1: Matrix Biol. 2005 Oct 14; [Epub ahead of print] Characterization of human Collagen XVIII promoter 2: Interaction of Sp1, Sp3 and YY1 with the regulatory region and a SNP that increases transcription in hepatocytes. Armelin-Correa LM, Lin CJ, Barbosa A, Bagatini K, Winnischofer SM, Sogayar MC, Passos-Bueno MR. Department of Genetics and Evolution Biology , Bioscience Institute, University of Sao Paulo, Sao Paulo, Brazil. Different levels of Collagen XVIII expression have been associated with several pathological processes such as cancer, liver fibrosis, diabetic retinopathy and Alzheimer's disease. Understanding the transcriptional regulation of Collagen XVIII might elucidate some pathways related to the progression of these diseases. The promoter 2 of COL18A1 gene is poorly understood and is responsible for the transcription of this gene in several adult tissues such as liver, eyes and brain. This study focused upon characterization of cis-regulatory elements interacting with human COL18A1 promoter 2 and identification of SNPs in this region in different ethnic groups. Our results show that there are five conserved regions (I to V) between human and mouse promoter 2 and that the human COL18A1 core promoter is located between nucleotides -186 and -21. Sp1 and Sp3 bind to conserved regions I and V, while Sp3 and YY1 interact with region II. We have verified that the SNP at position -700 (T>G) is embedded in two common haplotypes, which have different frequencies between European and African descendents. The allele -700G increases transcription and binding for a still unknown transcription factor. SNP -700 affects Sp3 and YY1 interaction with this region, even though it is not part of these transcription factors' predicted binding sites. Therefore, our results show for the first time that Sp3 and YY1 interact with human COL18A1 promoter 2, and that nucleotide -700 is part of a binding motif for a still unknown TF that is involved in the expression of this gene in hepatocytes. In addition, we also confirm the involvement of Sp1 in the regulation of this gene. PMID: 16229994 [PubMed - as supplied by publisher] --------------------------------------------------------------- 2: Physiol Genomics. 2005 Sep 27; [Epub ahead of print] Is there an autoimmune process in bone? Gene expression studies in diabetic and non-diabetic BB rats as well as BB rat-related and -unrelated rat strains. Kloting N, Follak N, Kloting I. Laboratory Animal Science, University of Greifswald, Medical Faculty, Karlsburg, Germany. It is well known that type 1 diabetes is associated with a decrease in bone mass and delayed healing of fractures in human and in animal models of type 1 diabetes. Using well and poorlycompensated diabetic BB/O(ttawa) K(arlsburg) rats spontaneously developing insulindependent type 1 diabetes, it was recently shown that, in contrast to all other tissues studied, bone is most influenced by metabolic state and seems to be regulated in a manner different from other organs. Therefore, we studied the expression of additional genes (Bmp-1, Bmp-4, Vegf, Bglap, Il-1b, Ifng, Tnfa, Calca, Sp1, Yy1) in bone of non-diabetic BB rats compared with newly diagnosed, well- and poorly-compensated diabetic rats as well as in 2 nondiabetes-prone, congenic BB.SHR rats, BB rat-related (WOKW) and -unrelated rat strains (F344). Six males of each group were euthanized, the tibial bone was removed, total RNA was extracted, transcribed in complementary DNA, and used for real-time PCR. Comparing non-diabetic with diabetic groups, the relative gene expression was reduced by more than 80% in newly diagnosed and in well-compensated diabetic BB/OK rats. The gene expression in poorly-compensated rats increased significantly in 7 out of 10 genes and was comparable with those of non-diabetic BB/OK rats. Comparing gene expression between diabetes-prone BB/OK and non-diabetes-prone BB.1K, BB.4S, WOKW and F344, there were no significant differences between newly diagnosed and well-compensated BB/OK diabetic rats and non-diabetic BB.1K, BB.4S, WOKW and F344 rats. Based on these findings we concluded that spontaneous diabetes influences bone gene expression in BB/OK rats, which may be attributed to the genetically determined autoimmune process not only affecting pancreatic beta-cells but also bone formation and resorption. PMID: 16189279 [PubMed - as supplied by publisher] --------------------------------------------------------------- 3: J Virol. 