synthesis, as it is not blocked by the addition of cycloheximide. The effect is due, however, to an increase in intracellular calcium, suggesting that any vasoconstrictor which results in an increase in intracellular calcium may act in this manner. Following the induction of the early growth response genes there is delayed induction of the platelet derived growth factor A-chain gene. Data from our laboratory and from that of others has shown in preliminary studies that blockade of either the Ang II-induced increases in c-fos or in the platelet-derived growth factor A-chain increases smooth muscle cell protein synthesis. This suggests that Ang II and other vasoconstrictors may play an important role in vascular smooth muscle growth, in hypertension and also in atherosclerosis and following balloon injury of the arterial wall.(ABSTRACT TRUNCATED AT 250 WORDS) Publication Types: Review Review, Tutorial PMID: 1380617 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 862: Brain Res Mol Brain Res. 1992 Jan;12(1-3):215-23. Changes in immediate early gene expression during postnatal development of cat cortex and cerebellum. McCormack MA, Rosen KM, Villa-Komaroff L, Mower GD. Department of Neurology, Children's Hospital, Boston, MA. Postnatal brain development involves interactions between extracellular signals and preprogrammed genetic events. Immediate early genes (IEGs) are a group of genes that are induced by extracellular signals and their protein products alter transcription by binding regulatory elements in other genes. Using Northern and slot blot analysis of total RNA isolated from visual cortex, frontal cortex, and cerebellum of cats, we have determined the postnatal development patterns of mRNA expression for 5 of these genes, c-fos, erg-1, c-jun, jun-B, and c-myc. Each gene had a distinct developmental pattern of mRNA expression, and for a given gene, these patterns were often different in different brain structures. These results suggest that temporal changes in the combinatorial interaction of different IEGs during early postnatal life are important for normal brain development. PMID: 1372068 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 863: Cancer Treat Res. 1992;63:301-11. Fos and Jun: inducible transcription factors regulating growth of normal and transformed cells. Holt J. Publication Types: Review Review, Tutorial PMID: 1363363 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 864: EMBO J. 1992 Jan;11(1):115-26. Autocrine growth induced by kinase type oncogenes in myeloid cells requires AP-1 and NF-M, a myeloid specific, C/EBP-like factor. Sterneck E, Muller C, Katz S, Leutz A. Zentrum fur Molekulare Biologie Heidelberg, University of Heidelberg, FRG. The nuclear oncogenes v-myc or v-myb specifically transform avian myeloid cells. In both cases, the transformed cells remain dependent on chicken myelomonocytic growth factor (cMGF). This factor dependence can be relieved by expression of kinase-type oncogenes such as v-mil or v-erbB, leading to expression of cMGF and autocrine growth stimulation. In erythroid cells the same kinase-type oncogenes cause transformation but do not induce cMGF expression. Here we investigated the molecular mechanisms of the observed lineage specific oncogene collaboration. We found that kinase-type oncogenes and TPA activate the cMGF promoter via AP-1 like transcription factors. The activation of the cMGF promoter is, however, strictly dependent on the binding of nuclear proteins to both halves of an inverted repeat adjacent to the AP-1 binding site. These proteins are related to C/EBP. They are expressed exclusively in myeloid cells and were therefore termed NF-M. Our results indicate that the lineage specific cooperation of kinase type oncogenes with v-myb or v-myc in leukemia formation is based on the concerted action of AP-1 and NF-M on the cMGF promoter. PMID: 1346759 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 865: J Membr Biol. 1992 Jan;125(2):163-70. Serum independence of low K+ induction of Na,K-ATPase: possible role of c-fos. Cayanis E, Russo JJ, Wu YS, Edelman IS. Center for Reproductive Sciences, College of Physicians and Surgeons, Columbia University, New York, New York 10032. Cultured ARL15 cells respond to abnormally low extracellular K+ concentrations by increasing the abundance of Na,K-ATPase (the Na/K pump). This response is preceded by significant increases in the mRNAs of the alpha 1 and beta 1 subunits of this enzyme, implying transcriptional or post-transcriptional regulation in the response. The present study concerned the possible participation of serum factors in low K+ induction of Na,K-ATPase. In normal K+ (4.5 mM) or low K+ (0.68 mM) the presence of 10% calf serum had no effect on Na,K-ATPase activity. The serum independence of the response to low K+ raised the possibility that low K+ may itself elicit a "growth" response. Accordingly, the effect of low K+ on mRNA abundances of four proto-oncogenes (c-fos, c-myc, c-jun and c-ski) was evaluated in the early phase of the response by quantitative Northern blot analysis. The mRNA for c-fos was transiently elevated by low K+, with a peak at 30 min. In contrast, low K+ had no measurable effect on the abundances of c-myc, c-jun and c-ski, for up to 2 hr of exposure. The early elevation of c-fos mRNA makes it a candidate mediator in this signal-transduction pathway. Induction of c-fos mRNA by the phorbol ester, PMA, or by dioctanoyl glycerol, however, had no effect on Na,K-ATPase activity. These results indicate that an increase in c-fos mRNA alone is not sufficient to induce Na,K-ATPase. Whether induction of c-fos is necessary for the response to low K+ remains to be determined in future studies. PMID: 1313115 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 866: J Virol. 1992 Jan;66(1):53-61. Polyomavirus large and small T antigens cooperate in induction of the S phase in serum-starved 3T3 mouse fibroblasts. Ogris E, Mudrak I, Wintersberger E. Institut fur Molekularbiologie, Universitat Wien, Austria. The induction of an S phase in the host cell is a prerequisite for the lytic replication cycle of polyomavirus. This function was attributed to proteins coded for by the early region of the viral DNA, the T antigens. A consideration of the role of the T antigens in the initiation of a mitogenic response of the host cell has to take into account the recent discovery that virus adsorption is sufficient to induce the synthesis of proteins which are known to appear early after quiescent cells are stimulated by the addition of serum, namely fos, jun, and myc (J. Zullo, C.D. Stiles, and R.L. Garcea, Proc. Natl. Acad. Sci. USA 84:1210-1214, 1987; G. M. Glenn and W. Eckhart, J. Virol. 64:2193-2201, 1990). This induction is followed by an initiation of DNA synthesis. It is therefore important to dissociate the effects of the T antigens on the host cell from those of virus adsorption. To do so, we used dexamethasone-regulated versions of the large and small T antigens of polyomavirus stably integrated into the genome of Swiss 3T3 cells to study their function in S-phase induction. When the production of the large or small T antigen in serum-starved 3T3 mouse fibroblasts was activated, only a small fraction of cells was able to leave G0/G1 despite the synthesis of considerable amounts of the respective T antigen. Activation of both T antigens within the same cell, on the other hand, resulted in S-phase induction in a notable percentage of cells, suggesting that the two proteins cooperate in this activity. Polyomavirus T antigens appear to bypass the pathway of growth regulation involving the activation of c-fos. These results are discussed in relation to other known functions of the two virally coded proteins. PMID: 1309261 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 867: Am J Respir Cell Mol Biol. 1991 Dec;5(6):548-55. Regulation of bovine bronchial epithelial cell proliferation and proto-oncogene expression by growth factors. Takizawa H, Beckmann JD, Yoshida M, Romberger D, Rennard SI. Department of Internal Medicine, University of Nebraska Medical Center, Omaha 68198-2465. The proto-oncogenes are thought to play important roles in the regulation of cellular growth and differentiation. In order to evaluate the role of proto-oncogenes in the regulation of growth of bronchial epithelial cells, we studied steady-state levels of fos, jun, and myc transcripts in response to fetal calf serum, bovine pituitary extract, and insulin. Extensively quiescent populations of bovine bronchial epithelial cells in growth factor-free medium were stimulated to divide by each of these three additives. We observed rapid but transient increases of fos, jun, and myc expression in association with such growth stimulation. There were no changes in tubulin mRNA levels over the same time periods. Other "growth factors" (epidermal growth factor, hydrocortisone, epinephrine, triiodothyronine, and transferrin) were also studied and did not affect either cell growth or expression of fos, jun, or myc. We further examined the effect of transforming growth factor-beta1 (TGF-beta1) on the above stimulatory effects. TGF-beta1 consistently inhibited the growth induced by fetal calf serum, bovine pituitary extract, or insulin and, interestingly, reduced proto-oncogene myc mRNA level without altering that of fos and jun. In conclusion, proto-oncogenes fos, jun, and myc appear to play a role in the regulation of growth response in bovine bronchial epithelial cells. It is also possible that TGF-beta1 exerts its growth inhibitory effect, at least in part, through the processes that involve the regulation of proto-oncogene myc transcription in these cells. PMID: 1958382 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 868: Leukemia. 1991 Dec;5(12):1099-109. Interleukin-3 dependent mitogenesis in murine cells involves a predominant non-protein kinase C (pKC) dependent pathway for c-myc transcription. Role of a myc expression vector in rescuing pKC dependent mitogenesis. Nahreini TS, Litz-Jackson S, Burgess GS, Helvering LM, Manolagas SC, Boswell HS. Department of Medicine, Indiana University School of Medicine, Indianapolis. The signaling pathways used by interleukin-3 (IL-3) and by active phorbol ester (12-0-tetradecanoyl phorbol-13-acetate, TPA) to stimulate mitogenesis in the growth factor dependent myeloid cell line FDC-P1 were studied by 'reporter' analysis of nuclear proto-oncogene expression. These studies revealed that IL-3 strongly stimulated c-myc expression by a transcriptional mechanism but IL-3 poorly stimulated c-jun expression, a measure of protein kinase C dependent signals. On the other hand, the protein kinase C agonist, TPA, strongly activated c-jun expression but poorly promoted expression (transcription) of c-myc in FDC-P1. These findings appeared to correlate with the poor mitogenic capacity of TPA for FDC-P1. However, stable transfection of FDC-P1 with a c-myc expression vector driven by a human methallothionein IIA promoter containing the TPA responsive element (TRE), led to a cell clone, FDMT myc.A1, in which TPA mediated selective transcription of the transfected TRE driven c-myc vector and down-regulated expression of the endogenous c-myc gene. IL-3 selectively failed to stimulate expression of the TRE driven c-myc vector in FDMT myc.A1. Augmented TPA dependent vector derived c-myc expression was accompanied by enhanced mitogenesis of the cell line FDMT myc.A1 compared with FDC-P1. In addition, TPA mediated expression of the transfected c-myc gene in FDMT myc.A1 was accompanied by augmented transcription of c-jun and c-fos in response to TPA. These studies show the importance of a non-protein kinase C dependent pathway for IL-3 mediated c-myc transcription. However, these studies reveal that protein kinase C mediated pathways can be promitogenic, especially when complemented by unregulated c-myc expression (in this case driven by an alternative, TRE containing promoter). PMID: 1774959 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 869: Am J Physiol. 1991 Dec;261(6 Pt 1):C994-1000. Production of transforming growth factor-alpha by normal rat small intestine. Barnard JA, Polk WH, Moses HL, Coffey RJ. Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2576. Transforming growth factor-alpha (TGF-alpha) and epidermal growth factor (EGF) are similar in structure and biological activity. In the present study, the distributions of TGF-alpha mRNA, TGF-alpha immunoreactivity, and TGF-alpha-EGF receptor mRNA were examined in epithelial and nonepithelial compartments of the jejunum, and the effect of TGF-alpha on growth of a jejunal crypt cell line (IEC-6) was determined. Epithelial cells eluted from the rat jejunal cryptvillus axis expressed TGF-alpha mRNA at twofold higher levels in the villus tip than in the crypt and EGF receptor mRNA at sevenfold higher levels in the villus tip. Expression of these two mRNA transcripts in the subepithelium was low. Immunohistochemical staining showed TGF-alpha immunoreactivity predominantly in the epithelium and muscularis. Immunostaining of villus cells was uniform, whereas crypt cells did not stain. IEC-6 cells bound 125I-EGF to a single class of high-affinity (dissociation constant = 833 pM) receptors. EGF and TGF-alpha (10 ng/ml) only modestly stimulated IEC-6 cell growth in the presence of 5% serum but increased expression of the protooncogenes c-jun and c-myc threefold over control cells. These findings suggest that, among the potential physiological roles for TGF-alpha produced by the jejunal epithelium, promotion of cell migration and modulation of fluid and electrolyte transport may be as relatively important as stimulation of cell proliferation. PMID: 1767826 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 870: Cell Growth Differ. 1991 Dec;2(12):645-52. Selective induction of c-jun and jun-B but not c-fos or c-myc during mitogenesis in SV40-transformed cells at the predifferentiation growth arrest state. Wang H, Wang JY, Johnson LR, Scott RE. Department of Pathology, University of Tennessee Medical Center, Memphis. Insulin has recently been reported to function as a complete mitogen for SV40 large T antigen-transformed 3T3 T-cells, designated CSV3-1, but not for nontransformed 3T3 T-cells (H. Wang and R. E. Scott, J. Cell. Physiol., 147: 102-110, 1991). It is now reported that sodium orthovanadate mimics this effect of insulin. For example, when exposed to 1-5 microM vanadate, most predifferentiation growth-arrested CSV3-1 cells undergo DNA synthesis within 24 h, but neither vanadate nor insulin induces mitogenesis in nontransformed 3T3 T-cells. To investigate the possible mechanisms by which mitogenesis is induced in CSV3-1 cells, the effects of insulin and vanadate on the expression of growth-related genes were examined. Whereas insulin and vanadate had no effect on the expression of c-fos, c-myc, c-jun, jun-B, or ornithine decarboxylase activity in nontransformed 3T3 T-cells, insulin and vanadate showed different effects on the expression of these genes in CSV3-1 cells. Insulin induced a rapid and transient accumulation of c-fos mRNA followed by induction of c-myc, c-jun, jun-B, and ornithine decarboxylase. In contrast, vanadate induced the expression of c-jun, jun-B, and ornithine decarboxylase without inducing c-fos and c-myc. These observations suggest that SV40 large T antigen may play an important role in insulin- and vanadate-induced mitogenesis and that insulin and vanadate may mediate their mitogenic effects by different signal transduction pathways. PMID: 1667087 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 871: Biokhimiia. 1991 Nov;56(11):1960-8. [Regulation of Ca(2+)-dsRNA for proliferation and terminal differentiation processes of human fibroblasts and HeLa cells] [Article in Russian] Zakharian RA. Ca2+ complexes of dsRNA, poly(dA) and poly(dT) of yeast low molecular weight RNA produce a pronounced mitogenic effect on human fibroblasts at early stages of fibroblast proliferation in culture. At later stages of cell cultivation Ca(2+)-dsRNA stimulates terminal differentiation by inducing the synthesis of proteins characteristic of the postmitotic population of human fibroblasts undergoing terminal differentiation. Ca(2+)-dsRNA produces a stimulating effect on c-fos and c-jun gene transcription in fibroblasts and HeLa-S-3. PMID: 1805979 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 872: Oncogene. 1991 Nov;6(11):2129-35. v-erbA oncogene abrogates growth inhibition of chicken embryo fibroblasts induced by retinoic acid. Desbois C, Pain B, Guilhot C, Benchaibi M, Ffrench M, Ghysdael J, Madjar JJ, Samarut J. Immuno-Virologie Moleculaire et Cellulaire, Universite Claude Bernard Lyon-1/CNRS UMR 30, Faculte de Medecine Alexis Carrel, Lyon, France. Retinoic acid inhibits chicken embryo fibroblast (CEF) proliferation by altering the G1 phase of the cell cycle with induction of a strong increase in the generation time. This growth-inhibitory response to retinoic acid is abrogated by expression of the v-erbA oncogene, suggesting an interference between retinoic acid receptors and the v-ErbA oncoprotein. Moreover, CEF expressing either the v-src, v-jun or v-fos oncogenes are also insensitive to retinoic acid treatment. In contrast, CEF expressing either the v-myc, v-myb-ets, v-mil, v-sea or v-erbB oncogenes are still sensitive to retinoic acid. These data strongly suggest functional interferences between the retinoic acid receptors and the AP-1 transcription factor complex in the control of expression of genes involved in CEF proliferation. PMID: 1682867 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 873: Br J Haematol. 1991 Oct;79 Suppl 1:14-6. Possible mechanism of action of interferon alpha in chronic B-cell malignancies. Heslop HE, Brenner MK, Ganeshaguru K, Hoffbrand AV. Department of Hematology/Oncology, St Jude Children's Research Hospital, Memphis, Tennessee. Recent evidence suggests that tumour necrosis factor alpha (TNF) is an autocrine growth factor for the chronic B-cell malignancies hairy cell leukaemia (HCL) and some cases of B-chronic lymphocytic leukaemia (B-CLL). Incubation with TNF in vitro has been shown to increase viability, DNA synthesis and the expression of the protooncogenes myc, fos and jun in the tumour cells from these patients. TNF in vitro also increases expression of TNF-mRNA, suggesting the existence of an autocrine growth loop for TNF in these cells. Current experiments are compatible with the hypothesis that interferon alpha (IFN) interferes with this autocrine growth loop in HCL and B-CLL by stimulating degradation of messenger RNAs (mRNAs) for a number of cytokines including that of TNF. This RNA degradation may be mediated through induction of the enzyme 2,5 oligo-A synthetase with consequent increased synthesis of 2,5 oligo-A which is known to stimulate the activity of a latent ribonuclease capable of degrading cytokine mRNAs. Circulating tumour-derived TNF may also contribute to the pancytopenia in HCL and B-CLL. Whether cytokine autocrine growth loops are important in other B-cell malignancies, e.g. myeloma and non-Hodgkin's lymphoma, and subject to IFN-stimulated breakdown needs further study. Publication Types: Review Review, Tutorial PMID: 1931702 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 874: Oncogene. 1991 Oct;6(10):1767-73. v-jun oncogene prevents terminal differentiation and suppresses muscle-specific gene expression in ASV-17-infected muscle cells. Grossi M, Calconi A, Tato F. Dipartimento di Biologia Cellulare e dello Sviluppo, Universita di Roma La Sapienza, Italy. Infection of replicating quail myoblasts with avian sarcoma virus 17 (ASV-17) results in the inhibition of terminal differentiation into multinucleated myotubes and in the acquisition of anchorage-independent proliferation. Expression of v-jun, the ASV-17 oncogene, concomitantly leads to the accumulation of the gag-jun polyprotein P65 in the nucleus and to the lack of expression of typical differentiation-specific genes such as myosin heavy chain (MHC) and alpha-actinin. Surprisingly, expression of desmin, the muscle-specific subunit of intermediate filaments, is conserved in ASV-17-transformed myoblasts. Analysis of clonal strains of transformed myoblasts suggests that (i) suppression of morphological and biochemical differentiation depends on the absence of muscle-specific gene transcripts; (ii) inhibition of muscle differentiation by v-jun does not depend on the transcriptional silencing of MyoD, a muscle-specific regulatory gene; (iii) expression of desmin is compatible with proliferation of ASV-17-transformed cells and is independent of v-jun and MyoD levels of expression. The present data suggest that nuclear localization of v-jun prevents terminal differentiation in myoblasts and selectively down-regulates muscle-specific genes in terminally differentiated myotubes. In this respect, the behaviour of v-jun is quite different from that of v-myc, thus suggesting that these two oncogenes, although both encoding nuclear proteins, may have different mechanisms of action. PMID: 1923502 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 875: Mol Cell Biol. 1991 Oct;11(10):4952-8. Early gene responses to transforming growth factor-beta in cells lacking growth-suppressive RB function. Zentella A, Weis FM, Ralph DA, Laiho M, Massague J. Howard Hughes Medical Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021. The growth-suppressive function of the retinoblastoma susceptibility gene product, RB, has been implicated in the mediation of growth inhibition and negative regulation of certain proliferation related genes by transforming growth factor-beta 1 (TGF-beta 1). Early gene responses to TGF-beta 1 were examined in order to determine their dependence on the cell cycle and on the growth-suppressive function of RB. TGF-beta 1, which rapidly elevates the steady-state level of junB and PAI-1 mRNAs and decreases that of c-myc mRNA, induces these responses in S-phase populations of Mv1Lu lung epithelial cells containing RB in a phosphorylated state. Since in this state RB is presumed to lack growth-suppressive activity, the response to TGF-beta 1 was also examined in DU145 human prostate carcinoma cells whose mutant RB product lacks growth-suppressive function. In these cells, TGF-beta 1 also decreases c-myc expression at the transcription initiation level. These results suggests that the c-myc, junB, and PAI-1 responses to TGF-beta 1 are not restricted to the G1 phase of the cell cycle and that down-regulation of c-myc expression by TGF-beta 1 can occur through a mechanism independent from the growth-suppressive function of RB. PMID: 1922028 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 876: Exp Cell Res. 1991 Oct;196(2):279-86. Dimethyl sulfoxide inhibits the expression of early growth-response genes and arrests fibroblasts at quiescence. Srinivas S, Sironmani TA, Shanmugam G. Cancer Biology Division, School of Biological Sciences, Madurai Kamaraj University, India. We have previously shown that dimethyl sulfoxide (DMSO) treatment of mouse embryo fibroblasts (MEF) at the early hours of mitogenic stimuli resulted in the inhibition of DNA and protein synthesis; delayed treatment of serum-stimulated cells with DMSO had little effect on the synthesis of these macromolecules. Here, we demonstrate the specific inhibition of expression of early growth response genes by DMSO in serum-stimulated MEF. The expression of interleukin 6, and of oncogenes c-myc and c-fos were inhibited when the cells were treated with 2% DMSO from the beginning of serum-stimulated growth but not after 3 h of mitogenic stimuli. Although the actin gene is an early serum-response gene, its expression was not affected by DMSO. The synthesis of another serum-induced protein, the plasminogen activator inhibitor-1 was blocked during concurrent and delayed (after 3 h of stimulation) treatment of serum-stimulated fibroblasts with DMSO. The expression of glyceraldehyde-3-phosphate dehydrogenase gene was not affected by DMSO. These results indicate that the expression of non-growth-related genes are either not affected or affected nonspecifically both at early and late stages of serum-induced growth of mouse embryo fibroblasts. The serum-induced expression of c-fos gene was abolished by DMSO treatment of MEF while the phorbol 12-myristate 13-acetate-induced expression of fos gene was not, indicating that the PMA signaling pathway was refractory to DMSO. Treatment of cells with medium containing 2% DMSO for 24-48 h prevents them from progression into cell cycle by preventing the expression of genes involved in G0-G1 transition of quiescent cells. PMID: 1909967 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 877: Biochemistry. 1991 Oct 1;30(39):9523-30. Enrichment of a second class of native acceptor sites for the avian oviduct progesterone receptor as intact chromatin fragments. Horton M, Landers JP, Subramaniam M, Goldberger A, Toyoda H, Gosse B, Spelsberg TC. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905. Several classes of specific progesterone receptor (PR) nuclear binding sites (acceptor sites) have previously been identified in avian oviduct chromatin on the basis of different binding affinities. Recently, two classes of acceptor proteins (AP) that are associated with these binding sites in the avian oviduct have been identified. These APs were termed receptor binding factors (RBF-1 and -2), and one (RBF-1) has been purified [Schuchard et al. (1991) Biochemistry 30, 4535-4542]. The RBF-1 is associated with the highest affinity class of sites in the intact chromatin, and the RBF-2 is associated with the second highest affinity class of sites. The PR binding sites and their associated RBF-2 protein remain with the residual chromatin fraction following extraction by 4 M Gdn-HCl. This Gdn-HCl-treated chromatin has been termed nucleoacidic protein (NAP). This paper describes the 200-fold enrichment of the native RBF-2 class of PR acceptor sites beginning with the DNase I digestion of NAP to obtain DNase-resistant fragment (NAPf) containing approximately 150 bp of DNA. The PR binding sites are further enriched by high-performance or fast protein liquid chromatography and chromatofocusing. Anti-RBF-1/RBF-2 protein antibodies identify antigens that coelute with the PR binding activity. Hybridization analysis of the DNAf from the enriched NAPf demonstrates sequence homologies with the nuclear matrix DNA as well as with genomic sequences of the rapid steroid responding nuclear protooncogenes c-myc and c-jun. However, comparative analyses of the whole genomic DNA with the nuclear matrix DNA indicate that the RBF-2 (NAPf) is largely nonnuclear matrix.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1892851 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 878: Biochemistry. 1991 Oct 1;30(39):9516-22. Nuclear matrix localization and specific matrix DNA binding by receptor binding factor 1 of the avian oviduct progesterone receptor. Schuchard M, Subramaniam M, Ruesink T, Spelsberg TC. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905. A chromatin acceptor protein for the avian oviduct progesterone receptor (PR), termed receptor binding factor 1 (RBF-1), has recently been shown to (1) be a component of the nuclear binding sites (acceptor sites) for PR and (2) generate high-affinity binding sites (termed the RBF-1 class of sites) on avian genomic DNA [Schuchard et al. (1991) Biochemistry 30, 4535-4542]. A second class of sites and its associated protein (termed RBF-2) were also identified. This paper demonstrates that RBF-1 and also the PR nuclear binding sites are localized in the oviduct nuclear matrix. RBF-1 is found in abundance in the nuclear matrix of liver but only in traces in the nuclear matrix of spleen. Extraction of the nuclear matrix with 4.0 M Gdn-HCl results in the complete removal of RBF-1 as occurs with whole chromatin. Interestingly, a second class of specific PR binding, termed RBF-2, remains on the nuclear matrix after the removal of all RBF-1. Southern blot analysis indicates that the nuclear matrix DNA contains sequences homologous with the 5'-flanking domains of the rapidly steroid regulated c-myc and c-jun protooncogenes and the beta-actin gene, but not genomic sequences of the late sex steroid regulated gene, ovalbumin, or the alpha-actin gene. A specific, small region in the 5'-flanking domain of the c-myc gene appears to be associated with the nuclear matrix. Southwestern blot analysis using partially purified RBF-1 shows a marked affinity and specificity of the RBF-1 for the nuclear matrix DNA.(ABSTRACT TRUNCATED AT 250 WORDS) PMID: 1892850 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 879: Rinsho Ketsueki. 1991 Sep;32(9):986-90. [Altered expression of protooncogenes during clinical course in an AML case transformed from MDS] [Article in Japanese] Fujikawa T, Horiguchi J, Iizuka K, Nemoto T, Iwase S, Yamamura S, Inaba S, Yamazaki Y, Sano S, Yamada H. Department of Internal Medicine, Aoto Hospital, Jikei University School of Medicine. The changes of expression of oncogenes in the mononuclear cells of MDS case was studied during his clinical course, in series. His bone marrow was considered to maintain its function partly in initial stage, since both peripheral blood and bone marrow responded to clinical episodes. However, his hematopoietic function was gradually impaired with the disease evolution to AML. We examined the expression of four oncogenes in the mononuclear cells of his three clinical stages, early RAEB-t, RAEB-t and AML, to study the cause of transformation from MDS to AML. Early RAEB-t cells expressed all oncogenes studied other than c-myb, while only c-myc was weakly observed in RAEB-t. AML cells expressed c-myc, c-jun and c-myb, except for c-fms. The expression of c-fms and c-jun of early RAEB-t was considered to reflect the monocytosis induced by infections, and the expressions of c-myb and c-myc of AML cells were regarded as one of malignant signs of tumor transformation. These findings suggest that the evolutional transformation of MDS to AML was affected by the altered expression of oncogenes. Publication Types: Case Reports PMID: 1942545 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 880: Oncogene. 1991 Sep;6(9):1531-7. Imbalanced expression of cellular nuclear oncogenes caused by v-sis/PDGF-2. Sonobe MH, Bravo R, Armelin MS. Departamento de Bioquimica, Universidade de Sao Paulo, Brazil. Two murine cell lines that overexpress v-sis/PDGF-2 were used to study the mechanism of cell transformation by SSV (simian sarcoma virus). In contrast to the parental cells that are phenotypically normal and serum-dependent for growth, v-sis-overexpressing cells grow in PDGF-free plasma medium, are unable to enter the G0 state and are highly tumorigenic. Analysis of the expression of some growth factor-induced early response genes in v-sis-overexpressing cells revealed: (a) high and constitutive c-myc mRNA levels in SSV-NRK cells; (b) unaltered levels of fra-1, fos B, jun B and krox 20 transcripts; (c) high and constitutive FOS staining due to c-FOS and FOS-related protein(s); (d) constitutive c-JUN and higher JUN D expression. These results are compatible with a model in which endogenous production of v-sis/PDGF-2 leads to deregulated expression of key cellular transregulators that, in turn, alter the cells' transcriptional program leading to the transformed state and malignancy. PMID: 1923519 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 881: Lab Invest. 1991 Sep;65(3):324-33. Temporal and spatial patterns of proto-oncogene expression at early stages of toxic liver injury in the rat. Herbst H, Milani S, Schuppan D, Stein H. Institute of Pathology, Klinikum Steglitz, Free University of Berlin, Germany. Sequential and transient expression of c-fos, c-jun, c-myc, c-Ha-ras and c-Ki-ras proto-oncogene RNA transcripts with zonal heterogeneity was demonstrated in virtually all hepatocytes of adult rat liver by in situ hybridization with single-stranded, [35S]-labeled cRNA probes at various time points after intraperitoneal administration of a single dose of carbon tetrachloride (CCl4). After a brief interval, elevated RNA levels of these genes were also observed in nonparenchymal cells. A second phase of proto-oncogene expression was characterized by high RNA levels in only a fraction of parenchymal cells with preference of mediolobular areas. Distribution and number of these cells were comparable tl hepatocytes expressing the proliferation-associated nuclear antigen Ki-67 72 hours after toxic injury. Oncogene expression in the nonparenchymal compartment correlated with distinct morphologic changes preceding type I procollagen gene expression by desmin-positive perisinusoidal cells, accumulating together with numerous c-fms expressing cells in the areas of hepatocellular necrosis. We conclude that zonal hepatic destruction by carbon tetrachloride induces proto-oncogene expression with distinct temporal and spatial patterns initiated by the most severely damaged hepatocytes. Proto-oncogene products thus represent valuable markers of cellular activation preceding and accompanying various aspects of tissue repair reactions. PMID: 1890812 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 882: Brain Res Mol Brain Res. 1991 Sep;11(2):115-24. Differential expression of immediate early genes in the hippocampus in the kindling model of epilepsy. Simonato M, Hosford DA, Labiner DM, Shin C, Mansbach HH, McNamara JO. Division of Neurology, Duke University, Durham, NC 27710. Kindling is a phenomenon in which brief afterdischarges (ADs) evoked by periodic electrical stimulation of the brain eventually result in generalized clonic motor seizures. Once present, the enhanced sensitivity to electrical stimulation is lifelong. The mechanism by which brief ADs produce this long-lasting effect may involve a change in gene expression. To begin to investigate changes in gene expression that occur during kindling, we used in situ hybridization histochemistry to examine the time course of expression of mRNAs of the immediate early genes (IEGs) c-fos, c-jun, NGFI-A, and c-myc within the dorsal hippocampus of rats following a kindling AD. Three principal findings resulted from this study. First, the expression of all mRNAs except c-myc was significantly increased (P less than 0.05) within discrete neuronal populations. Second, the time course of expression of the IEGs differed markedly within the same neuronal population. Third, for a given IEG, the time course and anatomic pattern of expression were strikingly different among different neuronal populations of the hippocampus. The prolonged and distinctly different patterns of IEG expression suggest that target genes are differentially regulated in these neuronal populations for prolonged periods following a kindling AD. PMID: 1661808 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 883: Endocrinology. 1991 Sep;129(3):1243-9. Regulation of c-fos, c-jun, jun-B, and c-myc messenger ribonucleic acids by gonadotropin and growth factors in cultured pig Leydig cell. Hall SH, Berthelon MC, Avallet O, Saez JM. INSERM U307, Hopital Debrousse, Lyon, France. The nuclear protooncogenes have been implicated in the coordinate regulation of gene expression during cell proliferation and differentiation. Previous work has shown that LH and human h CG as well as several growth factors including epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), transforming growth factor-beta, and insulin-like growth factor-I play a role in Leydig cell differentiated functions. To evaluate the possibility that protooncogenes mediate long term effects of these factors, their action on the levels of c-fos, c-jun, jun-B, and c-myc messenger (m) RNAs was studied. hCG (10(-9) M) produced a time-dependent increase in c-fos (9-fold), jun-B (18-fold) and c-myc (5-fold) mRNA levels but did not affect c-jun. The concentration of hCG required for half-maximal stimulation (ED50 = 7 +/- 4 x 10(-12) M) was similar to that required to induce half-maximal testosterone production. At optimal concentrations, the effects of EGF and bFGF on c-fos and jun-B mRNAs were lower than those induced by hCG, but their effects on c-myc mRNA were higher. In addition, they stimulated c-jun. Moreover, EGF and bFGF potentiated the effects of hCG on c-fos and jun-B, whereas hCG potentiated the action of growth factors on c-jun. Transforming growth factor-beta increased only jun-B mRNA, whereas insulin-like growth factor I increased c-fos, jun-B, and c-myc but less effectively than hCG. Lastly, the phorbol ester phorbol 12-myristate 13-acetate increased the level of the four protooncogene mRNAs, and its effects on c-fos and c-myc were significantly higher than those produced by hCG. These data indicate that the regulation of protooncogene mRNAs in normal Leydig cells is multifactorial. They also show differential responsiveness of the members of the Jun family to several factors. Our results are consistent with the hypothesis that the Fos and Jun families of regulatory proteins could play a role in mediating long term responses to the complex array of hormones and growth factors to which Leydig cells are exposed in vivo. PMID: 1651842 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 884: J Biol Chem. 1991 Aug 15;266(23):15277-85. Pro-Leu-Ser/Thr-Pro is a consensus primary sequence for substrate protein phosphorylation. Characterization of the phosphorylation of c-myc and c-jun proteins by an epidermal growth factor receptor threonine 669 protein kinase. Alvarez E, Northwood IC, Gonzalez FA, Latour DA, Seth A, Abate C, Curran T, Davis RJ. Department of Biochemistry and Molecular Biology, University of Massachusetts Medical School, Worcester 01605. A growth factor-stimulated (MAP2-related) protein kinase, ERT, that phosphorylates the epidermal growth factor receptor at Thr669 has been purified from KB human tumor cells by Northwood and co-workers (Northwood, I. C., Gonzalez, F. A., Wartmann, M., Raden, D. L., and Davis, R. J. (1991) J. Biol. Chem. 266, 15266-15276). The ERT protein kinase has a restricted substrate specificity, and the structural determinants employed for substrate recognition by this enzyme have not been defined. As an approach toward understanding the specificity of substrate phosphorylation, we have used an in vitro assay to identify additional substrates for the ERT protein kinase. In this report we describe two novel substrates: (a) the human c-myc protein at Ser62 and (b) the rat c-jun protein at Ser246. Alignment of the primary sequences surrounding the phosphorylation sites located within the epidermal growth factor receptor (Thr669), Myc (Ser62), and Jun (Ser246) demonstrated a marked similarity. The observed consensus sequence was Pro-Leu-Ser/Thr-Pro. We propose that this sequence forms part of a substrate structure that is recognized by the ERT protein kinase. PMID: 1651323 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 885: J Med Virol. 1991 Aug;34(4):241-7. Activation of cellular oncogenes by clinical isolates and laboratory strains of human cytomegalovirus. Boldogh I, AbuBakar S, Fons MP, Deng CZ, Albrecht T. Department of Microbiology, University of Texas Medical Branch, Galveston 77550. The effect on cellular (c) oncogene RNA levels was investigated after infection of permissive cells with cell culture adapted strains (AD-169, C-87, Davis) and unadapted clinical isolates (82-1, 84-2, 85-1) of human cytomegalovirus (HCMV). The results indicate that both adapted and unadapted strains of HCMV induce substantial increases in c-oncogene RNA levels for fos, jun, and myc measured by Northern blot hybridization. Elimination of immediate early (IE) protein synthesis between 0 and 3 hrs or reduction of virus infectivity (99.99%) by UV-irradiation did not reduce the increase in c-oncogene RNA levels. Inhibition of viral and cellular protein synthesis by cycloheximide resulted in a high abundance (superinduction) of specific RNAs which hybridized to c-oncogene probes after infection with either adapted or unadapted strains of HCMV. These data suggest that IE viral gene expression is not essential for activation of c-oncogenes. Inhibition of DNA-dependent RNA synthesis by blocking RNA elongation with actinomycin-D or by inhibiting the activity of RNA polymerase II with alpha-amanitin significantly reduced the increase in c-oncogene RNA levels, suggesting that activation of cellular genes by HCMV is controlled at the level of transcription. Activation of c-oncogenes by HCMV may be particularly important because their protein products appear to be involved in initiation and regulation of viral and cellular gene expression. PMID: 1719130 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 886: J Immunol. 1991 Jul 15;147(2):520-7. An NF-kappa B-like transcription factor mediates IL-1/TNF-alpha induction of gro in human fibroblasts. Anisowicz A, Messineo M, Lee SW, Sager R. Division of Cancer Genetics, Dana-Farber Cancer Institute, Boston, MA 02115. Normal human foreskin fibroblasts were used to examine transcriptional induction by IL-1 and TNF-alpha of the novel cytokine gro (melanoma growth-stimulating activity). Gro mRNA was expressed at levels 100-fold above background within 45 min of exposure to either IL-1 or TNF-alpha, in growing or serum-starved cells and a similar response was shown by IL-6. In contrast, as shown previously, gro mRNA was elevated only 10-fold by serum in starved but not in growing cells, similar to fos. Thus gro expression appears to be regulated by at least two signal transduction systems: a cytokine pathway, and a growth-related pathway. Three closely related gro genes (alpha, beta, and gamma) have been described. Their proximal 5' regulatory sequences presented here show close similarity in the region to -136, which includes the NF-kappa B site at -66 to -76 in gro alpha and gro gamma, and -64 to -74 in gro beta, and sequence diversity further upstream. Transient transfection of HeLa cells with CAT constructs localized the cytokine response to a region between -84 and -65 in gro beta. Gel retardation studies with FS-2 cells identified a cytokine-induced protein binding at the NF-kappa B site in all three gro genes as shown by competition studies with a pair of oligonucleotides representing wild-type and mutant sequences of the NF-kappa B binding site. Neither serum nor PMA induced a detectable gel shift at NF-kappa B or upstream to position -723. These results demonstrate conservation of the cytokine response element, NF-kappa B, in the three genes, consistent with the conservation of sequence in this region; and suggest that differential expression of the three gro genes may depend upon interactions with other sites located in the divergent upstream region. PMID: 1906501 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 887: Oncogene. 1991 Jul;6(7):1259-68. The stimulation of quiescent rat fibroblasts by v-src and v-fps oncogenic protein-tyrosine kinases leads to the induction of a subset of immediate early genes. Jahner D, Hunter T. Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92186. The stimulation of quiescent murine fibroblasts by growth factors and by phorbol esters results in a rapid and transient transcriptional activation of a large group of so-called immediate early genes. Several such genes were found to be induced in chicken embryo fibroblasts following activation of a temperature sensitive (ts) Rous sarcoma virus v-src mutant following temperature shift (Simmons et al., 1989). In contrast, the classical immediate early genes c-myc, c-fos and c-jun were essentially uninducible upon activation of a ts v-src mutant in rat-1 fibroblasts (Welham et al., 1990). We have cloned 9 cDNAs of genes that are rapidly and transiently inducible in rat fibroblasts by ts v-src mutants, and by a ts Fujinami sarcoma virus v-fps mutant. Six of these cDNAs are derived from the known immediate early genes NGFI-A, KC, c-fos, tissue factor, PC4 and ornithine decarboxylase; the other three cDNAs have not been described before. These 9 genes showed individual profiles of inducibility by fetal calf serum, epidermal growth factor (EGF) and by phorbol esters. Their response to the retroviral oncogenic protein-tyrosine kinases correlated best with the one to EGF, suggesting a common pathway of signal transduction. c-fos did not respond strongly to this pathway but was well induced by fetal calf serum. NGFI-A, however, was induced to a similar extent by all activators tested. Furthermore, we demonstrated that the induction of several of these genes by the retroviral oncogenic protein-tyrosine kinases is rapid, direct and occurs at the transcriptional level. PMID: 1861868 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 888: Cell Regul. 1991 Jun;2(6):479-89. Requirement of the adenovirus E1A transformation domain 1 for inhibition of PC12 cell neuronal differentiation. Heasley LE, Benedict S, Gleavy J, Johnson GL. Division of Basic Sciences, National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado. Expression of the adenovirus early gene E1A inhibits the nerve growth factor (NGF)-induced differentiation of PC12 pheochromocytoma cells. Expression of the 12S form of E1A, which lacks the transcription activation region, also inhibited PC12 cell differentiation in a manner similar to the wild-type gene. Three cellular proteins--the retinoblastoma susceptibility gene product referred to as 105(Rb)-, 107-, and 300-kDa proteins--stably interacted with the different E1A polypeptides. Analysis of the association of these cellular proteins with mutant E1A polypeptides demonstrated that a functional domain 1, which is minimally involved in the association of the 300-kDa protein with E1A, was sufficient to inhibit neuronal differentiation. Deletion of transformation domain 2, which encodes sequences necessary for the binding of the 105(Rb)- and 107-kDa proteins, did not influence the ability of the mutant E1A polypeptide to inhibit PC12 cell differentiation. E1A was also shown to alter the expression of mRNAs for the early response genes c-fos, c-myc, egr-1, and c-jun and their regulation in response to NGF. In clones expressing either 12S or 13S E1A, NGF stimulation of c-fos and c-myc was repressed. In contrast, basal mRNA levels for c-jun and egr-1 were constitutively elevated and not significantly affected further by challenge with NGF. Simply expressing c-jun by gene transfer, however, did not mimic the action of E1A because constitutively expressing c-jun clones differentiated in response to NGF. Thus, expression of the E1A polypeptide disrupts NGF control of early transcription events that have been shown to be critical for PC12 cell neuronal differentiation. PMID: 1832020 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 889: Oncogene. 1991 Jun;6(6):1049-56. T antigens' role in polyomavirus transformation: c-myc but not c-fos or c-jun expression is a target for middle T. Rameh LE, Armelin MC. Universidade de Sao Paulo, Instituto de Quimica, Brasil. Polyoma virus (Py) causes neoplastic transformation in vitro and multiple tumors in vivo. The role played by large and middle T antigens (LT, MT) and their mechanisms of action are focused here. Py-transformed Balb-3T3 cells become independent of platelet-derived growth factor (PDGF) for growth. JE, c-fos, c-jun and c-myc are 'immediate early' genes induced in response to PDGF. To test whether these cellular genes play a role in malignant transformation by Py, we generated a number of transfectant cell lines overexpressing LT, MT or both. Characterization of these cell lines revealed that: (a) MT but not LT causes morphological transformation, ability to grow in agarose suspension; (b) cooperation between LT and MT is evident in vitro, however, high and simultaneous LT and MT expression does not warrant tumorigenic potential; (c) MT expression does not correlate with tumorigenic potential but alters the probability of eliciting tumors; (d) JE and c-myc (but not c-fos or c-jun) are constitutively expressed in MT transfectants. MT induction is followed by c-myc induction 1.5 h later. We conclude that some of the 'immediate-early' genes may play pivotal roles in Py transformation. PMID: 1648700 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 890: Proc Natl Acad Sci U S A. 1991 May 1;88(9):3530-4. Analysis of the genes involved in the insulin transmembrane mitogenic signal in Chinese hamster ovary cells, CHO-K1, utilizing insulin-independent mutants. Mamounas M, Ross S, Luong CL, Brown E, Coulter K, Carroll G, Englesberg E. Department of Biological Sciences, University of California, Santa Barbara 93106. CHO-K1 cells, wild type (WT), grow in a defined medium with insulin as the only essential hormone. When starved for insulin, these cells accumulate in G0/G1 stage. Insulin binding to its receptor stimulates DNA synthesis and cell division and induces an increase in abundance of mRNA for c-fos, c-jun, Krox-20, Krox-24 (zif/268), fra-1, jun-B, c-myc, and JE. The kinetics of induction of these genes are similar to that shown with serum induction of 3T3. These genes show maximum stimulation at insulin concentrations of 20, 160, or 320 ng/ml and their expression is inhibited at higher concentrations. The addition of cycloheximide results in superinduction. The WT and insulin-independent mutants show no detectable signal for KC, fos-b, or nur77 and no increase over the basal level of pI-15, probably eliminating these genes as participants in the insulin mitogenic signal. These mutants synthesize DNA in the absence of insulin at rates that vary from 4 to 12 times that of the quiescent (insulin unstimulated) WT and are further inducible by insulin. The mutants have "constitutive" levels of Krox-24 (zif/268), fra-1, jun-B, c-myc, and JE (INS-type 2 genes) mRNAs that vary from mutant to mutant, reaching a maximum of an 8-fold increase for fra-1 and JE over the quiescent WT levels. There were no detectable levels of mRNA for genes c-fos and Krox-20 and no increase in level of mRNA for c-jun (INS-type 1 genes) as compared to the quiescent WT. Thus, although these INS-type 1 and type 2 genes may be involved in the full insulin mitogenic signal, the constitutive up-regulation of only genes in INS-type 2 is sufficient for insulin-independent DNA synthesis and cell division. Analysis of hybrids constructed between WT and mutant 27 indicate that the mutant phenotype is recessive, pointing to the existence of a regulatory gene producing a negative regulator. PMID: 1902566 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 891: Kidney Int. 1991 May;39(5):946-53. Proto-oncogene expression in peripheral blood mononuclear cells in IgA nephropathy. Ebihara I, Nakamura T, Suzuki S, Tomino Y, Koide H. Department of Medicine, Juntendo University School of Medicine, Tokyo, Japan. We have investigated the expression of proto-oncogenes in mononuclear cells obtained from patients with IgA nephropathy using a RNA hybridization technique. Patients with IgA nephropathy expressed more c-myc, c-raf, c-fos, and c-jun proto-oncogene RNA than did normal controls. However, no significant expression of c-N-ras, c-mos or c-myb genes was found in the mononuclear cells of these patients. When the amount of urinary protein excretion was used as an indicator of disease activity (greater than 1 g/day), a positive correlation was found between c-myc, c-raf, c-fos, and c-jun expression and urinary protein excretion (P less than 0.01). The expression of these genes correlated also with the serum IgA concentration (P less than 0.01), IgA immune complex (P less than 0.01), and histopathological changes in renal tissues obtained from patients with IgA nephropathy (P less than 0.01). The results of this survey suggest that abnormally regulated proto-oncogene expression in mononuclear cells may play an important role in the progression of IgA nephropathy and may be useful as an indicator of disease activity and/or prognosis. PMID: 1712408 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 892: Neuroreport. 