2005 Oct;79(20):12852-60. Host cell nuclear proteins are recruited to cytoplasmic vaccinia virus replication complexes. Oh J, Broyles SS. Department of Biochemistry, Purdue University, West Lafayette, IN 47907, USA. The initiation and termination of vaccinia virus postreplicative transcription have been reported to require cellular proteins, some of which are believed to be nuclear proteins. Vaccinia virus replicates in the cytoplasmic compartment of the cell, raising questions as to whether vaccinia virus has access to nuclear proteins. This was addressed here by following the fate of several nuclear proteins after infection of cells with vaccinia virus. The nuclear transcription factors YY1, SP1, and TATA binding protein were found to colocalize with virus replication complexes in the cytoplasm of infected cells. In addition, the nuclear proteins RNA polymerase II, TAFIIp32, and histone deacetylase 8, but not the structural protein lamin B, also were found in the cytoplasm of the cell. The association of YY1 with replication complexes was dependent on DNA replication and required only the DNA binding domain of the protein, indicating that DNA binding alone may be responsible for the association of nuclear transcription factors with viral replication complexes in the cytoplasm. The cytoplasmic localization of YY1 was resistant to the nuclear export inhibitor leptomycin B. Evidence is presented indicating that nuclear import and export pathways were not adversely affected by vaccinia virus infection. These observations indicate that vaccinia virus replication complexes have ready access to nuclear proteins by allowing leakage from the nucleus. PMID: 16188987 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 4: FEBS J. 2005 Oct;272(19):5031-55. Mitochondrial biogenesis in mtDNA-depleted cells involves a Ca2+-dependent pathway and a reduced mitochondrial protein import. Mercy L, Pauw A, Payen L, Tejerina S, Houbion A, Demazy C, Raes M, Renard P, Arnould T. Laboratory of Biochemistry and Cellular Biology, University of Namur (FUNDP), Belgium. Alterations in mitochondrial activity resulting from defects in mitochondrial DNA (mtDNA) can modulate the biogenesis of mitochondria by mechanisms that are still poorly understood. In order to study mitochondrial biogenesis in cells with impaired mitochondrial activity, we used rho-L929 and rho(0)143 B cells (partially and totally depleted of mtDNA, respectively), that maintain and even up-regulate mitochondrial population, to characterize the activity of major transcriptional regulators (Sp1, YY1, MEF2, PPARgamma, NRF-1, NRF-2, CREB and PGC-1alpha) known to control the expression of numerous nuclear genes encoding mitochondrial proteins. Among these regulators, cyclic AMP-responsive element binding protein (CREB) activity was the only one to be increased in mtDNA-depleted cells. CREB activation mediated by a calcium-dependent pathway in these cells also regulates the expression of cytochrome c and the abundance of mitochondrial population as both are decreased in mtDNA-depleted cells that over-express CREB dominant negative mutants. Mitochondrial biogenesis in mtDNA-depleted cells is also dependent on intracellular calcium as its chelation reduces mitochondrial mass. Despite a slight increase in mitochondrial mass in mtDNA-depleted cells, the mitochondrial protein import activity was reduced as shown by a decrease in the import of radiolabeled matrix-targeted recombinant proteins into isolated mitochondria and by the reduced mitochondrial localization of ectopically expressed HA-apoaequorin targeted to the mitochondria. Decrease in ATP content, in mitochondrial membrane potential as well as reduction in mitochondrial Tim44 abundance could explain the lower mitochondrial protein import in mtDNA-depleted cells. Taken together, these results suggest that mitochondrial biogenesis is stimulated in mtDNA-depleted cells and involves a calcium-CREB signalling pathway but is associated with a reduced mitochondrial import for matrix proteins. PMID: 16176275 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 5: Nucleic Acids Res. 2005 Aug 22;33(15):4740-53. Print 2005. Characterization of cis- and trans-acting elements in the imprinted human SNURF-SNRPN locus. Rodriguez-Jato S, Nicholls RD, Driscoll DJ, Yang TP. Department of Biochemistry and Molecular Biology, University of Florida College of Medicine Gainesville, FL 32610, USA. The imprinted SNRPN locus is a complex transcriptional unit that encodes the SNURF and SmN polypeptides as well as multiple non-coding RNAs. SNRPN is located within the Prader-Willi and Angelman syndrome (PWS/AS) region that contains multiple imprinted genes, which are coordinately regulated by a bipartite imprinting center (IC). The SNRPN 5' region co-localizes with the PWS-IC and contains two DNase I hypersensitive sites, DHS1 at the SNRPN promoter, and DHS2 within intron 1, exclusively on the paternally inherited chromosome. We have examined DHS1 and DHS2 to identify cis- and trans-acting regulatory elements within the endogenous SNRPN 5' region. Analysis of DHS1 by in vivo footprinting and chromatin immunoprecipitation identified allele-specific interaction with multiple regulatory proteins, including NRF-1, which regulates genes involved in mitochondrial and metabolic functions. DHS2 acted as an enhancer of the SNRPN promoter and contained a highly conserved region that showed allele-specific interaction with unphosphorylated RNA polymerase II, YY1, Sp1 and NRF-1, further suggesting a key role for NRF-1 in regulation of the SNRPN locus. We propose that one or more of the regulatory elements identified in this study may also contribute to PWS-IC function. PMID: 16116039 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 6: J Invest Dermatol. 