1991 Apr;2(4):173-6. CDF/LIF selectively increases c-fos and jun-B transcripts in sympathetic neurons. Yamamori T. Biology Division, California Institute of Technology, Pasadena 91125. We recently demonstrated that the neuronal cholinergic differentiation factor (CDF) which switches the neurotransmitter phenotype of cultured sympathetic neurons from noradrenergic to cholinergic is identical to leukemia inhibitory factor (LIF). To elucidate some of the initial events leading to the phenotypic switch, the effects of CDF/LIF on the mRNA levels of several immediate early genes were examined in cultured neonatal rat sympathetic neurons. c-fos and jun-B were induced within 30 min of addition of CDF/LIF. In contrast, no effect on the expression of c-myc, fra-1, v-jun or actin mRNA was detected at this time. Thus, CDF/LIF may induce the expression of particular immediate early genes prior to its positive and negative effects on neurotransmitter and neuropeptide gene expression. PMID: 1909907 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 893: Oncogene. 1991 Apr;6(4):669-72. Transfected mouse c-jun can inhibit transformation of primary rat embryo fibroblasts. Ginsberg D, Hirai SI, Pinhasi-Kimhi O, Yaniv M, Oren M. Department of Chemical Immunology, Weizmann Institute of Science, Rehovot, Israel. The c-jun gene, which encodes a transcriptional regulatory protein, is the cellular homologue of the transforming gene of avian sarcoma virus 17. In an attempt to assess the biological activities of mouse c-jun, we studied the consequences of its overproduction in an in vitro transformation assay. A c-jun expression plasmid failed to cooperate with either ras, myc or mutant p53 in this focus formation assay. On the other hand, it dramatically inhibited the ability of various oncogene combinations to elicit foci upon transfection into primary rat embryo fibroblasts. Deletion plasmids lacking either the transactivating domain or the leucine repeat of c-jun still displayed a pronounced inhibitory activity. On the contrary, a plasmid encoding only the first 187 amino acids of c-jun had no such activity. The data suggests that enhanced c-jun expression may interfere with the induction or proliferation of transformed cells in this system, and that the inhibitory activity resides in the C-terminal half of the molecule. PMID: 1903196 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 894: Curr Opin Cell Biol. 1991 Apr;3(2):261-8. Temporal events regulating the early phases of the mammalian cell cycle. Naeve GS, Sharma A, Lee AS. University of Southern California School of Medicine, Los Angeles. It is proposed that the regulation of the pathways directing mammalian cell cycle progression involves several oncogenes. A summary of what is known about some of these regulatory oncogenes (fos, jun, myc, and Rb-1) and where they might function in the progression of a cell from G0 to G1 and G1 to S is presented. Data on two replication-dependent genes, those encoding histones and thymidine kinase, respectively, are also presented as models for describing transcriptional and post-transcriptional events at the G1-S border. Publication Types: Review Review, Tutorial PMID: 1883619 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 895: J Cell Physiol. 1991 Apr;147(1):121-7. The half-lives of platelet-derived growth factor A- and B-chain mRNAs are similar in endothelial cells and unaffected by heparin-binding growth factor-1 or cycloheximide. Gay CG, Winkles JA. Jerome H. Holland Laboratory for the Biomedical Sciences, American Red Cross, Rockville, Maryland 20855. Platelet-derived growth factor (PDGF) is mitogenic and chemotactic for vascular smooth muscle cells cultured in vitro, and, thus, may play a role in the smooth muscle cell proliferation and migration that occurs during atherosclerotic lesion development. Two related PDGF polypeptides, designated as the A and B chains, form functionally active PDGF-AA, AB, or BB dimers. The PDGF A- and B-chain genes are both transcribed in human umbilical vein endothelial (HUVE) cells and their expression is regulated by cytokines, growth factors, endotoxin, and phorbol ester. We reported previously that the angiogenic polypeptide heparin-binding growth factor (HBGF)-1 induces PDGF A-chain gene expression, but does not affect PDGF B-chain gene expression. In this study, we determined whether mRNA stabilization contributed to this induction by measuring the half-life of PDGF A-chain mRNA in quiescent, HBGF-1-stimulated, and proliferating HUVE cells. PDGF A-chain mRNA levels increase when quiescent HUVE cells are treated with the protein synthesis inhibitor cycloheximide; therefore, the effect of cycloheximide on PDGF A-chain mRNA decay was also investigated. The half-life of PDGF A-chain transcripts in quiescent cells was approximately 2.4 h and neither HBGF-1 nor cycloheximide significantly altered this decay rate. We also estimated the half-life of PDGF B-chain mRNA under the three different growth conditions and in the absence or presence of cycloheximide. The half-life in quiescent cells was approximately 1.8 h and was unaffected by HBGF-1 or protein synthesis inhibition. Therefore, the PDGF mRNAs have similar decay rates in HUVE cells, even though the 3' untranslated region of B-chain transcripts, but not A-chain transcripts, contains AU-rich sequence motifs postulated to confer rapid turnover in vivo. PMID: 1709940 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 896: Oncogene. 1991 Mar;6(3):455-60. Phorbol ester-mediated inhibition of growth and regulation of proto-oncogene expression in the human T cell leukemia line JURKAT. Makover D, Cuddy M, Yum S, Bradley K, Alpers J, Sukhatme V, Reed JC. Department of Internal Medicine, University of Pennsylvania, Philadelphia 19104. The expression and function of several proto-oncogenes were examined in a human acute T cell leukemia line, JURKAT, during phorbol ester-induced terminal differentiation. Treating JURKAT cells with the phorbol ester tetradecanoyl phorbol acetate (TPA) inhibited their proliferation and induced expression of the gene for the interleukin 2 receptor alpha chain (IL2R-alpha), consistent with previous reports. In unstimulated proliferating JURKAT cells, high levels of C-MYC, N-RAS, and BCL2 mRNAs were found that diminished rapidly following TPA-induced cessation of growth. In contrast, accumulation of mRNAs for the C-FOS, C-JUN, and EGR-1 genes increased markedly in TPA-treated cells and preceded the induction of IL2R-alpha mRNA. Expression of C-MYB, C-RAF-1, C-LCK, C-FYN, and C-FGR proto-oncogenes was relatively unchanged. To explore directly the function of two of these protooncogenes in regulating the growth of JURKAT T cells, we stably transferred C-MYC and BCL2 expression plasmids into these cells. Despite sustained expression of C-MYC, BCL2, or the combination of these protooncogenes, TPA continued to inhibit JURKAT cell growth and to induce IL2R expression. Thus, although C-MYC and BCL2 proto-oncogene expression correlated with proliferation in TPA-treated JURKAT cells, continuous over-expression of even the combination of these oncogenes was insufficient for abrogating the effects of TPA in these leukemic T cells. Because human lymphoid malignancies frequently contain chromosomal translocations that deregulate the expression of C-MYC and BCL2, our findings could have relevance for attempts to induce terminal differentiation of leukemic cells by in vitro exposure of patients' bone marrow cells to phorbol esters. PMID: 2011401 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 897: J Immunol. 1991 Mar 1;146(5):1509-15. IL-1 induces ornithine decarboxylase in normal T lymphocytes. Bristol LA, Smith MR, Bhat NK, Durum SK. Laboratory of Molecular Immunoregulation, Program Resources, Inc., Frederick, MD. IL-1 alpha regulation of ornithine decarboxylase (ODC) was examined in T cells because IL-1 is a costimulus for T cell proliferation and ODC is a critical enzyme in the metabolic events associated with cellular proliferation. In the present study, we demonstrate that IL-1 alpha induces ODC mRNA and ODC enzyme activity in T cells. Unlike many IL-1 actions on T cells, this did not require a costimulus from the TCR, IL-1 alone being sufficient to induce ODC. The mechanism of IL-1 induction of ODC probably operates at several levels, including transcription, mRNA stability, and translation. Previous studies have shown that IL-1 prepares T cells for replication by increasing the production of c-jun, c-fos, c-myc, growth factors, growth factor receptors, and the response to growth factors. From the present study, ODC induction can be added to the list of IL-1-induced replicative machinery in T cells. PMID: 1993840 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 898: J Virol. 1991 Mar;65(3):1568-71. Transcriptional activation of cellular oncogenes fos, jun, and myc by human cytomegalovirus. Boldogh I, AbuBakar S, Deng CZ, Albrecht T. Department of Microbiology, University of Texas Medical Branch, Galveston 77550. The mechanisms responsible for the human cytomegalovirus (HCMV)-induced increase in cellular oncogene RNAs for c-jun, c-fos, and c-myc in human embryo lung cells (I. Boldogh, S. AbuBakar, and T. Albrecht, Science 247:561-564, 1990) were investigated. Results of transcription assays indicated that the rapid increase in RNA levels for the above-noted oncogenes was controlled at the transcriptional level and was related to enhanced transcription. The maximum rates of transcription for c-jun and c-fos genes occurred at 40 min postinfection, while for the c-myc gene the maximum rate occurred at about 60 min. The magnitude of HCMV-induced activation of these cellular genes was similar to the activation induced by serum. The half-lives of the cellular oncogenes showed similar decay rates after either serum or HCMV activation when measured by dactinomycin chase. The half-life for c-fos or c-jun was about 20 min, and that for c-myc was about 40 min. Furthermore, inhibition of the RNA increase by dactinomycin or by alpha-amanitin suggested that the increase in RNA levels was due to an increase in the transcriptional activity of oncogenes triggered by HCMV. PMID: 1847472 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 899: Nucleic Acids Res. 1991 Feb 25;19(4):739-46. Non-leucine residues in the leucine repeats of Fos and Jun contribute to the stability and determine the specificity of dimerization. Schuermann M, Hunter JB, Hennig G, Muller R. Institut fur Molekularbiologie und Tumorforschung (IMT), Phillipps-Universitat Marburg, FRG. Various transcription factors, including C/EBP, GCN4 and members of the Fos, Jun and Myc families have been shown to form highly specific complexes via alpha-helical structures referred to as leucine zippers. Experimental evidence has suggested that dimerization involves the formation of hydrophobic bonds between leucine residues in laterally aligned coiled coil structures. However, the specificity of interaction between leucine zipper proteins is not understood. In this study, we show that amino acids, which are located in positions a, e, and g are instrumental in the formation of Fos/Jun heterodimers, presumably by establishing intermolecular electrostatic and hydrophobic interactions. These residues are highly conserved in proteins of the Fos or Jun families but completely different between Fos and Jun, suggesting that these residues determine the specificity of interaction. This conclusion is supported by the observation that the substitution of amino acids in position a or g in Fos with the corresponding Jun amino acids facilitates the association of two Fos leucine repeats. In addition, we show that a conserved histidine residue, located 7 amino acids (i.e., two alpha-helical turns) C-terminally to the 5th leucine in Fos and Jun, is also important for complex formation. PMID: 1901988 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 900: Mol Cell Biol. 1991 Feb;11(2):954-62. Intracellular leucine zipper interactions suggest c-Myc hetero-oligomerization. Dang CV, Barrett J, Villa-Garcia M, Resar LM, Kato GJ, Fearon ER. Hematology Division, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205. The physiological significance of in vitro leucine zipper interactions was studied by the use of two strategies which detect specific protein-protein interactions in mammalian cells. Fusion genes were constructed which produce chimeric proteins containing leucine zipper domains from several proteins fused either to the DNA-binding domain of the Saccharomyces cerevisiae GAL4 protein or to the transcriptional activation domain of the herpes simplex virus VP16 protein. Previous studies in mammalian cells have demonstrated that a single chimeric polypeptide containing these two domains will activate transcription of a reporter gene present downstream of the GAL4 DNA-binding site. Similarly, if the GAL4 DNA-binding domain of a chimeric protein could be complexed through leucine zipper interactions with the VP16 activation domain of another chimeric protein, then transcriptional activation of the reporter gene would be detected. Using this strategy for detecting leucine zipper interactions, we observed homo-oligomerization between leucine zipper domains of the yeast protein GCN4 and hetero-oligomerization between leucine zipper regions from the mammalian transcriptional regulating proteins c-Jun and c-Fos. In contrast, homo-oligomerization of the leucine zipper domain from c-Myc was not detectable in cells. The inability of the c-Myc leucine zipper to homo-oligomerize strongly in cells was confirmed independently. The second strategy to detect leucine zipper interactions takes advantage of the observation that the addition of nuclear localization sequences to a cytoplasmic protein will allow the cytoplasmic protein to be transported to and retained in the nucleus. Chimeric genes encoding proteins with sequences from a cytoplasmic protein fused either to the GCN4 or c-Myc leucine zipper domains were constructed. Experiments with the c-Myc chimeric protein failed to demonstrate transport of the cytoplasmic marker protein to the nucleus in cells expressing the wild-type c-Myc protein. In contrast, the cytoplasmic marker was translocated into the nucleus when the GCN4 leucine zippers were present on both the cytoplasmic marker and a nuclear protein, presumably as a result of leucine zipper interaction. These results suggest that c-Myc function requires hetero-oligomerization to an as yet undefined factor. PMID: 1990293 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 901: Leukemia. 1991 Feb;5(2):142-9. Childhood undifferentiated leukemia with early erythroid markers and c-myb duplication. Castaneda VL, Parmley RT, Saldivar VA, Cheah MS. Department of Pediatrics, University of Texas Health Science Center, San Antonio 78284-7810. Ultrastructural, flow cytometric, and molecular studies were performed on leukemia cells from bone marrow and pleural effusion of a 6-year-old boy diagnosed with undifferentiated (MO) leukemia, using routine histology and immunostains at diagnosis and relapse. Ultrastructurally, surface and/or intracellular ferritin particles were present on or in some blasts and the majority of blasts contained identifiable acid ferrocyanide reactive inorganic iron comparable to that seen in normal early erythroblasts. The cells lacked other evidence of differentiation, including diaminobenzidine-reactive or immunoreactive hemoglobin. Flow cytometric analysis of malignant cells showed a lack of lymphoid or myeloid markers. Anti-transferrin receptor antibody was positive on 93% of cells and antibody to glycophorin A reacted with 23% of cells. RNA blot analysis of leukemia cells with myeloperoxidase (MPO) showed an absence of appreciable levels of MPO mRNA. Chromosome analysis showed 51,XY, t(1;16)(p31;q24), +6, +10, +15, +19, +21. The oncogene c-myb, which is specifically expressed and regulated in hematopoietic cells and produces a DNA-binding protein responsible for myeloid differentiation, was found to be duplicated in the patient's tumor cells. Expression of c-jun, N-ras, c-myc, and p53 was normal. The data indicate that the malignant cells in this patient are of early erythroid lineage at diagnosis and relapse and that classification of cell lineage can be enhanced by ultrastructural Prussian blue staining. The failure of this otherwise undifferentiated leukemia to express or evolve into a myeloid phenotype is biologically and clinically distinct from previously described cases of erythroid and myeloid leukemia and may represent a previously unidentified phenotype which should be included in the spectrum of 'undifferentiated' childhood leukemia. Publication Types: Case Reports PMID: 1708434 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 902: Int J Cancer. 1991 Jan 21;47(2):227-37. Characterization of a highly invasive and spontaneously metastatic human malignant melanoma cell line. Welch DR, Bisi JE, Miller BE, Conaway D, Seftor EA, Yohem KH, Gilmore LB, Seftor RE, Nakajima M, Hendrix MJ. Division of Biology, Glaxo Research Laboratories, Research Triangle Park, North Carolina. Although the incidence of, and deaths due to, malignant melanoma are rising at a rapid rate, few experimental models mimic the highly metastatic properties associated with the pathogenesis of the human disease, making study of the disease difficult. Thus, new human models are required to understand melanoma biology, especially its metastatic properties. Here we describe C8161, a highly invasive and spontaneously metastatic human melanoma cell line, which grows progressively in the subcutis of athymic nude mice with an average doubling time of approximately 6 days. By the time the tumor reaches a diameter of 1 cm, amelanotic metastases in lymph nodes, skin, peritoneal wall, spleen and lungs have formed. By comparing C8161 to variants from other well-characterized human malignant melanomas (A375 and MeWo) with differing metastatic traits, properties presumed to be involved in metastatic propensity were examined. C8161 showed a 2- to 14-fold higher ability to invade reconstituted basement membrane barriers in the MICS and correspondingly high type-IV collagenase mRNA levels and collagenolytic activity, as compared with other melanoma cell lines. Likewise, differential adhesion to immobilized RBM or HUVEC monolayers was observed, but did not correlate to rank orders of malignant properties. Recently, a correlation between surface expression of ICAM-1 and secondary tumor formation by human melanomas has been described in several laboratories. Basal levels of ICAM-1 on C8161, A375 and MeWo human melanomas were compared, but no correlation with metastatic potential was noted. Proto-oncogene expression in C8161 cells was compared with A375P and A375M variants using Northern blot analysis. c-myc expression was 6-fold greater than both A375 variants; c-fos expression was 3.4-fold less than A375P and 1.7-fold less than A375M; c-jun in C8161 cells was 2.5-fold and 2.1-fold greater than expression in A375P and A375M, respectively. Because C8161 is so highly malignant, amenable to experimental manipulation, and its behavior in nude mice mimics the clinical course of malignant melanoma, this cell line will prove valuable for studying properties associated with human melanoma tumor progression. PMID: 1671030 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 903: Mol Carcinog. 1991;4(3):203-9. DNA methylation and oncogene expression in methapyrilene-induced rat liver tumors and in treated hepatocytes in culture. Hernandez L, Petropoulos CJ, Hughes SH, Lijinsky W. ABL-Basic Research Program, National Cancer Institute-Frederick Cancer Research and Development Center, Maryland 21702. Continued exposure of rats to carcinogenic doses of methapyrilene (MP) leads to elevated levels of 5-methyl-deoxycytidine (5MC) in liver DNA. Since gene expression often correlates with DNA methylation, we investigated these parameters in the MP-induced hepatocellular carcinomas of Fischer 344 rats. DNA was hypermethylated in liver tissue surrounding the tumors relative to liver tissue of untreated controls of the same age, while tumor DNA was not; DNA methylation declined to normal levels when MP treatment ceased. Gene expression analysis showed measurable levels of mRNA for c-Ki-ras, erb-B, erb-B2, hck, src, lyn, vav, trk, raf-1, l-myc, c-jun, c-yes, c-myc, c-abl, and p53. No significant differences in expression for these and other oncogenes were seen between tumors and surrounding livers, although erb-B2 and vav showed visible decreases compared with normal liver. Hypermethylation of DNA and expression of these oncogenes in MP-treated tissues were not correlated. Levels of mRNA for the same genes in MP-treated hepatocytes in culture were similar to in vivo levels; analysis of DNA synthesis levels showed that this gene expression pattern occurred in the absence of proliferation bursts or toxicity in these cells, thus suggesting that treatment in vivo may produce the same results. PMID: 2064726 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 904: Anticancer Res. 1991 Jan-Feb;11(1):473-80. Oncogene and MDR1 gene expression in rat rhabdomyosarcoma sublines of different metastatic potential. Hanania N, Boyano MD, Mangin C, Poupon MF. Biologie des Metastases, IRSC-CNRS, Villejuif, France. The expression of various protooncogenes (myc, Ki-ras, Ha-ras, erbB, fms, sis, jun and fos) and a gene implicated in multidrug resistance (mdr1) was investigated in cell sublines, isolated from a rat rhabdomyosarcoma cell line, and in the corresponding tumors induced by injection of these cells into syngeneic rats. These cell lines and tumors, selected or not by treatment with chlorozotocin (Czt) or adriamycin (Adr), differed in their tumorigenicities and metastatic potentials. Our results showed that 1) an increased expression of some protooncogenes could be correlated with the metastatic potential of tumors; 2) such a correlation was not observed in the cultured cells from which these tumors were derived; 3) mdr1 expression, similarly to that of protooncogenes, was correlated with metastatic potential in all tumors except the Adr-selected tumors. PMID: 2018384 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 905: IARC Sci Publ. 1991;(105):294-304. Role of oncogenes and tumour suppressor genes in human lung carcinogenesis. Harris CC, Reddel R, Pfeifer A, Iman D, McMenamin M, Trump BF, Weston A. Laboratory of Human Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, MD. Six families of activated protooncogenes, ras, raf, fur, neu, jun and myc have so far been associated with human lung cancer. Human bronchial epithelial cells in vitro are being used to investigate the functional role of these specific oncogenes and growth regulatory genes in carcinogenesis and tumour progression. When transferred into normal human bronchial epithelial cells by the highly efficient protoplast fusion method, the v-Ha-ras oncogene initiates a cascade of events leading to decreased responsiveness of these cells to inducers of squamous differentiation, aneuploidy and, less frequently, 'immortality' and tumorigenicity with metastasis in athymic nude mice. Transfection of the SV40 T antigen gene results in nontumorigenic cell lines that have a nearly normal pathway of terminal squamous differentiation and can be transformed into malignant cells by transfected Ha-ras, N-ras or Ki-ras. The combination of transfected c-myc and c-raf-1 also transforms human bronchial epithelial cells into neoplastic cells that exhibit some phenotypic traits found in small-cell carcinomas. These and other results indicate that proto-oncogenes dysregulate the pathways of growth and differentiation of human bronchial epithelial cells and play an important role in human carcinogenesis. Analyses of allelic deletion and somatic cell hybrids are being used to identify the chromosomal localization of tumour suppressor genes. We have examined 54 non-small-cell bronchogenic carcinomas with 13 polymorphic markers. Loss of heterozygosity was more frequent than among 23 squamous-cell carcinomas than among 23 adenocarcinomas or eight large-cell carcinomas. Loss of heterozygosity for chromosome 17p was found in 89% of cases of squamous-cell carcinoma and 18% of adenocarcinomas. Analysis of chromosome 11 for allelic deletions revealed two commonly deleted regions (11p13 and 11p15.5). Somatic cell hybrids between normal human bronchial epithelial cells and Hut292DM, a lung carcinoma cell line, had a finite lifespan in vitro and were nontumorigenic in athymic nude mice. Tumour suppressive effects of individual or combinations of specific human chromosomes on Hut292DM are being examined by formation of microcell-cell hybrids. Chromosome 11 has tumour suppressor activity in these hybrids. Both of these studies suggest that tumour suppressor genes play a dominant role in lung carcinogenesis and provide in-vitro model systems for isolating these genes by subtraction library and insertional mutagenesis techniques. Publication Types: Review PMID: 1855868 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 906: Dokl Akad Nauk SSSR. 1991;321(4):846-9. [Gene product p53 is involved in the regulation of activity of the c-fos proto-oncogene promotor] [Article in Russian] Vikhanskaia FL, Pospelova TV, Volkov IV, Kukushkin AN, Svetlikova SB, Poslepov VA. PMID: 1806357 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 907: Growth Factors. 1991;5(4):283-93. Immediate early gene responses of NIH 3T3 fibroblasts and NMuMG epithelial cells to TGF beta-1. Koskinen PJ, Sistonen L, Bravo R, Alitalo K. Department of Virology, University of Helsinki, Finland. Transforming growth factor beta has a wide range of physiological effects on cell growth and metabolism. We have previously reported on the rapid induction of jun transcription factors in TGF beta-treated cells. Here we show that the early genomic response to TGF beta-1 includes activation of a broad spectrum of serum-inducible genes both in NIH 3T3 fibroblasts and in NMuMG epithelial cells, which are growth-stimulated and growth-inhibited by TGF beta, respectively. Of particular interest is the presence of a putative nuclear DNA-binding receptor (N10) and zinc finger transcription factors (Krox 20 and Krox 24) among the TGF beta-induced genes. In addition to the stimulatory effects of TGF beta, expression of a few genes including c-myc is decreased in both types of cells. In cells transformed by neu or ras oncogenes the immediate early mRNA responses to TGF beta are deregulated. Our results suggest that certain transcription factors are required for both positive and negative regulation of cell proliferation by TGF beta, and that their relative concentrations may determine the subsequent cellular responses. PMID: 1777237 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 908: Mol Biol (Mosk). 1991 Jan-Feb;25(1):105-15. [The c-fos proto-oncogene promotor is not regulated by serum, epidermal growth factor, and phorbol ester in embryonal fibroblasts transformed by E1Aad5+cHa-ras-oncogenes] [Article in Russian] Pospelova TV, Medvedev AV, Kisliakova TV, Svetlikova SB, Pospelov VA. Regulation of c-fos protooncogene activity in rat embryonal fibroblasts (REF), E1Aad5-immortalized REF cells, and E1Aad5 + cHa-ras transformed REF cells has been investigated. The analysis of regulation of fos-promoter activity was done by means of transient and stable transfection of fos-CAT plasmid into immortalized and transformed cells. In parallel, the regulation of cellular c-fos as well as c-jun and c-myc genes expression has been studied. It has been found that in E1Aad5 + cHa-ras-transformed cells the expression of c-fos promoter has a constitutive, non-inducible character while in REF cells and cells immortalized by E1Aad5 the fos-promoter can be regulated by serum growth factors, EGF, and TPA. PMID: 1716733 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 909: Mol Carcinog. 1991;4(4):297-307. Isolation and partial characterization of a transformation-associated sequence from human nasopharyngeal carcinoma. Cao Y, Sun Y, Poirier S, Winterstein D, Hegamyer G, Seed J, Malin S, Colburn NH. Laboratory of Viral Carcinogenesis, National Cancer Institute, Frederick Cancer Research, Maryland 21702-1201. A transforming activity associated with Chinese nasopharyngeal carcinoma (NPC) cell line CNE2 DNA has been identified by transfer into nontransformed promotion-sensitive mouse JB6(P+) C141 cells. To clone this transformation-associated sequence, we carried out three cycles of transfection, followed by cloning of anchorage-independent transformants in soft agar. A tertiary CNE/JB6 clonal transfectant cell line 625 whose DNA showed transforming activity, as indicated in both soft-agar assay and nude-mice implantation, was used to make a genomic library in the vector lambda dash. Using the human repeated sequence Blur 8 to screen the library, we obtained 10 human Alu-positive clones. A cloned Alu-positive insert of 16 kbp, CNE 323, was characterized in detail. CNE 323 transferred moderate transforming activity when introduced into JB6 P+ cells and showed no homology to Ha-, Ki-, or N-ras genes; human promotion sensitivity genes; src, myb, jun, myc, fos, raf, or int-2 oncogenes; or epidermal growth factor receptor. The isolated CNE 323 DNA sequence appeared to preserve the genomic structure of the original sequence found in CNE2 cells and in nude mouse tumors induced by CNE2 cells or by CNE/JB6 transfectant cells, indicating that the cloned NPC sequence was activated during NPC carcinogenesis and not during transfection or construction of the library, and that the cloned sequence or a larger sequence of which it was part played a role in tumor formation. Finally, we identified a 1.3-kb mRNA that hybridizes to a subclone of the 16-kb NPC sequence in CNE2 cell poly (A)+ RNA. PMID: 1714741 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 910: Arch Virol. 1991;118(3-4):163-77. Cellular oncogene activation by human cytomegalovirus. Lack of correlation with virus infectivity and immediate early gene expression. Boldogh I, AbuBakar S, Millinoff D, Deng CZ, Albrecht T. Department of Microbiology, University of Texas Medical Branch, Galveston. The contribution of expression of human cytomegalovirus (HCMV) immediate early (IE) genes to the rapid and transient increase in cellular (c)-oncogene (fos, jun, myc) transcription following HCMV infection was investigated. A partial temporal overlap was observed between the increases in c-oncogene RNA levels and the increase in either transcripts from HCMV IE genes or the number of cells in which HCMV IE proteins were detected. The increases in c-oncogene RNA levels, however, slightly preceded the increase in the detection of HCMV IE transcripts or proteins. To distinguish between the temporal coincidence and a direct relationship between expression of HCMV IE genes and the increased transcription of c-oncogenes, the number of cells synthesizing HCMV IE proteins was reduced by infecting with virus stock enriched in defective particles. Alternatively, the synthesis of HCMV IE proteins was essentially eliminated by ultra-violet (UV) irradiation of virus stock or by inhibitors of protein synthesis. Virus stocks enriched in defective particles demonstrated a substantially reduced capacity to direct the synthesis of HCMV IE proteins, but were more efficient in activating c-oncogene expression than infectious virus stocks. Elimination of expression of HCMV IE genes by UV-irradiation of virus stock or by inhibiting de novo viral and/or cellular protein synthesis with cycloheximide (100 micrograms/ml) or anisomycin (100 micrograms/ml) did not eliminate the HCMV-induced increase in RNA levels of c-oncogenes. These data indicate that activation of these early response cellular genes is independent from de novo expression of HCMV IE proteins, and possibly involves biologically active virion proteins that are related to the induction of a cascade of cellular events associated with the binding of HCMV to its cellular receptor. PMID: 1712580 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 911: Oncogene Res. 1991;6(1):1-12. Interleukin-3 and phorbol esters induce different patterns of immediate-early gene expression in an interleukin-3 dependent cell line. McCubrey JA, Steelman LS, McKearn JP. Department of Microbiology & Immunology, East Carolina University School of Medicine, Greenville, North Carolina 27858. The myeloid interleukin-3 (IL-3) dependent cell line, FDC-P1, enters the G0 stage of the cell cycle after IL-3 deprivation for 24 hr. The expression of 13 protooncogenes and immediate-early genes was compared with 4 "control" genes after the addition of either IL-3 or phorbol myristate acetate (PMA) to IL-3-deprived cells. mRNA transcripts encoding c-myc and the T-cell receptor c-gamma gene were induced to high levels only after IL-3 addition, whereas c-fos, fos-B, c-jun, jun-B, Krox-20, and Krox-24 were induced transiently only after PMA addition. The remaining genes (fra-1, p53, jun-D, c-Ha-ras, c-Ki-ras, c-raf, beta-actin, ornithine decarboxylase, and histone 2B) were detected after culture with either IL-3 or PMA. When cells were serum- and IL-3-deprived, c-fos, fos-B, c-jun, jun-B, Krox-20, and Krox-24 were detected after exposure to either serum or PMA. Moreover, culture with cycloheximide and PMA resulted in superinduction of these genes, whereas cycloheximide and IL-3 addition did not. mRNAs encoding these genes had half-lives of 10-20 min after PMA treatment, whereas that of beta-actin was longer (greater than 2 hr), suggesting that short mRNA half-lives contributed to the transient nature of these genes. Although c-fos, fos-B, c-jun, jun-B, Krox-20, and Krox-24 expression can be detected in IL-3-dependent cells after exposure to either PMA or serum, these genes were not detected after IL-3 addition, which allows cell-cycle progression. These results document the existence of IL-3 and PMA-responsive genes and demonstrate that IL-3 and protein kinase C agonists can induce distinct patterns of gene expression. PMID: 1705318 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 912: Ann N Y Acad Sci. 1991;638:139-48. Mitogenic effects of fibroblast growth factors in cultured fibroblasts. Interaction with the G-protein-mediated signaling pathways. Paris S, Pouyssegur J. Biochemistry Center, CNRS, University of Nice, France. We have shown that FGF (basic or acidic) is mitogenic for quiescent hamster lung fibroblasts (CCL39 line). It is active alone but is much more efficient in synergistic combinations with G-protein-activating agents. When used alone, FGF appears to exert its mitogenic effects without involving any of the major G-protein-mediated signaling pathways. It causes no significant hydrolysis of phosphoinositides, it does not alter the activity of adenylate cyclase, and its mitogenicity is insensitive to pertussis toxin. It therefore seems likely that all pleiotropic actions of FGF are primarily mediated by the intrinsic protein tyrosine kinase of its receptors. However, FGF, acting through its receptor tyrosine kinase, and thrombin, acting through G-protein-coupled receptors, induce a common set of early responses detected within seconds or minutes at the level of membranes, cytoplasm, and nuclei. Typical examples of early responses are activation of Na/H antiporter and Na/K/Cl cotransporter, phosphorylation of ribosomal protein S6, and increased transcription of early-immediate genes (c-fos, c-jun, and c-myc). Not only various classes of growth factors acting via distinct transducing mechanisms activate common targets, but also their synergistic effects on reinitiation of DNA synthesis is reflected on the early responses. How does the coordination of these signaling events take place? A partial answer to this question is illustrated in Figure 6 in which "switch kinases" play the role of integrators of multiple extracellular signals. Raf and, perhaps more convincingly, MAP kinases that are activated by dual phosphorylation on tyrosine and threonine residues are potential good candidates for this integration. This hypothetical scheme could therefore explain, in part, the coordination and the synergy commonly observed in the mitogenic response. The synergy could be generated at the level of MAP kinases simply by dual activating phosphorylations. With the recent cloning of MAP kinases, these questions will be more easily addressed. Another important gap that will have to be filled in future studies is the identification of all the members of the kinase cascade. When used in synergistic combinations with G-protein-activating agents, FGF does exert in contrast some effects on the G-protein-mediated pathways. It potentiates the G-protein-mediated activations of both PIP2-PLC and adenylate cyclase.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 1664681 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 913: Biochem Biophys Res Commun. 1990 Dec 31;173(3):1322-30. Neurokinin A induces c-fos, c-jun, and c-myc expression in L6 rat myoblasts. Rahm M, Hultgardh-Nilsson A. Department of Medical Cell Genetics, Medical Nobel Institute, Karolinska Institutet, Stockholm, Sweden. Neurokinin A (NKA), a neuropeptide belonging to the tachykinin family, induced c-fos proto-oncogene mRNA expression in serum-deprived L6J1 rat skeletal myoblasts in vitro. The marked increase reached maximal levels after 15 to 30 min. In contrast to this, c-jun and c-myc proto-oncogene expression were only slightly induced, with peak levels after 30 min. NKA did not stimulate DNA synthesis or cell proliferation in serum-deprived L6J1 myoblasts. We demonstrate a relationship between NKA treatment and induction of c-fos, c-jun and c-myc mRNA expression in serum-deprived L6J1 rat myoblasts. The results on DNA synthesis and cell proliferation indicate that the induced proto-oncogene expression alone is not enough to induce a cellular response to NKA. Possible mechanisms of action are discussed. PMID: 2176488 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 914: J Biol Chem. 1990 Dec 25;265(36):22292-9. Strong and persistent activation of inositol lipid breakdown induces early mitogenic events but not Go to S phase progression in hamster fibroblasts. Comparison of thrombin and carbachol action in cells expressing M1 muscarinic acetylcholine receptors. Seuwen K, Kahan C, Hartmann T, Pouyssegur J. Centre de Biochimie-Centre National de la Recherche Scientifique, Universite de Nice-Sophia Antipolis, Nice, France. In order to evaluate the role of phosphoinositide turnover in growth factor action, we expressed human M1 muscarinic acetylcholine (Hm1) receptors in Chinese hamster lung fibroblasts (CCL39 cell line). In the transfected cells (39M1-81 clone), but not in wild type fibroblasts, the muscarinic agonist carbachol induced a release of inositol phosphates as strong as alpha-thrombin, a very potent growth factor and activator of phosphoinositide-specific phospholipase C (PLC) in this cell system. In contrast to thrombin, carbachol-stimulated PLC activity was not inhibited by pertussis toxin treatment of cells. At concentrations that elicited a comparable initial rate of inositol phosphate release (10 nM for thrombin and 0.1 mM for carbachol), both agents gave rise to an identical calcium signal and equally stimulated Na+/H+ exchange and the transcription of the early genes c-jun, c-fos, and c-myc. Surprisingly, however, carbachol is not a mitogen for 39M1-81 cells, and even if tested in association with insulin or fibroblast growth factor, its effects on cell proliferation remained weak when compared with thrombin. Also, the muscarinic agonist did not stimulate soft agar colony forming capacity and did not prevent growth arrest in Go upon serum deprivation of cycling 39M1-81 cells. The failure of carbachol to induce cell proliferation could not be attributed to rapid and complete desensitization of Hm1 receptors nor to the activation of inhibitory pathways like adenylyl cyclase stimulation. We conclude that strong and persistent activation of phosphoinositide turnover elicits early biochemical events generally associated with mitogenesis, but is not sufficient to stimulate or maintain continuous cell proliferation. On the basis of our results, we postulate that thrombin mitogenesis depends critically on signaling events different from phosphoinositide turnover, possibly the stimulation of a receptor tyrosine kinase or a Gi protein-activated tyrosine kinase. PMID: 2176213 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 915: J Immunol. 1990 Dec 15;145(12):4355-64. Transformation of T lymphocytes by the v-fos oncogene. Valge-Archer VE, de Villiers J, Sinskey AJ, Rao A. Division of Tumor Virology, Dana Farber Cancer Institute, Boston, MA 02115. Activation of T lymphocytes through the T cell antigen receptor has been shown to stimulate a rapid and transient accumulation of c-fos mRNA and protein. Transfection of a normal murine T lymphocyte clone with the FBJ-v-fos oncogene resulted in generation of a cell line that was morphologically transformed, had lost the requirement for IL-2 for proliferation, and was tumorigenic in adult syngeneic mice; however, the transformed cells retained the ability to proliferate in response to IL-2. The transformed cells did not show constitutive expression of IL-2 or c-fos mRNA, although the promoter regions of both IL-2 and c-fos genes contain AP-1 sites that are expected to be targets for binding of Fos/Jun complexes. In contrast, the transformed T cells showed increased constitutive expression of IL-2R alpha and c-myc mRNA; these genes may represent cellular targets for transformation by v-fos and physiologic activation by c-fos. We discuss the possibility that these transformed cells behave as cells partially activated through the TCR, and that transformation occurs through a mechanism independent of IL-2. PMID: 2124241 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 916: Eur J Biochem. 1990 Dec 12;194(2):527-32. Neurokinin A induces expression of the c-fos, c-jun, and c-myc genes in rat smooth muscle cells. Hultgardh-Nilsson A, Larsson SH, Jin P, Sejersen T, Ringertz NR. Department of Medical Cell Genetics, Medical Nobel Institute, Stockholm, Sweden. Neurokinin A, a member of the tachykinin family of neuropeptides, has been identified as a mitogen for cultured smooth muscle cells. Tachykinin-induced DNA synthesis has previously been shown to be mediated by a receptor-specific mechanism and to correlate with accumulation of phosphatidylinositol 4,5-bisphosphate breakdown products. In the present experiments, we have studied intracellular pH and expression of the proto-oncogenes c-myc, c-jun and c-fos in smooth muscle cells exposed to mitogenic concentrations of neurokinin A. Growth-arrested smooth muscle cells stimulated with neurokinin A responded with an amiloride-sensitive intracellular alkalinization, indicating Na+/H+ antiport activation. c-myc and c-jun mRNA expression was only slightly elevated by neurokinin A, while c-fos expression underwent a more pronounced increase. Maximal levels of c-fos transcripts were found after 15 min and 30 min following neurokinin A stimulation. The results demonstrate that neuropeptides may influence proto-oncogene expression in smooth muscle cells and suggest a mechanism by which peripheral neurons may modulate differentiation and growth of these cells. PMID: 2176599 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 917: Am Rev Respir Dis. 1990 Dec;142(6 Pt 2):S20-6. Gene amplification in human lung cancer. The myc family genes and other proto-oncogenes and growth factor genes. Bergh JC. Department of Oncology, University of Uppsala, Sweden. The development of human lung cancer may require multiple genetic deletions affecting a number of chromosomes, e.g., 1, 3, 11, 13, and 17. These genetic aberrations may induce the activation of proto-oncogenes (c-jun, ras, c-raf1) and the loss of tumor suppressor genes (p53). Some of the activated proto-oncogenes and tumor suppressor genes are more selectively expressed or absent in small-cell lung cancer (L-myc, c-myb, c-scr, Rb gene) or non-small-cell lung cancer (c-erbB-2, c-sis, c-fes). These genes may thus be of importance for selection of differentiation pathway. The c-myc oncogene is frequently amplified in small-cell lung cancer cell lines in a much higher frequency than in vivo. This indicates that c-myc seems to be related to tumor progression and a relatively late event in the lung cancer development. The uncontrolled production of multiple growth factors has been identified in human lung cancer cell lines. These factors can promote and inhibit the proliferation via paracrine and autocrine loops via specific receptors. The products from some of the activated proto-oncogenes (c-sis, c-erbB-2) are sequences homologous to a certain growth factor (PDGF) and a receptor (EGF) identified in lung cancer. The production and action of these growth factors may be of major importance for further activation of proto-oncogenes via intracellular signal transduction and specific oncogenic activation leading to further tumor progression. Publication Types: Review Review, Tutorial PMID: 2174659 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 918: J Cell Physiol. 1990 Dec;145(3):564-74. Suppression of programmed death and G1 arrest in B-cell hybridomas by interleukin-6 is not accompanied by altered expression of immediate early response genes. Sabourin LA, Hawley RG. Department of Experimental Oncology, Ottawa Regional Cancer Centre, University of Ottawa, Ontario, Canada. The murine B-cell hybridoma B9 requires interleukin-6 (IL-6) for its survival and proliferation in vitro. We show here that withdrawal of IL-6 from B9 cultures results in programmed death, concomitant with arrest of the cells in the G1 phase of the cell cycle. Unlike several other systems that undergo programmed cell death, no induction of transcripts corresponding to the testosterone-repressed message-2 or transglutaminase genes is observed during this process. Upon readdition of IL-6 to G1-arrested B9 cells, viability is maintained and entry into S phase occurs after a lag period of 10 to 12 hr. Northern blot analysis showed that the immediate-early mRNAs normally induced shortly after growth factor stimulation in quiescent fibroblasts (c-fos, c-jun, Egr-1, c-myc, JE, and KC), and other growth-related genes (2F1, c-Ha-ras, and p53), are either not induced or remain unchanged during G1 to S phase progression. A correlation was found, however, between the temporal pattern of expression of several G1/S phase genes (dihydrofolate reductase, thymidine kinase, transferrin receptor, and histone H3) and DNA synthesis. These results demonstrate that IL-6-induced viability and growth of hybridoma (and, presumably, plasmacytoma) cells is mediated via novel signal transduction pathways. PMID: 1703172 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 919: Cancer Res. 1990 Nov 15;50(22):7324-32. Differential response of nontumorigenic and tumorigenic human papillomavirus type 16-positive epithelial cells to transforming growth factor beta 1. Braun L, Durst M, Mikumo R, Gruppuso P. Department of Pathology and Laboratory Medicine, Brown University, Providence, Rhode Island 02912. The transforming growth factor (TGF) beta s are multifunctional polypeptide growth factors with diverse biological effects, including inhibition of epithelial cell proliferation both in vitro and in vivo. To investigate the possible role of TGF beta 1 in the regulation of papillomavirus infection and papillomavirus-associated transformation, we compared the response to TGF beta 1 of normal keratinocytes, human papillomavirus, type 16 (HPV 16)-positive-immortalized keratinocytes (nontumorigenic), and HPV 16-positive cervical carcinoma cells (tumorigenic) with respect to DNA synthesis and protooncogene expression. All HPV 16-immortalized cell lines were nearly as inhibited by TGF beta 1 as normal keratinocytes, whereas two cervical carcinoma cell lines (Caski and Siha) were refractory to growth inhibition by TGF beta 1. Cell surface receptors for TGF beta 1 were present on both normal and carcinoma cell lines. In all cases, growth inhibition by TGF beta 1 was accompanied by suppression of Steady-state levels of c-myc mRNA. In contrast, TGF beta 1 induced the expression of c-jun mRNA transcripts in normal, immortalized, and tumorigenic cells. We also studied the effect of TGF beta 1 on HPV 16 mRNA expression. Steady-state levels of HPV 16 mRNA transcripts were suppressed by TGF beta 1 in the nontumorigenic HPK cells but were unaffected in the tumorigenic lines. These findings suggest that TGF beta 1 may be an in vivo modulator of HPV infection and that loss of responsiveness to this growth inhibitory signal may be involved in HPV-associated malignant transformation. PMID: 2171761 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 920: Mol Cell Biol. 1990 Nov;10(11):5914-20. An amino-terminal c-myc domain required for neoplastic transformation activates transcription. Kato GJ, Barrett J, Villa-Garcia M, Dang CV. Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205. The product of the c-myc proto-oncogene is a nuclear phosphoprotein whose normal cellular function has not yet been defined. c-Myc has a number of biochemical properties, however, that suggest that it may function as a potential regulator of gene transcription. Specifically, it is a nuclear DNA-binding protein with a short half-life, a high proline content, segments that are rich in glutamine and acidic residues, and a carboxyl-terminal oligomerization domain containing the leucine zipper and helix-loop-helix motifs that serve as oligomerization domains in known regulators of transcription, such as C/EBP, Jun, Fos, GCN4, MyoD, E12, and E47. In an effort to establish that c-Myc might regulate transcription in vivo, we sought to determine whether regions of the c-Myc protein could activate transcription in an in vitro system. We report here that fusion proteins in which segments of human c-Myc are linked to the DNA-binding domain of the yeast transcriptional activator GAL4 can activate transcription from a reporter gene linked to GAL4-binding sites. Three independent activation regions are located between amino acids 1 and 143, a region that has been shown to be required for neoplastic transformation of primary rat embryo cells in cooperation with a mutated ras gene. These results demonstrate that domains of the c-Myc protein can function to regulate transcription in a model system and suggest that alterations of Myc transcriptional regulatory function may lead to neoplastic transformation. PMID: 2233723 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 921: J Invest Dermatol. 1990 Nov;95(5):7S-9S. Growth factor and proto-oncogene expression in psoriasis. Elder JT, Klein SB, Tavakkol A, Fisher GJ, Nickoloff BJ, Voorhees JJ. Department of Dermatology, University of Michigan, Ann Arbor 48109-0672. The expression of several proto-oncogenes and growth factors was analyzed in normal skin and psoriatic lesions by RNA blot hybridization. Isolation of intact RNA from frozen biopsy samples required immediate exposure to denaturants during tissue homogenization. Lipocortin II and cyclophilin transcripts were used as internal controls. These transcripts were abundant and slightly but significantly elevated in psoriatic lesions. When results were normalized according to these reference transcripts, there was no increase in the expression of c-myc, c-Ha-ras, c-erbB (EGF receptor), c-jun, or transforming growth factor-beta (TGF-beta) transcripts in psoriatic lesions, and lesional c-fos transcripts were decreased relative to normal skin. In contrast, expression of TGF-alpha mRNA transcripts were markedly increased in psoriatic lesions even after normalization. Placement of normal or psoriatic tissue in organ culture for 2 to 4 h resulted in strong induction of c-fos, c-jun, and c-myc transcripts, but not of the other genes studied. Thus, overexpression of proto-oncogenes may be more characteristic of the epidermal response to acute injury than of the steady-state hyperplasia characteristic of psoriasis. Interferon-gamma (IFN-gamma) increased TGF-alpha mRNA levels in cultured human KC at long time intervals (24-48 h). However, of various cytokines tested, only EGF and TGF-alpha induced TGF-alpha mRNA after short time intervals (2-4 h). These results as well as the selective overabundance of TGF-alpha mRNA in psoriatic lesions among various cytokines tested suggest that activation of the EGF receptor tyrosine kinase by TGF-alpha is important in the pathogenesis of psoriatic epidermal hyperplasia. Publication Types: Review Review, Tutorial PMID: 2230227 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 922: Cancer Res. 1990 Nov 1;50(21):7023-30. Induction of replicative competence ("priming") in normal liver. Mead JE, Braun L, Martin DA, Fausto N. Department of Pathology and Laboratory Medicine, Brown University, Providence, Rhode Island 02912. We have used a system of nutritional manipulation to investigate whether hepatocytes of the normal liver can be primed for replication in vivo. In this system, rats that are denied protein for 3 days undergo a burst of hepatic DNA synthesis and mitosis when they are refed amino acids, while normally fed or starved rats do not respond. To determine if hepatocytes of protein deprived (PD) rats have been "primed" for replication, we examined changes in protooncogene expression in livers of PD rats to see if they would mimic the pattern of gene expression that is induced early after partial hepatectomy. c-jun, c-myc, and p53 mRNAs were elevated in livers of PD rats, while c-fos and c-ras genes were not expressed. The administration of amino acids to PD rats stimulated hepatic DNA synthesis in a shorter period than is required after partial hepatectomy and induced p53 and c-ras expression. In culture, hepatocytes from PD rats had higher levels of c-myc mRNA, underwent morphological changes more rapidly, and reached maximum rates of DNA synthesis earlier than normal hepatocytes. In both normal and primed hepatocyte cultures, transforming growth factor alpha stimulated DNA synthesis more effectively than epidermal growth factor. We conclude that hepatocytes pass through a priming stage before they proliferate and that replicative competence without DNA synthesis can be induced in hepatocytes in the normal liver. PMID: 2208169 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 923: Mol Cell Biol. 1990 Nov;10(11):5814-21. Deprivation of a single amino acid induces protein synthesis-dependent increases in c-jun, c-myc, and ornithine decarboxylase mRNAs in Chinese hamster ovary cells. Pohjanpelto P, Holtta E. Department of Virology, University of Helsinki, Finland. Genes of higher eucaryotic cells are considered to show only a limited response to nutritional stress. Here we show, however, that omission of a single essential amino acid from the medium caused a marked rise in the mRNA levels of c-myc, c-jun, junB and c-fos oncogenes and ornithine decarboxylase (ODC) in CHO cells. There was no general accumulation of mRNAs in amino acid-starved cells, since the gamma-actin, beta-tubulin, protein kinase C, RNA polymerase II, and glyceraldehyde-3-phosphate dehydrogenase mRNAs and the total poly(A)+ mRNA were not increased. The levels of c-myc, ODC, and c-jun mRNAs were elevated more by amino acid starvation than by inhibition of protein synthesis with cycloheximide, which is known to increase the levels of these mRNAs. Importantly, however, cycloheximide present during amino acid starvation reduced the rise in the levels of the mRNAs down to the level obtained with cycloheximide alone. This implies that protein synthesis is required for the accumulation of c-myc, ODC, and c-jun mRNAs in amino acid-deprived cells. The junB and c-fos mRNAs, instead, were increased to the same extent or less by amino acid starvation than by cycloheximide treatment. The accumulation of the c-myc mRNA in amino acid-starved cells was due to both stabilization of the mRNA and increase of its transcription. The rise in the c-jun mRNA level seemed to be caused merely by stabilization of the mRNA. Further, despite the inhibition of general protein synthesis, amino acid starvation led to an increase in the synthesis of c-myc polypeptide. The results suggest that mammalian cells have a specific mechanism for registering shortages of amino acids in order to make adjustments compatible with cellular growth. PMID: 2122233 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 924: Blood. 1990 Nov 1;76(9):1830-7. Amplified expression of three jun family members inhibits erythroleukemia differentiation. Prochownik EV, Smith MJ, Snyder K, Emeagwali D. Department of Pediatrics, University of Michigan School of Medicine, Ann Arbor. Several different proto-oncogenes have been shown to influence cellular differentiation. One of the most widely studied model systems has been the Friend murine erythroleukemia cell (F-MELC) line, which can be induced to undergo erythroid differentiation by a variety of chemical agents. Constitutive overexpression of either the c-myc or c-myb proto-oncogenes has been previously shown to inhibit F-MELC differentiation, whereas c-myc antisense sequences accelerate the process. To investigate the potential involvement of other proto-oncogenes and immediate early response genes in F-MELC differentiation, we studied the expression of the three known members of the jun family as well as another gene, egr-1, which, like the jun family members, is expressed as an immediate early response gene in growth factor-stimulated quiescent cells. All four genes were expressed in F-MELC, although the levels of expression and modes of regulation differed. Transfection with amplifiable c-jun, junB, or junD expression plasmids inhibited differentiation, whereas transfection with an egr-1 expression plasmid was without effect. These results indicate that jun family members play a role in mediating F-MELC differentiation. The known inhibitory effect of phorbol ester tumor promoters on F-MELC differentiation may be the result of their known stimulation of jun expression. PMID: 2121297 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 925: Am J Pathol. 1990 Oct;137(4):761-5. Early proto-oncogene expression in rat aortic smooth muscle cells following endothelial removal. Miano JM, Tota RR, Vlasic N, Danishefsky KJ, Stemerman MB. Department of Pathology, New York Medical College, Valhalla 10595. To study the mechanism(s) of vascular smooth muscle cell proliferation in vivo, mRNA levels of c-fos, c-jun, and c-myc were determined by Northern blot analysis following vascular balloon de-endothelialization (BDE). Medial smooth muscle cells (SMC) were separated and studied by enzymatic digestion of the vessel wall. mRNA levels of c-fos and c-jun from aortic smooth muscle cells (SMC) were simultaneously induced within 30 minutes of BDE and declined to baseline by 1.5 hours, c-myc mRNA did not begin to increase until 1 hour after vascular injury. Levels of c-myc peaked at 2 hours and were sustained for an additional 4 hours before gradually declining. Smooth muscle cells derived from enzyme-treated control aortae that did not undergo BDE expressed c-fos and c-jun, but showed no evidence of c-myc message. In contrast, nonenzymatically treated, non-BDE whole aortae (containing both media and adventitia) demonstrated a prominent c-myc signal, but failed to express c-fos and c-jun. Corresponding examination of adventitia derived from enzyme-treated aortae showed this tissue to be a source of all three proto-oncogenes. The results of this study demonstrate the earliest in vivo molecular markers of vascular injury reported to date and implicate SMC proto-oncogene expression in the initiation of SMC proliferation. Furthermore these findings suggest two avenues for proto-oncogene induction, that are due to (1) vessel wall manipulation and (2) humoral stimulation. PMID: 2221010 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 926: Mol Cell Biol. 1990 Oct;10(10):5333-9. The leucine zipper of c-Myc is required for full inhibition of erythroleukemia differentiation. Smith MJ, Charron-Prochownik DC, Prochownik EV. Department of Pediatrics, University of Michigan School of Medicine, Ann Arbor 48109. The leucine zipper motif has been observed in a number of proteins thought to function as eucaryotic transcription factors. Mutation of the leucine zipper interferes with protein dimerization and DNA binding. We examined the effect of point mutations in the leucine zipper of c-Myc on its ability to dimerize in vitro and to inhibit Friend murine erythroleukemia (F-MEL) differentiation. Glutaraldehyde cross-linking studies failed to provide evidence for homodimerization of in vitro-synthesized c-Myc protein, although it was readily demonstrated for c-Jun. Nevertheless, whereas transfected wild-type c-myc sequences strongly inhibited F-MEL differentiation, those with single or multiple mutations in the leucine zipper were only partially effective in this regard. Since the leucine zipper domain of c-Myc is essential for its cooperative effect in ras oncogene-mediated transformation, this study emphasizes the close relationship that exists between transformation and hematopoietic commitment and differentiation. c-Myc may produce its effects on F-MEL differentiation through leucine zipper-mediated heterodimeric associations rather than homodimeric ones. PMID: 2204813 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 927: Oncogene. 1990 Oct;5(10):1511-9. Differential regulation and expression of jun, c-fos and c-myc proto-oncogenes during mouse liver regeneration and after inhibition of protein synthesis. Morello D, Lavenu A, Babinet C. Unite de Genetique des Mammiferes, Paris, France. In order to obtain information in vivo about the possible relationships between early response gene products, we have analysed the expression of c-myc, c-fos and jun proto-oncogenes in regenerating mouse liver. We show that c-myc, c-fos, jun B, c-jun and jun D mRNA expression is transiently increased soon after partial hepatectomy, jun and fos expression being induced earlier (30 min) than that of c-myc (1-2 h). C-fos, jun B and c-jun mRNA expression is dramatically enhanced (50 fold) while that of jun D and c-myc is weaker (less than 10 fold), but lasts longer. Moreover, the relative contributions of transcriptional and post-transcriptional regulations are unique for each proto-oncogene analysed. These results suggest that following the growth signal delivered by partial hepatectomy, the five proto-oncogenes analysed are all involved in the progression of hepatocytes through G1; however, due to their differential regulation and kinetics, they might play different roles in the changes in gene expression that occur during the transition from quiescence to proliferation. When protein synthesis is inhibited by injection of cycloheximide, the expression of c-myc, c-fos, jun B, c-jun and jun D mRNA is also transiently increased. Although this increase is mainly due to post-transcriptional mechanisms, c-myc, c-jun, jun D and, to a lesser extent, jun B transcription is enhanced, suggesting that labile repressor-like molecules may inhibit transcription of these genes in the quiescent liver. Moreover, the kinetics of c-myc, c-fos and jun mRNA induction are not identical, showing that different components are involved in their turnover or stability. PMID: 2123531 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 928: Mol Cell Biol. 1990 Oct;10(10):5536-40. Induction of the proto-oncogene c-jun by angiotensin II. Naftilan AJ, Gilliland GK, Eldridge CS, Kraft AS. Hypertension Program, University of Alabama, Birmingham 35294. Angiotensin (Ang) II causes hypertrophy of rat aortic smooth muscle cells in culture and results in the rapid activation of c-fos. This study demonstrated that Ang II also activated c-jun and, in addition, could activate the AP-1 enhancer element. These data add support for a role of Ang II as an important mediator of vascular smooth muscle cell growth. PMID: 2119001 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 929: Mol Cell Biol. 1990 Oct;10(10):5314-23. Effect of a dominant inhibitory Ha-ras mutation on mitogenic signal transduction in NIH 3T3 cells. Cai H, Szeberenyi J, Cooper GM. Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115. We used a dominant inhibitory mutation of c-Ha-ras which changes Ser-17 to Asn-17 in the gene product p21 [p21(Asn-17)Ha-ras] to investigate ras function in mitogenic signal transduction. An NIH 3T3 cell line [NIH(M17)] was isolated that displayed inducible expression of the mutant Ha-ras gene (Ha-ras Asn-17) via the mouse mammary tumor virus long terminal repeat and was growth inhibited by dexamethasone. The effect of dexamethasone induction on response of quiescent NIH(M17) cells to mitogens was then analyzed. Stimulation of DNA synthesis by epidermal growth factor (EGF) and 12-O-tetradecanoylphorbol-13-acetate (TPA) was completely blocked by p21(Asn-17) expression, and stimulation by serum, fibroblast growth factor, and platelet-derived growth factor was partially inhibited. However, the induction of fos, jun, and myc by EGF and TPA was not significantly inhibited in this cell line. An effect of p21(Asn-17) on fos induction was, however, demonstrated in transient expression assays in which quiescent NIH 3T3 cells were cotransfected with a fos-cat receptor plasmid plus a Ha-ras Asn-17 expression vector. In this assay, p21(Asn-17) inhibited chloramphenicol acetyltransferase expression induced by EGF and other growth factors. In contrast to its effect on DNA synthesis, however, Ha-ras Asn-17 expression did not inhibit fos-cat expression induced by TPA. Conversely, downregulation of protein kinase C did not inhibit fos-cat induction by activated ras or other oncogenes. These results suggest that ras proteins are involved in at least two parallel mitogenic signal transduction pathways, one of which is independent of protein kinase C. Although either pathway alone appears to be sufficient to induce fos, both appear to be necessary to induce the full mitogenic response. PMID: 2118993 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 930: J Cell Physiol. 1990 Oct;145(1):46-52. Butyrate blocks the accumulation of CDC2 mRNA in late G1 phase but inhibits both the early and late G1 progression in chemically transformed mouse fibroblasts BP-A31. Charollais RH, Buquet C, Mester J. INSERM U55, Hopital St. Antoine, Paris, France. Sodium butyrate (6 mM) blocks the resumption of the cell division cycle in serum-deprived chemically transformed Balb/c-3T3 mouse fibroblasts (BP-A31). The inhibition of G1 progression by sodium butyrate is not restricted to a specific mitogenic signaling pathway and is equally effective when tetradecanoyl phorbol acetate (TPA), insulin, or fetal calf serum (FCS) is used as inducer. The inhibitor acts in early as well as late G1 phase as indicated by experiments in which inhibitor was added and withdrawn at different times after restimulation of quiescent cells by FCS. At the gene expression level, sodium butyrate does not affect the inducibility of early cell cycle-related genes (c-myc, c-jun) while blocking the induction of cdc 2 mRNA, a late G1 marker. We conclude that sodium butyrate does not interfere with the growth factor signaling pathways regulating the (early) cell cycle-related gene expression. However, the presence of sodium butyrate early in G1 phase inhibits the cascade of events leading eventually to the expression of late G1-characteristic genes such as cdc2. The antimitogenic activity of sodium butyrate may be related to its interference with an (unknown) process involved in the "mitogenic" cascade. PMID: 1976640 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 931: Gastroenterol Jpn. 1990 Sep;25 Suppl 2:43-8. Integration of hepatitis virus DNA near c-myc in woodchuck hepatocellular carcinoma. Hsu TY, Fourel G, Etiemble J, Tiollais P, Buendia MA. Unite de Recombinaison et Expression Genetique (INSERM U.163, Institut Pasteur, Paris, France. A total of 33 hepatocellular carcinomas, induced in woodchucks by chronic infection with woodchuck hepatitis virus (WHV), a virus closely related to the human hepatitis B virus, were analyzed for the state of viral DNA, the expression of viral genes and of different cellular proto-oncogenes. Low levels of viral replication and presence of integrated viral forms including sequences of the enhancer element, appeared as a general rule in these tumors. Enhanced expression of one or more of the nuclear protooncogenes: c-myc, N-myc, c-fos, c-jun and jun-B was frequently observed. In two hepatomas, elevated expression and allelic alterations of c-myc were subsequent to integration of WHV DNA near the c-myc coding domain. The viral strategy for insertional activation of c-myc in these tumors appeared basically identical to that of mammalian retroviruses in T-cell lymphomas of mice and rats. Whether insertional mutagenesis of different oncogenes may be more generally linked to liver oncogenesis induced by WHV and hepatitis B viruses remains to be determined. PMID: 2172071 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 932: Cancer Lett. 1990 Sep;53(2-3):205-12. Increased expression of c-jun gene during spontaneous hepatocarcinogenesis in LEC rats. Suzuki H, Fujita H, Mullauer L, Kuzumaki N, Konaka S, Togashi Y, Takeichi N, Kawamukai Y, Uchino J. Laboratory of Molecular Genetics, University School of Medicine, Sapporo, Japan. We have studied the expressions of nine proto-oncogenes (c-myc, N-myc, c-fos, C-jun, p53, H-ras, N-ras, c-raf, hst) and two other genes (PCNA, GST-P) during the spontaneous development of hepatocellular carcinomas (HCCs) in LEC rats. Expression of c-myc, H-ras, N-ras, C-raf, p53 and PCNA genes was detected, but this did not significantly change during the development of HCCs in LEC rats. Expression of N-myc and hst genes was not detectable. Expression of c-fos gene was detected in one HCC case out of four. Significantly increased expression of c-jun gene was observed in the liver tissues of LEC rats aged 8 months. This high expression was decreased with the development of HCCs. On the other hand, the expression of GST-P gene increased in parallel with the clinical course of the development of HCCs in LEC rats. The pattern of c-jun mRNA augmentation was different from that of GST-P mRNA. These observations suggest that c-jun gene may play a role in the spontaneous development of HCCs in LEC rats. PMID: 1976434 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 933: Biochem Biophys Res Commun. 1990 Aug 31;171(1):287-92. Activation of 'immediate-early' genes by estrogen is not sufficient to achieve stimulation of DNA synthesis in rat uterus. Persico E, Scalona M, Cicatiello L, Sica V, Bresciani F, Weisz A. Istituto di Patologia Generale e Oncologia, Prima Facolta di Medicina e Chirurgia, Universita di Napoli, Italy. 17 beta-estradiol, a long acting estrogen that is mitogenic for rat uterus in vivo, or the short acting estrogens estriol and 16 alpha-estradiol, not mitogenic on their own, were injected into adult, castrated rats and their effect on uterine gene expression and rate of DNA synthesis were compared. All three compounds increased steady-state mRNA concentration of c-fos, c-jun and c-myc proto-oncogenes to comparable levels (2 hrs after treatment), whereas only 17 beta-estradiol was found to stimulate significantly DNA synthesis (20-22 hrs later). Based on the different retention time of the tested estrogens in rat tissues, it is concluded that a short exposure to the hormone is sufficient to render uterine cells competent to progress through the cell cycle, via activation of 'immediate-early' genes expression, but that stimulation of DNA synthesis requires further changes, achieved via a prolonged exposure of the cells to the estrogenic stimulus. PMID: 2118345 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 934: J Biol Chem. 1990 Aug 25;265(24):14061-4. Cadmium induces transcription of proto-oncogenes c-jun and c-myc in rat L6 myoblasts. Jin P, Ringertz NR. Department of Medical Cell Genetics, Medical Nobel Institute, Karolinska Institutet, Stockholm, Sweden. The effect of cadmium on the expression of oncogenes c-jun, c-fos, and c-myc was studied by exposing L6J1 myoblast cultures to different doses of cadmium chloride and then analyzing the abundance of oncogene transcripts. Cadmium induced a transient accumulation of c-jun and c-myc mRNA with maximum expression at 2-4 h. At the same time, the level of c-fos transcripts remained below the detection level. Both the c-fos and c-jun genes could. However, be induced by treating the myoblasts with insulin. Cadmium induction of c-jun and c-myc mRNA occurred in a concentration-dependent manner with maximum stimulation at 5-10 microM. In the presence of cycloheximide, c-jun and c-myc genes were superinduced by the addition of cadmium. Under these conditions there was also a marked increase in c-fos transcripts. Induction of c-myc and c-jun by cadmium and c-fos by a combination of cadmium and cycloheximide could be abolished by blocking transcription with actinomycin D. The cadmium-induced increase in c-jun and c-myc mRNA was enhanced in myoblasts stably transfected with a mouse c-fos gene under a metallothionein promoter. Our present data suggest that cadmium has the potential to deregulate the expression of several important oncogenes. PMID: 1696944 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 935: Cell Growth Differ. 