2005 Jun;124(6):1206-14. Transcriptional regulation of ATP2C1 gene by Sp1 and YY1 and reduced function of its promoter in Hailey-Hailey disease keratinocytes. Kawada H, Nishiyama C, Takagi A, Tokura T, Nakano N, Maeda K, Mayuzumi N, Ikeda S, Okumura K, Ogawa H. Atopy (Allergy) Research Center, Juntendo University School of Medicine, Tokyo, Japan. Hailey-Hailey disease (HHD) is a blistering skin disease caused by malfunction of the Ca2+-dependent ATPase, ATP2C1. In this study, key regulatory regions necessary for the expression of the gene encoding human ATP2C1 were investigated. The transient reporter assay demonstrated that region +21/+57 was necessary for activation of the ATP2C1 promoter, and the electrophoretic mobility shift assay demonstrated that the region was recognized by the transcription factors, Sp1 and YY1. In accordance with this result, when Sp1 or YY1 was overexpressed in keratinocytes, an obvious increase in ATP2C1 promoter activity was observed, which was in contrast with the case where a mutant promoter lacking the binding sites for Sp1 and YY1 was used as the reporter. Ca2+-stimulation signal increased nuclear Sp1 proteins and ATP2C1 mRNA levels in normal keratinocytes. In contrast, both these increases were suppressed in keratinocytes from HHD patients. These results indicate that Sp1 and YY1 transactivate the human ATP2C1 promoter via cis-enhancing elements and that incomplete upregulation of ATP2C1 transcription contributes to the keratinocyte-specific pathogenesis of HHD. This is a report describing the regulation of the expression of ATP2C1. PMID: 15955096 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 7: Genome Biol. 2005;6(4):R33. Epub 2005 Mar 29. Promoter features related to tissue specificity as measured by Shannon entropy. Schug J, Schuller WP, Kappen C, Salbaum JM, Bucan M, Stoeckert CJ Jr. Center for Bioinformatics, University of Pennsylvania, Philadelphia, PA 19104, USA. jschug@pcbi.upenn.edu BACKGROUND: The regulatory mechanisms underlying tissue specificity are a crucial part of the development and maintenance of multicellular organisms. A genome-wide analysis of promoters in the context of gene-expression patterns in tissue surveys provides a means of identifying the general principles for these mechanisms. RESULTS: We introduce a definition of tissue specificity based on Shannon entropy to rank human genes according to their overall tissue specificity and by their specificity to particular tissues. We apply our definition to microarray-based and expressed sequence tag (EST)-based expression data for human genes and use similar data for mouse genes to validate our results. We show that most genes show statistically significant tissue-dependent variations in expression level. We find that the most tissue-specific genes typically have a TATA box, no CpG island, and often code for extracellular proteins. As expected, CpG islands are found in most of the least tissue-specific genes, which often code for proteins located in the nucleus or mitochondrion. The class of genes with no CpG island or TATA box are the most common mid-specificity genes and commonly code for proteins located in a membrane. Sp1 was found to be a weak indicator of less-specific expression. YY1 binding sites, either as initiators or as downstream sites, were strongly associated with the least-specific genes. CONCLUSIONS: We have begun to understand the components of promoters that distinguish tissue-specific from ubiquitous genes, to identify associations that can predict the broad class of gene expression from sequence data alone. PMID: 15833120 [PubMed - in process] --------------------------------------------------------------- 8: J Biol Chem. 2005 Apr 22;280(16):16508-13. Epub 2005 Mar 8. Induction of endoplasmic reticulum-induced stress genes in Panc-1 pancreatic cancer cells is dependent on Sp proteins. Abdelrahim M, Liu S, Safe S. Institute of Biosciences and Technology, Texas A and M University System Health, Science Center, Houston, Texas 77030-3303, USA. Endoplasmic reticulum (ER) stress plays a critical role in multiple diseases, and pharmacologically active drugs can induce cell death through ER stress pathways. Stress-induced genes are activated through assembly of transcription factors on ER stress response elements (ERSEs) in target gene promoters. Gel mobility shift and chromatin immunoprecipitation assays have confirmed interactions of NF-Y and YY1 with the distal motifs of the tripartite ERSE from the glucose-related protein 78 (GRP78) gene promoter. The GC-rich nonanucleotide (N(9)) sequence, which forms the ER stress response binding factor (ERSF) complex binds TFII-I and ATF6; however, we have now shown that in Panc-1 pancreatic cancer cells, this complex also binds Sp1, Sp3, and Sp4 proteins. Sp proteins are constitutively bound to the ERSE; however, activation of GRP78 protein (or reporter gene) by thapsigargin or tunicamycin is inhibited after cotransfection with small inhibitory RNAs for Sp1, Sp3, and Sp4. This study demonstrates that Sp transcription factors are important for stress-induced responses through their binding to ERSEs. PMID: 15760841 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 9: BMC Evol Biol. 2005 Feb 13;5(1):15. The architecture of mammalian ribosomal protein promoters. Perry RP. Fox Chase Cancer Center Philadelphia, PA 19111, USA. RP_Perry@fccc.edu BACKGROUND: Mammalian ribosomes contain 79 different proteins encoded by widely scattered single copy genes. Coordinate expression of these genes at transcriptional and post-transcriptional levels is required to ensure a roughly equimolar accumulation of ribosomal proteins. To date, detailed studies of only a very few ribosomal protein (rp) promoters have been made. To elucidate the general features of rp promoter architecture, I made a detailed sequence comparison of the promoter regions of the entire set of orthologous human and mouse rp genes. RESULTS: A striking evolutionarily conserved feature of most rp genes is the separation by an intron of the sequences involved in transcriptional and translational regulation from the sequences with protein encoding function. Another conserved feature is the polypyrimidine initiator, which conforms to the consensus (Y)2C+1TY(T)2(Y)3. At least 60 % of the rp promoters contain a largely conserved TATA box or A/T-rich motif, which should theoretically have TBP-binding capability. A remarkably high proportion of the promoters contain conserved binding sites for transcription factors that were previously implicated in rp gene expression, namely upstream GABP and Sp1 sites and downstream YY1 sites. Over 80 % of human and mouse rp genes contain a transposable element residue within 900 bp of 5' flanking sequence; very little sequence identity between human and mouse orthologues was evident more than 200 bp upstream of the transcriptional start point. CONCLUSIONS: This analysis has provided some valuable insights into the general architecture of mammalian rp promoters and has identified parameters that might coordinately regulate the transcriptional activity of certain subsets of rp genes. PMID: 15707503 [PubMed - in process] --------------------------------------------------------------- 10: Gene. 2004 Oct 27;341:141-8. The antiapoptotic gene Ian4l1 in the rat: genomic organization and promoter characterization. Andersen UN, Markholst H, Hornum L. Hagedorn Research Institute, Niels Steensens Vej 6, Gentofte DK-2820, Denmark. Rat immune-associated nucleotide 4-like 1 (Ian4l1) encodes an antiapoptotic protein, which is essential for T-cell survival. A frameshift mutation at codon 85 in the biobreeding diabetes-prone (BBDP) rat is the cause of their life-long T-cell lymphopenia, which includes lack of regulatory T-cells--a prerequisite for spontaneous autoimmune destruction of their beta-cells. This study reports the identification of seven Ian4l1 mRNA variants. The genomic organization of the exons indicates three promoter regions. The promoter of two of the mRNAs was characterized. Rapid amplification of cDNA ends (RACE) and ribonuclease protection assay (RPA) demonstrated multiple transcription start sites (TSS) with two major sites. The localization of the core promoter and regulatory regions was identified by a luciferase assay of the 2.7-kb upstream of the TSS. The regulatory regions functioned similarly in two cell lines--one expressing Ian4l1 and one not expressing it. This indicates that the cell-specific expression is controlled by regions outside the 2.7-kb region, or by the chromatin structure or chromatin methylation level. The core promoter is TATA-less and initiator element-less, and contains putative binding sites for YY1, Sp1, and MED-1, the latter being an element believed to be important for transcription from TATA-less promoters. PMID: 15474297 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 11: Bull Exp Biol Med. 2004 May;137(5):485-9. Regulation of ceruloplasmin gene in mammals. Gyulikhandanova NE, Tsymbalenko NV, Platonova NA, Babich VS, Puchkova LV. St. Petersburg State Technological university. ludmila@usr8.iem.ras.spb.ru A site of rat DNA (about 1800 b. p.) adjacent to the first ceruloplasmin gene contains, apart from regulatory sequences common for all eukaryotic promotors, cis-elements, which are potential binding sites for soluble nuclear receptors of some hormones. Sequences characteristic of genes expressed in liver cells and mammary gland cells during lactation were detected. Full-length fragment of this locus of ceruloplasmin gene (1800 b. p.) was synthesized