1990 Aug;1(8):351-9. Regulation of the epidermal growth factor receptor gene expression in a morphological variant isolated from an epidermal growth factor receptor-deficient small cell lung carcinoma cell line. Gamou S, Shimosato Y, Shimizu N. Department of Molecular Biology, Keio University School of Medicine, Tokyo, Japan. Most cell lines derived from small cell lung carcinoma grow in an anchorage-independent manner; they neither possess epidermal growth factor binding activity nor express epidermal growth factor receptor (EGFR) mRNA. A variant AD320, which grew in an anchorage-dependent manner with altered morphology, was isolated from the small cell lung carcinoma cell line Lu134 by treatment with the demethylating agent 5-azacytidine. The analysis, using methylation-sensitive restriction enzymes, revealed that the methylation pattern was altered only in the structural region of the EGFR gene; EGFR mRNA and epidermal growth factor binding activity could be detected in the variant. In addition, drastic changes in gene expression including a decrease of creatine kinase B mRNA and an increase of c-myc mRNA were observed. The EGFR in the variant appeared to be an active part of the transmembrane signaling machinery since c-fos and c-jun mRNA accumulated after epidermal growth factor treatment, followed by EGFR and c-myc mRNA accumulation. A potent tumor promoter, 12-O-tetradecanoylphorbol-13-acetate, also induced EGFR mRNA. Thus, the inducible regulatory mechanism for the EGFR gene was activated in the variant even though the EGFR gene was constitutively expressed. PMID: 1980602 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 936: Hematol Oncol. 1990 Jul-Aug;8(4):191-204. Heterogeneous expression of proto-oncogenes in Hodgkin's disease derived cell lines. Jucker M, Schaadt M, Diehl V, Poppema S, Jones D, Tesch H. Genetisches Labor Med. Klinik I, Universitat Koln, Cologne, FRG. The expression of 20 proto-oncogenes was analysed by Northern blotting in four cell lines derived from patients with Hodgkin's disease (L428, L540, CO and DEV) and compared to lymphoid and myeloid leukemia cell lines and normal hematopoietic cells. Expression of the proto-oncogenes c-myc, p53, c-jun, pim-1, lck, c-syn, c-raf and N-ras were detected in Hodgkin's disease derived cell lines and in normal hematopoietic cells. Transcripts of the proto-oncogene c-met were detected in the Hodgkin's derived cell lines L428 and L540 but not in the lymphoid or myeloid leukemia cell lines or in tonsil cells, peripheral blood mononuclear cells and granulocytes. Expression of the proto-oncogenes N-myc and lck were observed in the Hodgkin's derived cell line CO which express T cell receptor genes and in the T cell lines JM and CEM. L428 cells and CO cells expressed aberrant transcripts of the c-fes proto-oncogene. Thus Hodgkin's disease derived cell lines are heterogeneous in their expression pattern of proto-oncogenes, expressing normal and aberrant transcripts of proto-oncogenes which are not found in untransformed hematopoietic cells. PMID: 2210688 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 937: Mol Endocrinol. 1990 Jul;4(7):1041-50. Estrogen stimulates transcription of c-jun protooncogene. Weisz A, Cicatiello L, Persico E, Scalona M, Bresciani F. Istituto di Patologia Generale e Oncologia, Prima Facolta di Medicina e Chirurgia, Universita di Napoli, Italy. Estrogen is a mitogen for the rat uterus, where it induces transient activation of c-fos and c-myc protooncogene expression, followed by increases in DNA synthesis and cell proliferation. JUN-C, the product of the c-jun protooncogene, is a nuclear protein that can interact with FOS to modulate the activity of AP-1-responsive promoters. To test whether c-jun is a target for estrogen regulation, we measured the effects of 17 beta-estradiol on the expression of this gene in rat uterus. A human c-jun cDNA probe detects in rat uterus two mRNA species of 2.5 and 3.2 kilobases. Treatment of the animals with estrogen results in a rapid transient increase in the concentrations of these mRNAs; a 4- to 5-fold increase over the prestimulation level was detected starting 30 min after estrogen injection and lasting for 2 h, with a return to the prestimulation level after 4 h. In accordance with the results obtained by analysis of the mRNA, we found that estrogen increases 3- to 4-fold c-jun gene transcription in the uterus, at the same time it induces its mRNA accumulation. The ability of estrogen to induce c-jun gene expression was not abolished by the protein synthesis inhibitor cycloheximide, suggesting that transcriptional activation of this protooncogene is a primary response to the hormone. Furthermore, we found that in the estrogen-responsive MCF-7 human mammary carcinoma cells, estrogen stimulates transcription of a reporter gene containing four copies of a jun/AP-1 response element. These data demonstrate that c-jun gene expression is regulated by estrogen and suggest that JUN-C could play a role in the activation of cell proliferation by estrogen. PMID: 2126598 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 938: J Immunol. 1990 Jun 15;144(12):4835-40. The expression of c-fos, c-jun, and c-myc genes is regulated by heat shock in human lymphoid cells. Bukh A, Martinez-Valdez H, Freedman SJ, Freedman MH, Cohen A. Division of Immunology/Rheumatology, Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada. The effect of heat shock on the expression of the nuclear protooncogenes c-fos, c-jun, and c-myc was studied in human lymphoid cells. Heat shock caused an increase in c-fos and c-jun mRNA levels and a decrease in c-myc mRNA levels in pre-B (Hyon) and T (DND-41) cell lines as well as in freshly isolated normal human thymocytes. The changes in the mRNA levels of these protooncogenes in Hyon cells were most pronounced at 42 and 43 degrees C; kinetic analysis demonstrated that the changes could be detected within 30 min of heat shock. Altered transcription of c-fos and c-myc genes was the primary effect of heat shock. Secondarily, heat shock of Hyon cells stabilized the c-myc mRNA level by increasing its half-life from 24 to 45 min. The overall effect of heat shock on c-myc mRNA level, however, was a marked inhibition of its transcription. These results demonstrate that the transcription of nuclear protooncogenes is regulated by heat shock indicating a role for nuclear protooncogenes in the stress response of lymphoid cells. PMID: 2112575 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 939: Mol Endocrinol. 1990 Jun;4(6):920-30. Erratum in: Mol Endocrinol 1990 Jul;4(7):1016. Multiple adenosine 3',5'-cyclic [corrected] monophosphate response element DNA-binding proteins generated by gene diversification and alternative exon splicing. Hoeffler JP, Meyer TE, Waeber G, Habener JF. Massachusetts General Hospital, Howard Hughes Medical Institute, Harvard Medical School, Boston 02114. The protein sequence deduced from the open reading frame of a human placental cDNA encoding a cAMP-responsive enhancer (CRE)-binding protein (CREB-327) has structural features characteristic of several other transcriptional transactivator proteins including jun, fos, C/EBP, myc, and CRE-BP1. Results of Southwestern analysis of nuclear extracts from several different cell lines show that there are multiple CRE-binding proteins, which vary in size in cell lines derived from different tissues and animal species. To examine the molecular diversity of CREB-327 and related proteins at the nucleic acid level, we used labeled cDNAs from human placenta that encode two different CRE-binding proteins (CREB-327 and CRE-BP1) to probe Northern and Southern blots. Both probes hybridized to multiple fragments on Southern blots of genomic DNA from various species. Alternatively, when a human placental c-jun probe was hybridized to the same blot, a single fragment was detected in most cases, consistent with the intronless nature of the human c-jun gene. The CREB-327 probe hybridized to multiple mRNAs, derived from human placenta, ranging in size from 2-9 kilobases. In contrast, the CRE-BP1 probe identified a single 4-kilobase mRNA. Sequence analyses of several overlapping human genomic cosmid clones containing CREB-327 sequences in conjunction with polymerase chain reaction indicates that the CREB-327/341 cDNAs are composed of at least eight or nine exons, and analyses of human placental cDNAs provide direct evidence for at least one alternatively spliced exon. Analyses of mouse/hamster-human hybridoma DNAs by Southern blotting and polymerase chain reaction localizes the CREB-327/341 gene to human chromosome 2. The results indicate that there is a dichotomy of CREB-like proteins, those that are related by overall structure and DNA-binding specificity as well as those that are related by close similarities of primary sequences. PMID: 2146494 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 940: Mol Cell Biol. 1990 Jun;10(6):3185-93. c-myc, c-fos, and c-jun regulation in the regenerating livers of normal and H-2K/c-myc transgenic mice. Morello D, Fitzgerald MJ, Babinet C, Fausto N. Department of Immunology, Institut Pasteur, Paris, France. We investigated the mechanisms of regulation of c-myc, c-fos, and c-jun at the early stages of liver regeneration in mice. We show that the transient increase in steady-state levels of c-myc mRNA at the start of liver regeneration is most probably regulated by posttranscriptional mechanisms. Although there was a marked increase in c-myc transcriptional initiation shortly after partial hepatectomy, a block in elongation prevented the completion of most transcripts. To gain further information on the mechanism of regulation of c-myc expression during liver regeneration, we used transgenic mice harboring the human c-myc gene driven by the H-2K promoter. In these animals, the murine c-myc responded to the growth stimulus generated by partial hepatectomy, whereas the expression of the transgene was constitutive and did not change in the regenerating liver. However, the mRNA from both genes increased markedly after cycloheximide injection, suggesting that the regulation of c-myc mRNA abundance in the regenerating liver differs from that occurring after protein synthesis inhibition. Furthermore, we show that in normal mice c-fos and c-jun mRNA levels and transcriptional rates increase within 30 min after partial hepatectomy. c-fos transcriptional elongation was restricted in nongrowing liver, but the block was partially relieved in the regenerating liver. Nevertheless, for both c-fos and c-jun, changes in steady-state mRNA detected after partial hepatectomy were much greater than the transcriptional increase. In the regenerating liver of H-2K/c-myc mice, c-fos and c-jun expression was diminished, whereas mouse c-myc expression was enhanced in comparison with that in nontransgenic animals. PMID: 2111449 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 941: Oncogene. 1990 May;5(5):683-9. The leucine zipper domain of avian cMyc is required for transformation and autoregulation. Crouch DH, Lang C, Gillespie DA. Beatson Institute for Cancer Research, Bearsden, Glasgow, UK. Small deletions of 7 to 48 amino acids have been generated in the leucine zipper domain of the avian cMyc protein and the mutant cMyc proteins expressed using an avian retroviral vector. Retrovirally encoded cMyc protein transforms primary chick embryo fibroblasts and leads to abnormal regulation of the endogenous c-myc gene. Deletion of the most C-terminal leucine of the zipper motif confers a partial phenotype affecting some but not all parameters of transformation. Complete loss of transforming activity results from deletion of further leucine residues, including one which is not part of the heptad repeat. In cMyc transformed cells endogenous c-myc mRNA is expressed at a low level and is abnormally refractory to induction by serum stimulation. In contrast, a non-transforming cMyc protein which lacks the zipper does not affect normal c-myc expression. These results demonstrate that the leucine zipper domain of avian cMyc is required for both transformation and autoregulation, and suggests that essential leucine residues within the motif may be spaced differently from those in the zippers of Fos and Jun. PMID: 2189105 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 942: J Virol. 1990 May;64(5):2193-201. Transcriptional regulation of early-response genes during polyomavirus infection. Glenn GM, Eckhart W. Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92138-9216. Infection of quiescent BALB/c 3T3 cells with polyomavirus leads to the biphasic accumulation of RNA from several early-response genes. The steady-state levels of RNA of c-fos, c-myc, and c-jun were detected at 0 to 1 and 12 to 30 h after infection but not at 6 h postinfection. Infections with mutant viruses suggest that in the context of a virus infection with other tumor (T) antigens present, large T and middle T antigens are dispensable for the effect and small t antigen is important for the regulation, perhaps in conjunction with large T or middle T antigens. These results are in agreement with those of Zullo et al. (Proc. Natl. Acad. Sci. USA 84:1210-1214). We have further found that this regulation occurs in NIH 3T3 cells and primary mouse embryo fibroblasts. DNA synthesis is not required for this effect. Half-lives of c-fos, c-myc, and c-jun were similarly short at both early and late times after infection, as determined by dactinomycin chase. The regulation of expression occurs at the transcriptional level. Nuclear run-on experiments showed increased rates of transcription both early and late after infection. Also, the polyomavirus early region can transactivate the c-fos promoter in transient transfection assays. PMID: 1691311 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 943: Immunology. 1990 Mar;69(3):490-3. Patterns of nuclear proto-oncogene expression during induced differentiation and proliferation of human B chronic lymphocytic leukaemia cells. Murphy JJ, Tracz M, Norton JD. Department of Haematology, Royal Free Hospital School of Medicine, Hampstead, London. Phorbol ester-induced differentiation of human B-chronic lymphocytic leukaemic cells was found to be preceded by a rapid transient induction in expression of the c-jun proto-oncogene, which paralleled that of c-fos. Induced expression of c-myc but not of c-fos/c-jun proto-oncogenes was markedly higher in a proliferating variant leukaemic cell population compared with that seen in typical lymphocytic leukaemia cells. These data suggest that the c-fos/c-jun nuclear oncogenes play a role in induced differentiation, whilst c-myc is more important in the proliferative response of B lymphocytes. PMID: 2312171 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 944: Anticancer Res. 1990 Mar-Apr;10(2A):423-32. Retinoic acid inhibits human melanoma tumor cell invasion. Wood WR, Seftor EA, Lotan D, Nakajima M, Misiorowski RL, Seftor RE, Lotan R, Hendrix MJ. Department of Anatomy, College of Medicine, University of Arizona, Tucson 85724. The anticancer effects of retinoids have been recognized both in vivo and in vitro; however, little is known about their mechanism of action. Our study evaluated the effects of retinoic acid on the invasiveness of four human melanoma cell lines in vitro and showed a time-dependent inhibition of the ability of these cells to penetrate matrigel-coated filters. The possible mechanisms of action responsible for the anti-invasive effect were further investigated, and the data showed that retinoic acid-treated cells: (a) secreted lower levels of collagenolytic enzymes detected in type IV collagen-containing polyacrylamide gels compared with control cells, which was demonstrated by a decreased ability to degrade [3H]proline-labeled type IV collagen substrate; (b) showed a reduction in PA activity, primarily in the form of tPA, as demonstrated by chromogenic analysis; (c) showed a heterogeneous response with regard to c-myc, c-fos and c-jun mRNA expression, as determined by Northern blot analysis; and (d) demonstrated a decrease in B-actin levels and an increase in vimentin, as demonstrated by Northern blot analysis and SDS-PAGE transblot analysis. Collectively, these data suggest that RA causes an inhibitory effect on tumor cell invasion through a reconstituted basement basement membrane matrix by suppressing type IV collagenolytic activity and PA activity, which is probably triggered through a complex series of oncogene trans-acting factors, ultimately affecting cytoskeletal expression. PMID: 2161200 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 945: Cell Growth Differ. 1990 Mar;1(3):113-7. Transcriptional down-regulation of c-myc expression by protein synthesis-dependent and -independent pathways in a human T lymphoblastic tumor cell line. Bading H, Moelling K. Max-Planck-Institut fuer Molekulare Genetik, Berlin, Federal Republic of Germany. We show that in the human T lymphoblastic tumor cell line Molt4 c-myc mRNA and protein expression is down-regulated after exposure to dimethyl sulfoxide, to phorbol myristate acetate, or to the calcium ionophore A23187, which raises the intracellular calcium concentration. A block to RNA elongation is largely responsible for decreased c-myc transcription. Although negative regulation by dimethyl sulfoxide takes place even when protein synthesis is inhibited by cycloheximide, the phorbol myristate acetate effect is blocked to some extent only by cycloheximide. The calcium ionophore-induced c-myc suppression, however, strictly requires de novo protein synthesis. Therefore, two different negative regulatory pathways are involved in c-myc regulation: one which is independent and one which depends on de novo protein synthesis. The latter one appears to be mediated by a rapidly calcium-dependent induced gene product. PMID: 2127692 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 946: Hokkaido Igaku Zasshi. 1990 Mar;65(2):201-9. [A study on expression of various oncogenes and tumor-associated genes in LEC rats spontaneously developing hepatitis and hepatoma] [Article in Japanese] Suzuki H. First Department of Surgery, Hokkaido University, School of Medicine, Sapporo, Japan. The expression of nine proto-oncogenes (c-myc, N-myc, c-fos, c-jun, p53, H-ras, N-ras, c-raf, hst) and other three genes (AFP, PCNA, GST-P) were investigated during spontaneous development to hepatocellular carcinomas (HCCs) in LEC rats. Expressions of c-myc, H-ras, N-ras, c-raf, p53, and PCNA genes were detected but did not significantly change during the development to HCCs in LEC rats. Expressions of N-myc, hst, and AFP genes were not detectable since 5 weeks after birth. Expression of c-fos gene was detected in one out of four HCCs. Significantly increased expression of c-jun gene was observed in the liver tissues of LEC rats aged 8 months. The high expression was decreased in HCCs. On the other hand, the expression of GST-P gene increased in parallel with the clinical course of the development to HCCs in LEC rats. The increased expression of GST-P gene was observed in the liver tissues of LEC rats aged 8 months, and HCCs showed very high expression of GST-P gene. These observations suggest that both c-jun and GST-P genes may play a role in the spontaneous development to HCCs in LEC rats. PMID: 1694810 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 947: J Biol Chem. 1990 Feb 25;265(6):3284-92. Heparin-binding growth factor-1 stimulation of human endothelial cells induces platelet-derived growth factor A-chain gene expression. Gay CG, Winkles JA. Laboratory of Molecular Biology, Jerome H. Holland Laboratory for the Biomedical Sciences, American Red Cross, Rockville, Maryland 20855. Heparin-binding growth factor-1 (HBGF-1), also known as acidic fibroblast growth factor, is a potent mitogen for a variety of cell types including vascular endothelial and smooth muscle cells. Studies using murine 3T3 fibroblasts have shown that HBGF-1 induces numerous cellular responses such as the tyrosine phosphorylation of specific polypeptides and the increased expression of actin mRNA. Here we report that the addition of HBGF-1 to quiescent human umbilical vein endothelial cells increases the level of platelet-derived growth factor (PDGF) A-chain mRNA but not PDGF B-chain mRNA. In contrast, factors that inhibit endothelial cell proliferation such as phorbol myristate acetate and the cytokines interleukin-1, interleukin-6, and tumor necrosis factor-alpha increase both PDGF A-chain and B-chain mRNA levels. HBGF-1 induction of PDGF A-chain mRNA expression occurs in the presence of the protein synthesis inhibitor cycloheximide and thus does not require de novo protein synthesis. HBGF-1 also increases c-fos, c-jun, and c-myc mRNA levels; in the presence of cycloheximide, PDGF A-chain and protooncogene mRNA accumulation kinetics are similar. Nuclear run-on experiments indicate that the transcription rate of the PDGF A-chain gene transiently increases after HBGF-1 addition. Immunoprecipitation analysis using PDGF A-chain-specific antibodies indicates that HBGF-1-stimulated cells synthesize and secrete an increased amount of PDGF relative to unstimulated cells. If HBGF-1 can regulate PDGF expression by vascular endothelial cells in vivo, then HBGF-1 availability would be an important component of smooth muscle cell growth control. For example, HBGF-1 within the vessel wall would promote smooth muscle cell proliferation by (a) direct interaction with smooth muscle cell HBGF-1 receptors, and (b) increasing the amount of endothelial cell-derived PDGF available for binding to smooth muscle cell PDGF receptors. PMID: 1689299 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 948: Science. 1990 Feb 2;247(4942):561-4. Activation of proto-oncogenes: an immediate early event in human cytomegalovirus infection. Boldogh I, AbuBakar S, Albrecht T. Department of Microbiology, University of Texas Medical Branch, Galveston 77550. A rapid increase in the RNA levels of the proto-oncogenes c-fos, c-jun, and c-myc was detected after human cytomegalovirus infection. Neither inactivation of viral infectivity with ultraviolet irradiation (with or without psoralen), nor inhibition of translation with cycloheximide or anisomycin adversely affected the enhanced expression of proto-oncogenes, even though these treatments substantially reduced or eliminated the detection of immediate early viral antigens. The increase in the RNA levels of the proto-oncogenes was prevented in the presence of alpha-amanitin or actinomycin D. Thus, expression of these oncogenes appears to be induced by events occurring before the onset of viral protein synthesis, perhaps by the interaction of viral particles with the cell surface. PMID: 1689075 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 949: Oncogene. 1990 Feb;5(2):161-9. Mitogenesis induced by pp60v-src is not accompanied by increased expression of immediate early response genes. Welham MJ, Wyke JA, Lang A, Wyke AW. Imperial Cancer Research Fund Laboratories, St. Bartholomew's Hospital, Bartholomew Close, London, UK. Rat-1 cells infected with a temperature sensitive mutant of RSV (ts LA 29 Rat-1) can be rendered quiescent by serum deprivation at restrictive temperature. Shift to permissive conditions activates the v-src protein tyrosine kinase within 10 minutes and either this stimulus, or serum addition at restrictive temperature, leads to progression of the cell from G0 to G1, S-phase and mitosis. The effects of serum and temperature shift are not synergistic, suggesting that they may operate by convergent mechanisms. However, the characteristic serum-stimulated transient increases in transcripts of three immediate early response genes, c-fos, c-jun and c-myc are absent or much reduced when mitogenesis in ts LA 29 Rat-1 is induced by pp60v-src. Nonetheless, upon activating the pp60v-src protein kinase there is a marked and rapid increase in the ability of ts LA 29 Rat-1 nuclear extracts to retard the gel migration of oligonucleotides containing the AP-1 binding site, indicating that pp60v-src activity leads to an enhanced functioning of Fos and Jun related proteins that may, in turn, affect their transcriptional activation. Furthermore, these findings, and comparison with those of other laboratories, suggest that the mitogenic and transforming activities of pp60v-src have different effects on the transcription of immediate early response genes. PMID: 2108404 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 950: Oncogene. 