by PCR and used in gel shift experiments. It was found that soluble proteins extracted from purified nuclei of mammary gland cells during lactation and from the liver of adult and newborn rats, contain proteins specifically interacting with the PCR product. A fragment of chromosome gene containing exons encoding the central part of rat ceruloplasmin was cloned in pTZ19 bacterial vector. Gel shift assay showed that the cloned fragment contained binding sites for specific transcription factor YY1, whose level in nuclear protein fractions varied during ontogeny (according to immunoblotting data). Monoclonal antibodies detected protein YY1 in the complex of cloned DNA-nuclear proteins. Possible mechanisms of tissue-specific regulation of ceruloplasmin gene varying during ontogeny are discussed. PMID: 15455125 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 12: J Biol Chem. 2004 Sep 3;279(36):38055-61. Epub 2004 Jun 25. SOX7 and SOX17 regulate the parietal endoderm-specific enhancer activity of mouse laminin alpha1 gene. Niimi T, Hayashi Y, Futaki S, Sekiguchi K. Sekiguchi Biomatrix Signaling Project, ERATO, Japan Science and Technology Agency, Aichi Medical University, Karimata, Yazako, Nagakute, Aichi 480-1195, Japan. Laminin-1 is the major component of embryonic basement membrane and consists of alpha1, beta1, and gamma1 chains. The expression of laminin-1 is induced in mouse F9 embryonal carcinoma cells upon differentiation into parietal endoderm cells. We recently identified a parietal endoderm-specific enhancer in the mouse laminin alpha1 (Lama1) gene and showed that Sp1/Sp3 and YY1 transcription factors were involved in the enhancer activity. Although here we identified that NF-Y binds to the enhancer sequence between Sp1/Sp3- and YY1-binding sites, all these transcription factors are ubiquitously expressed and thus are not sufficient to explain parietal endoderm-specific enhancer activity. In the present study, we further showed that SOX7 and SOX17 are involved in the regulation of parietal endoderm-specific enhancer activity of the mouse Lama1 gene. Northern blot analysis revealed that the steady-state levels of mouse Sox7 and Sox17 mRNAs increased in parallel with that of Lama1 mRNA during the differentiation of F9 cells. Both SOX7 and SOX17 markedly trans-activated the transcription of the Lama1 enhancer-reporter construct in undifferentiated F9 cells in a manner dependent on high mobility group box-mediated DNA binding. Electrophoretic mobility shift assays and mutational analyses revealed that SOX7 and SOX17 bound specifically to two SOX-binding sites within the Lama1 enhancer, and that these SOX-binding sites functioned synergistically to confer the trans-activation by SOX7 and SOX17. Furthermore, this trans-activation was dependent on the integrity of the binding sites for Sp1/Sp3 and NF-Y located at upstream of the two SOX-binding sites. These results indicate that the transcription of the mouse Lama1 gene during the differentiation of F9 cells is controlled by a combination of the actions of the ubiquitous factors, Sp1/Sp3 and NF-Y, and the parietal endoderm-specific factors, SOX7 and SOX17. PMID: 15220343 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 13: Genomics. 2004 May;83(5):873-82. Clusters of regulatory signals for RNA polymerase II transcription associated with Alu family repeats and CpG islands in human promoters. Oei SL, Babich VS, Kazakov VI, Usmanova NM, Kropotov AV, Tomilin NV. Institute of Biochemistry, Free University of Berlin, Thielallee 63, D-14195, Berlin-Dahlem, Germany. Primate genomes contain a very large number of short interspersed GC-rich repeats of the Alu family, which are abundant in introns and intergenic spacers but also present in 5' flanking regions of genes enriched in binding motifs (BMs) for transcription factors and frequently containing CpG islands. Here we studied whether CpG islands located in promoters of human genes overlap with Alu repeats and with clusters of BMs for the zinc-finger transcription factors Sp1, estrogen receptor alpha, and YY1. The presence of estrogen-response elements in Alu was shown earlier and here we confirm the presence in the consensus Alu sequence of the binding sites for Sp1 and YY1. Analyzing >5000 promoters from the two databases we found that Alu sequences are underrepresented in promoters compared to introns and that approximately 4% of CpG islands located within the -1000 to +200 segments of human promoters overlap with Alu repeats. Although this fraction was found to be lower for proximal segments of promoters (-500 to +100), our results indicate that a significant number (>1000) of all human genes may be controlled by Alu-associated CpG islands. Analysis of clustering of potential BMs for the indicated transcription factors within some promoters also suggests that the Alu family contributed to the evolution of transcription cis-regulatory modules in the human genome. It is important that among Alu sequences overlapping with CpG islands in promoters a large fraction of members of the old Alu subfamilies is found, suggesting extensive retroposon-assisted regulatory genome evolution during the divergence of the primates. PMID: 15081116 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 14: Biochim Biophys Acta. 2003 Jul 9;1628(1):22-9. Identification of functional regulatory regions of the connexin32 gene promoter. Field JM, Tate LA, Chipman JK, Minchin SD. School of Biosciences, The University of Birmingham, Edgbaston, B15 2TT, Birmingham, UK. Connexin32 (Cx32) is the predominant gap junction protein expressed in adult rat hepatocytes. This study investigated transcriptional regulation of the rat Cx32 gene in MH(1)C(1) rat hepatoma cells using transient expression assays in conjunction with promoter mutagenesis and 5' nested deletion analysis. Site-directed mutagenesis of the -736 and -187 hepatocyte nuclear factor-1 (HNF-1) sites, the -196 and -116 Sp1 sites, and the -729 and -329 Yin Yang 1 (YY1) sites all significantly reduced promoter activity. We have defined the contribution of each individual site to promoter activity in the intact cell. A novel upstream region of the Cx32 promoter (-1042 to -758) was cloned and shown to contain negative regulatory elements. The transcription factors HNF-1 and Sp1 have important functional roles in the transcriptional regulation of basal and cell-specific Cx32 expression. The multifunctional transcription factor YY1 is also implicated. PMID: 12850269 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 15: J Neurosci Res. 2003 Jul 1;73(1):73-80. Cloning and expression of the rat BACE1 promoter. Lange-Dohna C, Zeitschel U, Gaunitz F, Perez-Polo JR, Bigl V, Rossner S. Department of Neurochemistry, University of Leipzig, Leipzig, Germany. The pathogenic processing of the amyloid precursor protein (APP) into beta-amyloid peptides, which give rise to beta-amyloid plaques in the brains of Alzheimer's disease patients, requires the enzymatic activity of the beta-site APP-cleaving enzyme 1 (BACE1). We report the cloning and sequence of a 1.5-kb DNA fragment upstream of the coding sequence of the rat BACE1 gene and the construction of a BACE1 promoter/luciferase reporter construct. The basal activity of this promoter construct was highest in neuronal cell lines such as BE(2)-C and PC12 and in the pancreatic cell line AR42J, somewhat lower in rat primary neurons, and astrocytic and microglial cultures, very low in hepatocytes, and almost absent in fibroblasts and in the monocyte-macrophage cell line RAW264.7. The first 600 bp of this promoter are highly conserved among rat, mouse, and human, suggesting that this region contains regulatory elements that modulate BACE1 transcription. Indeed, this fragment contains several putative transcription factor binding sites such as MZF1, Sp1, four GATA-1 sites, and one YY1 site. Directed mutagenesis of GATA-1 elements led to altered luciferase expression, indicating that these sites are involved in the regulation of BACE1 transcription. Additionally, the analysis of promoter activities of deletion mutants suggests the presence of activators of BACE1 transcription between bases -514 to -753 and of suppressor elements between bases -754 and -1541. The BACE1 promoter sequence data and the constructs described here will be useful to identify factors that influence the expression of BACE1 in experimental paradigms in vitro. Copyright 2003 Wiley-Liss, Inc. PMID: 12815710 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 16: Biochim Biophys Acta. 2003 Apr 15;1626(1-3):1-9. HOXB7 expression is regulated by the transcription factors NF-Y, YY1, Sp1 and USF-1. Meccia E, Bottero L, Felicetti F, Peschle C, Colombo MP, Care A. Department of Hematology and Oncology, Istituto Superiore di Sanita, Viale Regina Elena, 299- 00161, Rome, Italy. Products of HOX genes are transcription factors responsible for developmental regulation and postnatal tissue homeostasis. Besides their well-established function played during embryonic development, we had previously demonstrated the direct role of HOXB7 in tumor progression through transactivation of several genes involved in the proliferative and angiogenic processes. This role is at first exerted through the deregulated, constitutive expression of this gene. To define the factors possibly responsible for such activation, we studied the molecular regulation of HOXB7 in embryonic and neoplastic cells. In a 1.9-kb 5' promoter region, we identified and functionally tested, at least in vitro, different regulatory sequences showing a direct binding by the NF-Y, YY1, Sp1/Sp3 and upstream stimulatory factor 1 (USF-1) transcription factors. Cell transfection and site-specific mutagenesis demonstrated Sp1/Sp3, NF-Y, YY1 and USF-1 binding to be functional and fundamental in driving HOXB7 expression. Disruption of the corresponding sites reduces gene expression of 65%, 78% and 55%, respectively. Because HOXB7 seems to play an important role in tumor proliferation and progression, the analysis of its regulatory sequences might represent an important step for gene targeting according to a new therapeutic strategy. PMID: 12697323 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 17: Exp Physiol. 