1990 Jan;5(1):85-95. Charge configurations in oncogene products and transforming proteins. Karlin S, Brendel V. Department of Mathematics, Stanford University, California 94305. Statistically significant charge clusters are of infrequent occurrence in all kinds of proteins. In the six standard classes of proto-oncogene products, all of the nuclear class contain a significant charge cluster and several, but not all, of the transmembrane class do, whereas significant charge clusters or patterns are not found in protooncogenes of primarily cytoplasmic location, nor in membrane-bound (src-like) proto-oncogenes, nor in those of the ras family. Among nuclear oncogene families, such as myc, jun, fos, myb, or ets-related, and among homologous proteins across species, the significant charge clusters are part of the most conserved region. These gene families generally have similar charge distributions embodying a significant charge cluster, not of an invariant sign, preceded by a substantial uncharged stretch of predominantly polar residues. The nuclear transforming proteins p53 and p68 also contain significant charge clusters together with long uncharged segments, suggestive of a modular structure of these proteins. The transmembrane oncogene c-mas contains a mixed charge cluster and c-fms displays an unusual (0, +)7 pattern, in both cases positioned within their intracellular activating domain. Distinctive charge configurations for excreted proto-oncogenes are of a mixed character. Possible functions, mechanisms, and associated experimental procedures for studying proteins with anomalous charge distributions are discussed. PMID: 2181379 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 951: Life Sci. 1990;47(17):1555-60. Physiological cyclic AMP stimulation and oncogene mRNA levels in HL-60 cells. Moens U, Bang BE, Aarbakke J. Virological Research Group, University of Tromso, Norway. Altered gene expression of the proto-oncogenes c-fos and c-myc is associated with differentiation of the human promyelocytic leukemia (HL-60) cells. We studied changes of cyclic AMP levels and c-fos and c-myc mRNA levels after stimulation with theophylline and theophylline plus isoproterenol. Reduced c-fos and c-myc mRNA levels were detected, but the reduction could not, however, be related to the observed alternations in cyclic AMP concentrations. Expression of c-jun and c-Ha-ras was not affected under these conditions. PMID: 2174489 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 952: Mol Carcinog. 1990;3(5):302-8. Elevated expression of secondary, but not early, responding genes to phorbol ester tumor promoters in papillomas and carcinomas of mouse skin. Hashimoto Y, Tajima O, Hashiba H, Nose K, Kuroki T. Department of Cancer Cell Research, University of Tokyo, Japan. A single topical treatment of mouse skin with the potent tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) results in transient inductions of a variety of genes. Based on the time courses of their inductions, these genes can be classified into two main groups: "early" response genes whose mRNA expression reaches a maximum 0.5-2 h after TPA treatment and "secondary" response genes whose mRNA expression is maximal 4 h or more after treatment. The nuclear oncogenes c-fos, c-myc, and c-jun belong to the early response group, whereas the metallothionein, osteopontin, and urokinase genes belong to the secondary response group. The steady-state expressions of these early and secondary response genes are all very low in normal skin, except that of c-jun, which is relatively high. Steady-state levels of expression and inducibility of these genes by TPA were not altered in initiated skin or in apparently normal skin during tumor promotion. We examined the expressions of these genes in papillomas and carcinomas produced by two-stage (initiator-promoter) and three-stage (initiator-promoter-initiator) protocols in mouse skin. Steady-state expression of the early responding nuclear oncogenes in papillomas and carcinomas was found to remain at the same low level as in normal skin. However, all the secondary responding genes were found to be expressed constitutively at high levels in these tumors. Elevated expressions of the genes for transforming growth factor alpha and beta were also observed in papillomas and to varying extents in carcinomas. These observations suggest that the regulatory machinery for transcription by the protein kinase C-mediated pathway through nuclear oncogenes is altered during the processes of tumor promotion and progression. The genes whose expression is elevated may be associated directly or indirectly with tumor promotion and progression. PMID: 2123108 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 953: J Cardiovasc Pharmacol. 1990;16 Suppl 1:S1-3. Enhanced cell proliferation in essential hypertension. Baudouin-Legros M, Guicheney P, Meyer P. INSERM U7, Hopital Necker, Paris, France. In order to define the molecular mechanism involved in enhancement of spontaneously hypertensive rat (SHR) cell proliferation, we have compared the actions of fetal calf serum (FCS) and angiotensin II on both SHR and Wistar-Kyoto (WKY) rat aortic smooth muscle cells. Both compounds are more mitogenic in SHR cells than in controls. However, phospholipase C (PLC) hyperresponsiveness can be seen only under angiotensin stimulation, as are the expressions of c-jun, c-fos, and c-myc. Oncogene overexpression therefore appears to be more strongly related to PLC hyperreactivity than to enhanced proliferation of SHR aortic smooth muscle cells. PMID: 1706007 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 954: J Cardiovasc Pharmacol. 1990;15 Suppl 1:S1-6. Hypertension and atherosclerosis. Baudouin-Legros M, Meyer P. Department of Pharmacology, Hopital Necker, Paris, France. Membrane phospholipase C (PLC) activation is induced by the interaction of numerous vasoactive hormones and growth factors with their receptors. Two products are liberated: inositol triphosphate (IP3) and diacyglycerol (DG). The first product liberates intracellular calcium from its stores in the sarcoplasmic reticulum and the second one activates a phosphokinase, which triggers a transmembrane Na+/H+ exchange. A cascade of metabolic events secondary to these chemical changes impinges on the expression of nuclear proto-oncogenes, which determines cell growth. Studies conducted in spontaneously hypertensive rats (SHRs) have shown that PLC is hyperreactive to various agonists and that the phenomenon is present within a variety of cells, fibroblasts, platelets, and myocytes. Therefore, it is likely that hypertension in SHRs is characterized by a diffuse and intrinsic cellular defect that cannot be considered a consequence of the hemodynamic changes of hypertension. On the one hand, enhanced intracellular calcium mobilization may play a role in arterial tone and contraction whereas, on the other hand, enhanced activation of proto-oncogenes, myc, fos, and jun, may be involved in the mechanisms of arteriosclerosis. The pattern of an evolution towards arterial cell proliferation with acquisition of a secretory phenotype with collagen production was indeed observed in cultured cells from the arterial wall. Publication Types: Review Review, Tutorial PMID: 1695295 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 955: J Invest Dermatol. 1990 Jan;94(1):19-25. Protooncogene expression in normal and psoriatic skin. Elder JT, Tavakkol A, Klein SB, Zeigler ME, Wicha M, Voorhees JJ. Department of Dermatology, University of Michigan, Ann Arbor. The expression of the c-myc, c-fos, c-jun, c-erbB, and c-Ha-ras protooncogenes was compared by Northern blot analysis of total RNA extracted from keratome biopsies of normal skin and psoriatic plaques. Isolation of intact RNA from frozen tissue required careful attention to technique during the early stages of extraction. Densitometric analysis revealed 1.5- to 2.5-fold elevations of c-myc transcript levels in lesional psoriatic relative to normal epidermis. Similar increases in cyclophilin and lipocortin II transcripts were also observed and may reflect characteristic differences in RNA preparations from normal and psoriatic epidermis. C-myc, c-jun, c-erbB, c-fos, and c-Ha-ras transcript levels were not significantly increased in lesional psoriatic epidermis when protooncogene mRNA levels were normalized to those of the cyclophilin or lipocortin genes. In contrast, transforming growth factor-alpha (TGF-alpha) transcripts were significantly increased (10- to 20-fold) with or without prior normalization. C-myc, c-fos, and c-jun transcripts were significantly induced over in vivo levels 2-4 h after organ culture of normal or psoriatic keratome biopsies, demonstrating that these genes can be highly expressed in the context of tissue injury. Our results suggest that overexpression of these protooncogenes per se is not central to the pathogenesis of psoriatic epidermal hyperplasia. PMID: 1688596 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 956: J Hypertens Suppl. 1989 Dec;7(6):S114-5. Role of nuclear proto-oncogenes in the proliferation of aortic smooth muscle cells in spontaneously hypertensive rats. Baudouin-Legros M, Paquet JL, Brunelle G, Meyer P. Inserm U 7, Hopital Necker, Paris, France. In order to define the molecular mechanism involved in the enhancement of spontaneously hypertensive rat (SHR) cell proliferation, we compared the actions of fetal calf serum and angiotensin II on both SHR and Wistar-Kyoto rat (WKY) aortic smooth muscle cells. Both compounds were more mitogenic on the SHR cells than on the controls. However, phospholipase C hyper-responsiveness was present only after angiotensin stimulation. This was also true of the expression of c-jun, c-fos and c-myc. Oncogene overexpression therefore appears to be more strongly related to phospholipase C hyperreactivity than to enhanced proliferation of SHR aortic smooth muscle cells. PMID: 2632689 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 957: New Biol. 1989 Dec;1(3):297-304. Stimulation of protein kinase C or protein kinase A mediated signal transduction pathways shows three modes of response among serum inducible genes. Mechta F, Piette J, Hirai SI, Yaniv M. URA 152 CNRS, Departement de Biologie Moleculaire, Institut Pasteur, Paris, France. Activation of the signal transduction pathways mediated by protein kinase A (PKA) or protein kinase C (PKC) led to different responses of several serum inducible genes including the jun gene family, c-fos, c-myc, krox 20 and krox 24. Whereas all of these genes were stimulated by the phorbol ester TPA, a chemical activator of protein kinase C, they were differently regulated upon cAMP stimulation of the PKA dependent pathway. The proto-oncogenes jun B, c-fos, and to a lesser extent jun D were stimulated by increasing the intracellular concentration of cAMP, whereas the TPA stimulation of c-jun and c-myc was inhibited under these conditions. Krox 20 and krox 24 were insensitive to this second messenger. This study allowed us to classify these growth stimulated genes into three distinct groups distinguished by their sensitivity to elevated concentrations of intracellular cAMP. The inhibition of c-jun and c-myc expression in the presence of increased cAMP levels may be at least partially responsible for the growth inhibitory effect of this agent in Balb/c-3T3 cells. PMID: 2562123 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 958: Cell Tissue Kinet. 1989 Nov;22(6):405-24. Oncogenes, growth, and the cell cycle: an overview. Studzinski GP. Department of Pathology, UMDNJ-New Jersey Medical School, Newark 07103-2757. In spite of the complexity of the network of regulatory factors which control the balance between the cell cycle and quiescence, a picture is emerging, if only in outline. Several dozens of protooncogenes participate in growth signal transduction and integration, and, when expressed inappropriately, generate growth signals that may override other cellular controls. Some of these controls are provided by the negatively regulating growth factors, and when these are lost (e.g. by chromosomal deletion), or inactivated (e.g. by binding to an inactive analogue or a DNA viral oncoprotein), cell cycle activity is favoured over quiescence. Embryonic tissues are rapidly growing, so their cells are actively cycling and expression of proto-oncogenes is usually observed (Schuuring et al., 1989). As embryonic and stem cells in adult tissues mature, expression of the active proto-oncogenes is generally lost, but other proto-oncogenes may now be expressed (e.g. Muller et al., 1982). These changes in proto-oncogene expression are not achieved by modulation of transcriptional rates alone; transcriptional attenuation, message processing and stability, and post-translational protein modifications are all known to be important for the regulation of proto-oncogene expression during the transition from growth to the differentiated state. When quiescent cells re-enter the cell cycle approximately 60 genes become up-regulated, including proto-oncogene c-fos, the jun family, and c-myc (Zipfel et al., 1989). Evidence is strong that fos and jun proteins are transcriptional regulators. Terminal differentiation, on the other hand, is sometimes accompanied by the up-regulation of the ras gene family, as well as of several other proto-oncogenes. Proto-oncogene function is essential to the cell cycle traverse, but the genes involved are different in various cell types, and the precise order of oncogene expression may not turn out to be important. This is because cell cycle traverse appears to be more dependent on a critical threshold of growth signals propagated by parallel pathways, rather than on a strict order of predetermined steps. The participation of proto-oncogenes in growth signal transduction offers opportunities for errors, and abnormal growth may result from aberrant oncogene products generating a persistent or excessive growth signal, which shifts the balance of input to the integrating genes from quiescence to an active cell cycle. Thus, cancer may result from an entirely normal processing of growth signals that are abnormal.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review Review, Tutorial PMID: 2692830 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 959: J Biol Chem. 1989 Oct 5;264(28):16905-9. The antimitogenic action of tumor necrosis factor is associated with increased AP-1/c-jun proto-oncogene transcription. Dixit VM, Marks RM, Sarma V, Prochownik EV. Department of Pathology, University of Michigan School of Medicine, Ann Arbor 48109. Tumor necrosis factor alpha (TNF) mediates its in vivo antineoplastic effect primarily through its action on the endothelial surfaces of tumor vessels. Among other changes, endothelial cells increase the expression of surface procoagulant molecules which allow for the genesis of microthrombi and the eventual anoxic killing of the tumor tissue. TNF is also a potent antimitotic factor for endothelial cells. We investigated the molecular basis for these diverse actions of TNF by differential screening of a cDNA library prepared from TNF and cycloheximide-treated human umbilical vein endothelial (HUVE) cells. One of the cDNA clones identified encodes the transcription factor and proto-oncogene AP-1/c-jun. The expression of AP-1/c-jun following TNF treatment is mediated largely at the transcriptional level. Neither c-myc nor c-fos transcripts were induced by TNF. Since AP-1/c-jun has been previously identified as one of the earliest transcripts expressed in quiescent cells that have been stimulated by growth factors, our results suggest that mitogenic and antimitogenic stimuli evoke distinct yet overlapping sets of molecular responses and that the coordinate expression of AP-1/c-jun and c-fos is not mandatory. AP-1/c-jun transcript induction is thus one of the initial events mediated by TNF treatment of HUVE cells. PMID: 2506186 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 960: Proc Natl Acad Sci U S A. 1989 Oct;86(20):7711-5. v-maf, a viral oncogene that encodes a "leucine zipper" motif. Nishizawa M, Kataoka K, Goto N, Fujiwara KT, Kawai S. Department of Tumor Virus Research, University of Tokyo, Japan. We have molecularly cloned the provirus of the avian musculoaponeurotic fibrosarcoma virus AS42. Nucleotide sequence analysis of a biologically active clone of AS42 showed that this virus encodes a viral oncogene, maf. The deduced amino acid sequence of the v-maf gene product contains a "leucine zipper" motif similar to that found in a number of DNA binding proteins, including the gene products of the fos, jun, and myc oncogenes. However, unlike these oncogenes, the cellular maf gene was not transcriptionally activated by growth stimulation of cultured cells. PMID: 2554284 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 961: J Protein Chem. 1989 Oct;8(5):679-88. Conformational energy analysis of the leucine repeat regions of C/EBP, GCN4, and the proteins of the myc, jun, and fos oncogenes. Brandt-Rauf PW, Pincus MR, Chen JM, Lee G. Division of Environmental Sciences, Columbia-Presbyterian Medical Center, New York, New York 10032. It has been recently proposed that certain DNA binding proteins (including C/EBP, GCN4 and the myc, jun, and fos oncogene proteins) share a common structural motif based on helix-promoting regions containing heptad repeat sequences of leucines. It has been suggested that this structure is critical to the biological activity of these proteins, since it facilitates the formation of functional dimers held together by interdigitating leucine side-chains along the hydrophobic interfaces between long alpha-helical regions of the polypeptide chains in a configuration termed the "leucine zipper." In this paper, conformational energy analysis is used to deterrmine the preferred three-dimensional structures of the leucine repeat regions of these proteins. The results indicate that, in all cases, the global minimum energy conformation for these regions is an amphipathic alpha-helix with the leucine side-chains arrayed on one side in such a way to favor "leucine zipper" dimerization. Furthermore, amino acid substitutions in these regions (such as Pro for Leu), that are known to inhibit dimer formation and prevent DNA binding, are found to produce significant conformational changes that disrupt the amphipathic helical structure. Thus, these results provide support for the proposed "leucine zipper" configuration as a critical structural feature of this class of DNA binding proteins. PMID: 2532887 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 962: EMBO J. 1989 Oct;8(10):2795-801. The O2 gene which regulates zein deposition in maize endosperm encodes a protein with structural homologies to transcriptional activators. Hartings H, Maddaloni M, Lazzaroni N, Di Fonzo N, Motto M, Salamini F, Thompson R. Instituto Sperimentale per la Cerealicoltura, Sezione di Bergamo, Italy. The structure of the zein regulatory gene Opaque 2 of Zea mays has been determined by sequence analysis of genomic and cDNA clones. The size of O2 mRNA is 1751 bp [poly(A) tail not included] containing a major open reading frame (ORF) of 1380 bp preceded by three short ORFs of 3, 21 and 20 amino acid residues. The main ORF comprises 1362 bp and is composed of six exons ranging in size from 465 to 61 bp and five introns of 678 bp to 83 bp. A putative protein 454 amino acids long was derived by the theoretical translation of the genomic sequences corresponding to exons. The opaque 2 protein contains a domain similar to the leucine zipper motif identified in DNA binding proteins of animal protooncogenes such as fos, jun and myc, and in the transcriptional activators GCN4 and C/EBP. The region of 30 amino acid residues next to the leucine repeats towards the N terminus is rich in basic amino acids and is also homologous to a domain present in fos, jun and GCN4. Moreover, in the carboxy terminal region an amino acid motif closely resembling a metal binding domain is present. PMID: 2479535 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 963: Cancer Lett. 1989 Sep 15;47(1-2):1-9. Does NF-kappa B relieve the transcription block in c-myc? Renan MJ. Division of Biophysical Sciences, National Accelerator Centre, Faure, South Africa. In this report, it is shown that the mRNA of the c-myc oncogene is capable of forming an extensive stem-and-loop structure, with a free energy of delta G (25 degrees C) = -34 kcal. This secondary structure is situated at the 3' end of the first exon, immediately upstream of an elongation block. It is shown that this region contains potential binding sites for 3 different activator proteins, namely AP-1, AP-2, and nuclear factor-kappa B (NF-kappa B). From an analysis of the properties of these proteins, NF-kappa B could be identified as a candidate for the trans-acting factor involved in relieving the block to transcription. Publication Types: Review PMID: 2517588 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 964: Nature. 1989 Sep 7;341(6237):74-6. Changing fos oncoprotein to a jun-independent DNA binding protein with GCN4 dimerization specificity by swapping "leucine zippers". Sellers JW, Struhl K. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115. A structural motif for DNA-binding proteins, the 'leucine zipper', has been proposed for the jun, fos and myc gene products, the yeast transcriptional activator GCN4, and the C/EBP enhancer-binding protein. These proteins all contain a region with four or five leucine residues spaced exactly seven amino acid residues apart whose sequence is consistent with the formation of an amphipathic alpha-helix. It has been proposed that the leucine zipper consists of two interdigitated alpha-helices, one from each monomer, that constitute the dimerization function necessary for high-affinity binding to DNA; an adjacent region of basic residues is thought to be responsible for specific protein-DNA contacts. In support of this model, substitution of the leucine residues within the motif can abolish dimerization and DNA-binding, and a synthetic peptide corresponding to the GCN4 leucine zipper forms alpha-helical dimers. Despite the conserved leucine residues, however, each protein has a distinct dimerization specificity. Specifically, GCN4 homodimer, Jun homodimer and Fos-Jun heterodimer proteins bind to the same DNA site, whereas Fos is unable to form homodimers, bind DNA, or interact with GCN4 (refs 8-14). Here, we alter the dimerization specificity of Fos by precisely replacing its leucine zipper with that from GCN4. This Fos-GCN4 chimaeric protein is able to bind to the target site in the absence of Jun, and can form DNA-binding heterodimers with GCN4 but not with Jun. These results indicate that the leucine zipper is sufficient to confer dimerization specificity and strongly suggest that Fos contacts DNA directly. PMID: 2505087 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 965: Anticancer Res. 1989 Sep-Oct;9(5):1265-79. The human breast carcinoma cell line SW 613-S: an experimental system to study tumor heterogeneity in relation to c-myc amplification, growth factor production and other markers (review). Lavialle C, Modjtahedi N, Lamonerie T, Frebourg T, Landin RM, Fossar N, Lhomond G, Cassingena R, Brison O. Laboratoire d'Oncologie Moleculaire, UA 1158 CNRS, Institut Gustave Roussy, Villejuif, France. Amplification of the c-myc gene has been frequently reported in breast carcinomas. However the precise function of the c-myc protein is still unknown and the nature of the selective advantage offered to a cell by an overexpression of such a protein is unclear. We are addressing this question using the SW 613-S human breast carcinoma cell line as a model system. This cell line harbours an amplified c-myc gene and a mutated c-Ki-ras gene. By various criteria the amplified c-myc gene of SW613-S cells appears undistinguishable from a normal human c-myc gene. The SW613-S cell line is heterogeneous: it contains cells with a high level of amplification and carrying the extra copies of the c-myc gene in double minute chromosomes (DMs) and cells with few c-myc genes integrated into chromosomes. DM-containing cells are progressively lost upon in vitro cultivation but are selected for during in vivo growth, as tumors in nude mice, or by cultivating the cells in a chemically defined, serum-free medium or under conditions preventing anchorage. Clones with different levels of amplification and different chromosomal localization of the c-myc copies were isolated from the SW 613-S cell population. Those with a high level of amplification and expression of the c-myc gene are tumorigenic in nude mice, whereas those with a low level are not. Introduction of c-myc gene copies by transfection confers tumorigenicity to the nontumorigenic clones, indicating that a high level of amplification of the c-myc gene contributes to the tumorigenic phenotype of SW 613-S cells. Tumorigenic clones grow unattached, are able to proliferate in a chemically defined medium, and produce high levels of several growth factors (e.g. TGF-alpha, IGF2). Nontumorigenic clones are more dependent upon anchorage for growth, show a restricted growth in defined medium, and produce low or undetectable level of the growth factors tested. We have identified several genes, besides c-myc, the expression level of which is markedly different in the two types of clones. TGF-alpha, IGF2, PDGF-A, int-2, cytokeratins K8 and K18 and ferritin H chain are overexpressed in tumorigenic clones. In contrast, c-erbB1 (EGF receptor), c-jun, vimentin and p53 are expressed at a higher level in the nontumorigenic clones. Finally the major histocompatibility class I antigens, ferritin L chain, TGF-beta and c-Ki-ras, are examples of genes expressed at the same level in both types of clones.