2003 Jan;88(1):33-40. Regulation and co-ordination of nuclear gene expression during mitochondrial biogenesis. Goffart S, Wiesner RJ. Institute of Vegetative Physiology, University of Koln, Robert-Koch-Strase 39, 50931 Koln, Germany. steffi.goffart@uni-koeln.de Biogenesis of mitochondria is happening constantly due to the physiological and developmental situation of a cell. As mitochondrial biogenesis is a complex process producing about 20 % of cellular protein, the expression of the 1000 genes involved is expected to be coordinated and regulated tightly. The variety of physiological stimuli and differentiation states lead to the idea of a complex network connecting many different regulatory pathways. By analysing nuclear encoded mitochondrial genes some of the factors involved in the regulation and coordination of mitochondrial gene expression were identified. These factors include general transcription factors such as Sp1 or YY1, as well as transcription factors specific for mitochondrial genes like the nuclear respiratory factors NRF1 and 2. An important control function linked to the physiological situation of a cell is triggered by hormones such as steroid and thyroid hormones. Even cell type-specific regulatory proteins like the myogenin transcription factor family have a strong influence on some mitochondrial genes in the specific cellular background. The regulatory function of most of these proteins can be modulated and enhanced by the coactivators PGC-1a and b and PRC. Although regulatory pathways have been characterized in more detail in recent years, no regulation mechanism has been shown to work on all analysed mitochondrial genes, and the general concept of mitochondrial regulation still remains unclear. Publication Types: Review Review, Tutorial PMID: 12525853 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 18: J Biol Chem. 2003 Mar 14;278(11):9332-8. Epub 2003 Jan 7. Identification of an upstream enhancer in the mouse laminin alpha 1 gene defining its high level of expression in parietal endoderm cells. Niimi T, Hayashi Y, Sekiguchi K. Sekiguchi Biomatrix Signaling Project, ERATO, Japan Science and Technology Corporation, Karimata, Yazako, Nagakute, Aichi 480-1195, Japan. Laminin-1 is the major component of the embryonic basement membrane and consists of alpha1, beta1, and gamma1 chains. The expression of laminin-1 is induced in mouse F9 embryonal carcinoma cells upon differentiation into parietal endoderm through transcriptional up-regulation of the genes encoding these subunits. Here, we identified a 435-bp enhancer in the 5'-flanking region of the mouse laminin alpha1 (LAMA1) gene that activated its transcription in a differentiation-dependent manner. This enhancer was also active in PYS-2 parietal yolk sac-derived cells but not in NIH/3T3 fibroblasts, indicating that it was a parietal endoderm-specific enhancer. This enhancer was also active in Engelbreth-Holm-Swarm (EHS) tumor-derived cells characterized by excessive production of laminin-1 and other basement membrane components, suggesting that EHS tumors have a transcriptional control mechanism similar to that of parietal endoderm cells. Electrophoretic mobility shift analyses revealed four protein binding sites (PBS1-PBS4) in the 435-bp region. However, these DNA-binding proteins were detected not only in parietal endoderm cells (i.e. differentiated F9 cells, PYS-2 cells, and EHS tumor-derived cells) but also in undifferentiated F9 cells and NIH/3T3 cells. Mutational analyses revealed that three of these binding sites (PBS2, PBS3, and PBS4) function synergistically to confer the parietal endoderm-specific enhancer activity. The proteins binding to PBS2 and PBS4 were identified as the Sp1/Sp3 family of transcription factors and YY1, respectively. PMID: 12519763 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 19: Blood. 2003 Feb 1;101(3):1111-7. Epub 2002 Sep 12. The gene encoding the transcriptional regulator Yin Yang 1 (YY1) is a myeloid transforming gene interfering with neutrophilic differentiation. Erkeland SJ, Valkhof M, Heijmans-Antonissen C, Delwel R, Valk PJ, Hermans MH, Touw IP. Department of Hematology, Erasmus University Medical Center, Rotterdam, The Netherlands. The genetic defects underlying the pathogenesis of acute myeloid leukemia (AML) are still largely unknown. Retroviral insertion mutagenesis in mice has become a powerful tool to identify candidate genes involved in the development of leukemia and lymphoma. We have used this strategy with the 1.4 strain of Graffi murine leukemia virus (MuLV), which predominantly causes myeloid leukemias. Here, we report that Graffi-1.4-induced