(ABSTRACT TRUNCATED AT 400 WORDS) Publication Types: Review PMID: 2686529 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 966: Proc Natl Acad Sci U S A. 1989 Sep;86(17):6474-8. Structure and expression of the murine retinoblastoma gene and characterization of its encoded protein. Bernards R, Schackleford GM, Gerber MR, Horowitz JM, Friend SH, Schartl M, Bogenmann E, Rapaport JM, McGee T, Dryja TP, et al. Whitehead Institute for Biomedical Research, Cambridge, MA 02142. We have isolated a cDNA clone of the murine homologue of the human retinoblastoma (Rb) susceptibility gene. DNA sequence analysis reveals a high degree of conservation with the human Rb sequence, both in the coding and in the noncoding regions. The predicted amino acid sequence of the mouse Rb protein shows 91% identity to that of the human protein. Both proteins were found to contain a peptide sequence reminiscent of a leucine-repeat motif ("leucine-zipper") that is also found in the myc, fos, and jun oncogenes. Synthetic peptide antiserum directed against a portion of the mouse Rb protein detects three proteins of 104-110 kDa in cells that were transiently transfected with a mouse Rb gene expression construct. In the mouse embryo the expression of Rb mRNA was ubiquitous, with maximal expression being observed around 13 days of gestation. In the embryo, the highest level of expression was observed in liver and brain. In contrast, the Rb gene was found to be expressed at a very low level in adult mouse liver with high levels being found in lung, thymus, and spleen. A shorter Rb transcript was detected in mouse testes. PMID: 2671991 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 967: Science. 1989 Aug 18;245(4919):740-3. Specific expression of nuclear proto-oncogenes before entry into meiotic prophase of spermatogenesis. Wolfes H, Kogawa K, Millette CF, Cooper GM. Dana-Farber Cancer Institute, Boston, MA. The expression of proto-oncogenes representative of several functional categories has been investigated during development of mouse male germ cells. The c-raf proto-oncogene and three members of the c-ras gene family were expressed in mitotically active stem cells, throughout the prophase of meiosis and to varying extents in post-meiotic cell types. In contrast, the nuclear proto-oncogenes c-fos, c-jun, and c-myc were specifically expressed at high levels in type B spermatogonia. High levels of c-myc and c-jun RNAs were also detected in spermatocytes early in the prophase of meiosis. The type B spermatogonia represent the last mitotic cell division before entry into meiotic prophase; therefore, these nuclear proto-oncogenes may be involved in altering programs of gene expression at this developmental transition. PMID: 2475907 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 968: J Virol. 1989 Aug;63(8):3220-6. Activation of endogenous c-fos proto-oncogene expression by human T-cell leukemia virus type I-encoded p40tax protein in the human T-cell line, Jurkat. Nagata K, Ohtani K, Nakamura M, Sugamura K. Department of Microbiology, Tohoku University School of Medicine, Sendai, Japan. We examined the ability of the trans-acting factor p40tax of human T-cell leukemia virus type I (HTLV-I), which is thought to be a crucial molecule in T-cell transformation by HTLV-I, to activate expression of a set of endogenous cellular genes related to T-cell proliferation. For this purpose we established a subclone (JPX-9) of Jurkat cells that was stably transfected with an expression plasmid containing the p40tax gene, whose expression is definitely dependent on heavy-metal ions. Expression of the interleukin-2 receptor alpha chain in JPX-9 cells was induced in response to the induction of p40tax expression, as has been demonstrated by others in transient transfection experiments with Jurkat cells. In addition, we found that significant enhancement of expression of the nuclear proto-oncogene c-fos was closely associated with expression of p40tax. Continous enhancement in the level of c-fos mRNA was observed in the presence of p40tax. In contrast, mRNA levels of other nuclear proto-oncogenes (c-myc, c-myb, and c-jun) were not appreciably effected by the expression of p40tax. These results suggest that (i) in addition to the interleukin-2-interleukin-2 receptor system, cellular genes such as c-fos, which regulate normal T-cell growth, are also activated directly or indirectly by p40tax and (ii) p40tax-induced modulation of gene expression plays a crucial role in T-cell transformation by HTLV-I. PMID: 2501514 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 969: J Biol Chem. 1989 May 25;264(15):8992-9. fos/jun and octamer-binding protein interact with a common site in a negative element of the human c-myc gene. Takimoto M, Quinn JP, Farina AR, Staudt LM, Levens D. Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland 20892. A negative element has previously been localized to a 57-base pair segment approximately 300 base pairs upstream of the human c-myc promoter P1. Within this element, a 26-base pair region was protected in vitro from DNase I digestion with a HeLa cell nuclear factor(s). Two specific DNA-protein complexes were identified in gel retardation assays using HeLa cell nuclear extracts and an oligonucleotide probe spanning the footprinted region. Exonuclease and chemical footprint analyses suggested that the binding sites for both complexes are almost entirely overlapping. One of the complexes was eliminated by oligonucleotide competitors possessing known AP-1 binding sites. This same complex reacted strongly with anti-fos immunoglobulin suggesting a role for c-fos in governing c-myc expression. Precipitation of fos protein bound to c-myc DNA that was immobilized on beads confirmed the involvement of c-fos in a specific complex with the c-myc upstream sequence. In contrast, the other complex seen by the c-myc probe could not be competitively inhibited by AP-1 binding sites and was not affected by anti-fos antibody. Instead, this complex was efficiently eliminated by unlabeled oligonucleotides containing the octamer DNA motif found in immunoglobulin gene promoters. Purified octamer-binding proteins formed stable complexes with the 26-base pair c-myc sequences. These results demonstrate that degeneracy in the consensus recognition sequences of these distinct factors allows each of them to bind the c-myc negative element. The interaction of known transcriptional activators with a negative element suggests that the same factors can mediate both transcriptional activation and repression. PMID: 2498322 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 970: Proc Natl Acad Sci U S A. 1989 Apr;86(7):2257-61. Deregulated expression of human c-jun transforms primary rat embryo cells in cooperation with an activated c-Ha-ras gene and transforms rat-1a cells as a single gene. Schutte J, Minna JD, Birrer MJ. NCI-Navy Medical Oncology Branch, National Cancer Institute, Bethesda, MD 20814. While the ability of the retroviral oncogene V-jun to transform chicken cells led to its discovery, the oncogenic potential of its cellular homologue, c-jun, which encodes a transcription factor, is unknown. We isolated a 1070-base-pair cDNA clone containing the unmutated entire open reading frame of c-jun from a human small cell lung cancer line. This cDNA as well as a 5.6-kilobase normal human genomic DNA fragment containing the c-jun gene were placed under the control of retroviral long terminal repeats and introduced into primary rat embryo cells (RECs), with or without other oncogenes, and into an immortal rat fibroblast cell line, Rat-1a, as a single gene. In Rat-1a cells the expression of human c-jun mRNA was associated with the ability to clone in soft agarose and form tumors in nude mice. When the c-jun cDNA or genomic DNA constructs were introduced into RECs, no foci of transformed cells were seen with c-jun alone or c-jun cotransfected with deregulated c-myc or L-myc protooncogenes. However, cotransfection of the c-jun constructs with an activated human c-Ha-ras gene led to foci of transformed cells which gave rise to immortalized cell lines that cloned in soft agarose and formed tumors in nude mice. Furthermore, formation of foci of transformed RECs by the c-jun/ras combination was augmented 3-fold by the tumor promoter phorbol 12-tetradecanoate 13-acetate. We conclude that deregulated expression of human c-jun can participate in malignant transformation of normal mammalian cells. PMID: 2648396 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 971: Science. 1989 Mar 31;243(4899):1681-8. The DNA binding domain of the rat liver nuclear protein C/EBP is bipartite. Landschulz WH, Johnson PF, McKnight SL. Howard Hughes Medical Institute, Carnegie Institution of Washington, Department of Embryology, Baltimore, MD 21210. C/EBP is a rat liver nuclear protein capable of sequence-specific interaction with DNA. The DNA sequences to which C/EBP binds in vitro have been implicated in the control of messenger RNA synthesis. It has therefore been predicted that C/EBP will play a role in regulating gene expression in mammalian cells. The region of the C/EBP polypeptide required for direct interaction with DNA has been identified and shown to bear amino acid sequence relatedness with the product of the myc, fos, and jun proto-oncogenes. The arrangement of these related amino acid sequences led to the prediction of a new structural motif, termed the "leucine zipper," that plays a role in facilitating sequence-specific interaction between protein and DNA. Experimental tests now provide support for the leucine zipper hypothesis. PMID: 2494700 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 972: Genes Dev. 1989 Mar;3(3):293-303. A FOS protein is present in a complex that binds a negative regulator of MYC. Hay N, Takimoto M, Bishop JM. Department of Microbiology and Immunology, University of California, San Francisco 94143. Regulation of the human proto-oncogene MYC apparently plays an important role in cellular proliferation and the genesis of diverse tumors. Transcription from MYC is governed principally by two promoters known as P1 and P2. Previously we have detected a negative regulator of these promoters upstream of MYC. We now report that this regulator comprises no more than 26 bp of DNA, with sequence that resembles the regulators of at least two other genes, and we describe nuclear factors that interact with the regulator. Nuclear extracts from human cells form three distinctive complexes with the negative regulator. One of these complexes includes the product of the proto-oncogene FOS or an antigenically related protein, and the FOS protein may, in turn, be associated with the product of the proto-oncogene JUN. Similarly, FOS and JUN proteins produced by translation in vitro bind cooperatively to the negative regulator. These results raise the possibility that FOS and JUN participate in the regulation of MYC. PMID: 2498162 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 973: Nature. 1989 Feb 16;337(6208):661-3. Prolonged activation of jun and collagenase genes by tumour necrosis factor-alpha. Brenner DA, O'Hara M, Angel P, Chojkier M, Karin M. Department of Medicine, University of California at San Diego, School of Medicine, La Jolla 92093. Tumour necrosis factor-alpha (TNF-alpha) is secreted by macrophages in response to inflammation, infection and cancer. Sublethal doses of recombinant TNF-alpha to rats causes cachexia, anaemia and inflammation. TNF-alpha plays a major part in tissue inflammation and remodelling by stimulating production of collagenase. Cellular responses to TNF-alpha are initiated by binding to high-affinity cell surface receptors. TNF-alpha then profoundly affects gene regulation, stimulating the fos, myc, interleukin-1 and interleukin-6 genes and inhibiting the type I collagen gene. Here we demonstrate that TNF-alpha also stimulates collagenase gene transcription; this stimulation is mediated by an element of the gene that is responsive to the transcription factor AP-1, the major component of which (jun/AP-1) is encoded by the jun gene; and that TNF-alpha stimulates prolonged activation of jun gene expression. This prolonged induction of jun contrasts with its transient activation by the phorbol ester TPA and provides a physiological example of the ability of jun/AP-1 to stimulate its own transcription. This may be a key mechanism for mediating at least some of the biological effects of TNF-alpha. PMID: 2537468 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 974: Mol Carcinog. 1989;2(4):208-16. Abolishment of c-fos inducibility in ras-transformed mouse osteoblast cell lines. Nose K, Itami M, Satake M, Ito Y, Kuroki T. Department of Cancer Cell Research, University of Tokyo, Japan. Proto-oncogene c-fos is induced by many types of cellular stimuli, such as 12-O-tetradecanoylphorbol-13-acetate (TPA), epidermal growth factor (EGF), serum (fetal bovine), calcium ionophore A23187, and dibutyryl cAMP (But2cAMP). In this study, c-fos induction was abolished in ras-transformed mouse osteoblast cells (MC3T3). Transformants of MC3T3 were isolated after transfection with Ki or Ha murine sarcoma virus DNA. All Ki- or Ha-ras transformed MC3T3 clones examined showed exceedingly low levels of c-fos induction by all inducers, as determined by the change in amounts of c-fos mRNA or its product. Induction of other TPA-responsive genes, such as metallothionein, was not altered in some ras-transformed cells; c-myc and c-jun expression was constitutively high in all the ras-transformed clones. Nuclear extracts and gel shift assay showed that the binding activity to c-fos enhancer element (serum response element) was altered in ras-transformed cells. These results indicate that transformation with ras oncogene induces modification of c-fos enhancer binding factors and that this modification is one cause for the decrease in c-fos induction. PMID: 2478147 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 975: Nature. 1988 Dec 15;336(6200):692-5. Direct interaction between fos and jun nuclear oncoproteins: role of the 'leucine zipper' domain. Sassone-Corsi P, Ransone LJ, Lamph WW, Verma IM. Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92138. Gene expression is modulated by the specific interactions of nuclear proteins with unique regulatory sequences in the genome. Proteins involved in transcriptional regulation seem to be either transcription factors or transcription modulators and their interactions are crucial in determining whether the expression of a specific gene is activated or repressed. Recently, the product of the proto-oncogene jun has been identified as the transcription factor AP-1, whereas nuclear oncoproteins fos and myc have been implicated in transcriptional transregulation of several promoters. Furthermore, the products of the fos and jun proto-oncogenes are associated in some transcription complexes. Although the nature of the association is unclear, the two proteins co-immunoprecipitate with fos antibodies in nuclear extracts. Here, we report studies that demonstrate that the fos protein directly modulates jun function by means of a heterodimer of fos and jun proteins. The fos 'leucine zipper' domain is necessary for the DNA binding of the heterodimer; a distinct domain, localized in the C-terminal region of the fos protein, is responsible for transcriptional regulation. PMID: 3143919 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 976: Nature. 1988 Dec 15;336(6200):646-51. The role of the leucine zipper in the fos-jun interaction. Kouzarides T, Ziff E. Department of Biochemistry, New York University Medical Center, New York 10016. Mutagenesis of the fos protein supports the hypothesis that a heptad repeat of leucine residues stabilizes the interaction between the fos and jun proteins. We show that the complex between fos and jun can bind to DNA more tightly than either protein alone and that basic residues adjacent to the leucine repeat of fos contribute to the DNA-binding potential of the complex. PMID: 2974122 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 977: Science. 1988 Dec 9;242(4884):1430-3. Cyclic AMP-responsive DNA-binding protein: structure based on a cloned placental cDNA. Hoeffler JP, Meyer TE, Yun Y, Jameson JL, Habener JF. Laboratory of Molecular Endocrinology, Massachusetts General Hospital, Boston. Cyclic AMP (cAMP) is an intracellular second messenger that activates transcription of many cellular genes. A palindromic consensus DNA sequence, TGACGTCA, functions as a cAMP-responsive transcriptional enhancer (CRE). The CRE binds a cellular protein of 38 kD in placental JEG-3 cells. A placental lambda gt11 library was screened for expression of specific CRE-binding proteins with the CRE sequence as a radioactive probe. A cDNA encoding a protein of 326 amino acids with the binding properties of a specific CRE-binding protein (CREB) was isolated. The protein contains a COOH-terminal basic region adjacent to a sequence similar to the "leucine zipper" sequence believed to be involved in DNA binding and in protein-protein contacts in several other DNA-associated transcriptional proteins including the products of the c-myc, c-fos, and c-jun oncogenes and GCN4. The CREB protein also contains an NH2-terminal acidic region proposed to be a potential transcriptional activation domain. The putative DNA-binding domain of CREB is structurally similar to the corresponding domains in the phorbol ester-responsive c-jun protein and the yeast transcription factor GCN4. PMID: 2974179 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 978: Cell. 1988 Dec 2;55(5):907-15. DNA binding activities of three murine Jun proteins: stimulation by Fos. Nakabeppu Y, Ryder K, Nathans D. Howard Hughes Medical Institute Laboratory, Baltimore, Maryland. Three members of the Jun/AP-1 family have been identified in mouse cDNA libraries: c-Jun, Jun-B, and Jun-D. We have compared the DNA binding properties of the Jun proteins by using in vitro translation products in gel retardation assays. Each protein was able to bind to the consensus AP-1 site (TGACTCA) and, with lower affinity, to related sequences, including the cyclic AMP response element TGACGTCA. The relative binding to the oligonucleotides tested was similar for the different proteins. The Jun proteins formed homodimers and heterodimers with other members of the family, and they were bound to the AP-1 site as dimers. When Fos translation product was present, DNA binding by Jun increased markedly, and the DNA complex contained Fos. The C-terminal homology region of Jun was sufficient for DNA binding, dimer formation, and interaction with Fos. Our general conclusion is that c-Jun, Jun-B, and Jun-D are similar in their DNA binding properties and in their interaction with Fos. If there are functional differences between them, they are likely to involve other activities of the Jun proteins. PMID: 3142691 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 979: Nature. 1988 Aug 18;334(6183):629-31. Induction of proto-oncogene JUN/AP-1 by serum and TPA. Lamph WW, Wamsley P, Sassone-Corsi P, Verma IM. Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92138. The response of a cell to mitogens and differentiation agents involves the transcriptional induction of several cellular genes. Prominent among these so-called 'immediate early' or 'competence' genes are the nuclear oncogenes fos and myc. Although the precise function of these early response genes in growth control is not understood, it is likely that many of them are involved in the transition from G0 to G1 in the cell cycle. The findings that the products of nuclear proto-oncogenes jun and erbA are transcriptional factors supports the notion of the role of the nuclear oncoproteins in the regulation of gene expression. Recently, it has been reported that the FOS protein is associated in transcriptional complexes with the product of the jun oncogene, the transcription factor AP-1. As the fos gene is induced in response to mitogens during initiation of cell growth, we investigated whether expression of the nuclear transcription factor AP-1 is also inducible. We report that mouse c-jun gene transcription is rapidly induced by serum and phorbol-ester 12-o-tetradecanoyl phorbol 13-acetate (TPA). Furthermore, induction is transient and the mRNA is superinduced by inhibitors of protein synthesis. PMID: 2457172 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 980: Nature. 1988 Aug 11;334(6182):535-7. Transcriptional activation of c-jun during the G0/G1 transition in mouse fibroblasts. Ryseck RP, Hirai SI, Yaniv M, Bravo R. European Molecular Biology Laboratory, Heidelberg, FRG. Before quiescent cells can respond to mitogens and progress through the G1 phase of cell growth, new messenger RNA synthesis is required. The G1 phase seems to be a critical point of control in the cell cycle, where normal cells deprived of growth factors halt cycling while transformed cells do not, suggesting that regulatory genes, uncontrolled in the neoplastic phenotype, are expressed during the G0 to G1 transition. Some of these may code for nuclear proteins that participate in the transactivation of genes required for the progression through G1. The observed changes in expression of the proto-oncogenes c-fos and c-myc, following stimulation of fibroblasts with growth factors, support this notion as recent evidence suggests that c-FOS and c-MYC proteins can function as transactivating factors. Moreover, the rapid induction of several genes in fibroblasts coding for putative transacting factors during the G0 to G1 transition has been recently reported. Here we present the nucleotide sequence of a mouse cDNA clone coding for a 334 residue protein which shows 80% similarity with v-JUN and more than 98% similarity with the human c-JUN sequence. We have demonstrated that in quiescent fibroblasts c-jun transcription is rapidly induced during the G0 to G1 transition. PMID: 3136397 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 981: EMBO J. 1988 Aug;7(8):2475-83. Transforming but not immortalizing oncogenes activate the transcription factor PEA1. Wasylyk C, Imler JL, Wasylyk B. Unite 184 INSERM, Laboratoire de Genetique Moleculaire des Eucaryotes du CNRS, Faculte de Medecine, Stasbourg, France. The transcription factor PEA1 (a homologue of AP1 and c-jun) is highly active in several fibroblast cell lines, compared to its low activity in a myeloma and an embryo-carcinoma (EC) cell line. Serum components are essential to attain these high levels of PEA1 activity in fibroblasts. This serum requirement is abrogated by transformation with the oncogenes c-Ha-ras, v-src and polyoma middle T (Py-MT) but not by immortalization with polyoma large T (Py-LT), v-myc, c-myc or SV40 large T (SV40T). Expression in myeloma cells of the same transforming oncogenes, as well as v-mos and c-fos, activates PEA1, whereas expression of the same immortalizing oncogenes and EIA does not. These results suggest that a common target for transforming oncogenes is PEA1. Serum components have no effect on PEA1 activity in the myeloma and EC cell lines. In contrast, retinoic acid treatment of F9 EC cells augments PEA1 activity. These results suggest that transforming oncogene expression compensates for the absence of cell type-specific factors which are required to activate PEA1. Activation of PEA1 may lead to altered transcription of a set of transformation-related genes. PMID: 3142763 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 982: Science. 1988 Jun 24;240(4860):1759-64. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Landschulz WH, Johnson PF, McKnight SL. Department of Embryology, Carnegie Institution of Washington, Baltimore, MD 21210. A 30-amino-acid segment of C/EBP, a newly discovered enhancer binding protein, shares notable sequence similarity with a segment of the cellular Myc transforming protein. Display of these respective amino acid sequences on an idealized alpha helix revealed a periodic repetition of leucine residues at every seventh position over a distance covering eight helical turns. The periodic array of at least four leucines was also noted in the sequences of the Fos and Jun transforming proteins, as well as that of the yeast gene regulatory protein, GCN4. The polypeptide segments containing these periodic arrays of leucine residues are proposed to exist in an alpha-helical conformation, and the leucine side chains extending from one alpha helix interdigitate with those displayed from a similar alpha helix of a second polypeptide, facilitating dimerization. This hypothetical structure is referred to as the "leucine zipper," and it may represent a characteristic property of a new category of DNA binding proteins. PMID: 3289117 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 983: Proc Natl Acad Sci U S A. 1987 May;84(10):3316-9. Homology between the DNA-binding domain of the GCN4 regulatory protein of yeast and the carboxyl-terminal region of a protein coded for by the oncogene jun. Vogt PK, Bos TJ, Doolittle RF. The product of the recently described oncogene jun shows significant amino acid sequence homology with the GCN4 yeast transcriptional activator protein. The similarity is restricted to the 66 carboxyl-terminal amino acids, thought to be the DNA-binding domain of the GCN4 protein. In these alpha-helix-permissive regions of the jun and GCN4 products there is also a lesser but still significant amino acid resemblance to the fos protein and a marginal degree of similarity to myc proteins. The amino acid sequence homology between GCN4 and jun gene products suggests that the jun protein may bind to DNA in a sequence-specific way and exert a regulatory function. PMID: 3554236 [PubMed - indexed for MEDLINE] ---------------------------------------------------------------