AML frequently harbors virus integrations in the gene encoding the transcription factor Yin Yang 1 (YY1). These integrations occurred in both orientations, and all were located in the 5' promoter region of the gene, 0.5 to 1.5 kb upstream of the major transcriptional start site. Luciferase reporter assays showed that virus integration in this region increases promoter activity and renders it independent of a functional binding site for Sp1, a major transcriptional regulator of YY1. We used the murine 32D model to study the consequence of perturbed YY1 expression for myelopoiesis. YY1 protein levels were high in 32D parental cells maintained in interleukin-3-containing medium, but they dropped when the cells were induced to differentiate by granulocyte-colony-stimulating factor (G-CSF). Strikingly, G-CSF-induced neutrophilic differentiation was reduced in 32D cell transfectants ectopically expressing YY1. In similar experiments on primary bone marrow cells, enforced YY1 expression blocked the outgrowth of CFU-GM colonies. Increased YY1 expression was seen in some cases of human AML. Collectively, these data imply a possible role of perturbed expression of YY1 in the development of AML through interference with the myeloid differentiation program in the leukemic progenitor cells. PMID: 12393438 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 20: Circulation. 2002 Sep 24;106(13):1652-8. Comment in: Circulation. 2003 Mar 25;107(11):e78-9; author reply e78-9. Resveratrol, a polyphenolic phytoalexin present in red wine, enhances expression and activity of endothelial nitric oxide synthase. Wallerath T, Deckert G, Ternes T, Anderson H, Li H, Witte K, Forstermann U. Department of Pharmacology, Johannes Gutenberg University, Mainz, Germany. BACKGROUND: Estrogens can upregulate endothelial nitric oxide synthase (eNOS) in human endothelial cells by increasing eNOS promoter activity and enhancing the binding activity of the transcription factor Sp1. Resveratrol, a polyphenolic phytoalexin found in grapes and wine, has been reported to act as an agonist at the estrogen receptor. Therefore, we tested the effect of this putative phytoestrogen on eNOS expression in human endothelial cells. METHODS AND RESULTS: Incubation of human umbilical vein endothelial cells (HUVEC) and HUVEC-derived EA.hy 926 cells with resveratrol for 24 to 72 hours upregulated eNOS mRNA expression in a time- and concentration-dependent manner (up to 2.8-fold). eNOS protein expression and eNOS-derived NO production were also increased after long-term incubation with resveratrol. Resveratrol increased the activity of the eNOS promoter (3.5-kb fragment) in a concentration-dependent fashion, with the essential trans-stimulated sequence being located in the proximal 263 bp of the promoter sequence. In addition, eNOS mRNA was stabilized by resveratrol. The effect of resveratrol on eNOS expression was not modified by the estrogen receptor antagonists ICI 182780 and RU 58668. In electrophoretic mobility shift assays, nuclear extracts from resveratrol-incubated EA.hy 926 cells showed no enhanced binding activity of the eNOS promoter-relevant transcription factors Sp1, GATA, PEA3, YY1, or Elf-1. In addition to its long-term effects on eNOS expression, resveratrol also enhanced the production of bioactive NO in the short-term (after a 2-minute incubation). CONCLUSIONS: In concert with other effects, the stimulation of eNOS expression and activity may contribute to the cardiovascular protective effects attributed to resveratrol. PMID: 12270858 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 21: Mol Ther. 2002 Sep;6(3):313-20. Erratum in: Mol Ther. 2002 Oct;6(4):563. Comment in: Mol Ther. 2003 Apr;7(4):438-40. Upstream conserved sequences of mouse leukemia viruses are important for high transgene expression in lymphoid and hematopoietic cells. Wahlers A, Kustikova O, Zipfel PF, Itoh K, Koester M, Heberlein C, Li Z, Schiedlmeier B, Skerka C, Fehse B, Baum C. Heinrich-Pette-Institute, Department of Cell and Virus Genetics, 20251 Hamburg, Germany. Highly conserved enhancer sequences located in the upstream part of the long terminal repeat (LTR) of murine leukemia retroviruses (MLV) were reported to compromise viral gene expression in multipotent embryonic cells in vitro and to reduce the likelihood for maintenance of retroviral gene expression in hematopoietic cells in vivo. We show that deletion of these sequences (nucleotides +37 to +95) attenuates rather than increases the transcriptional activity of retroviral vectors in hematopoietic cells almost independently of the developmental lineage (erythroid, myeloid, or lymphoid). Expression rates of modified vectors were reduced by as much as 34-65%, although the strong enhancer array located in the direct repeat of the LTR was preserved. Sequence analysis and electrophoretic mobility shift assays revealed the presence of a highly conserved binding site for NFAT (nuclear factor of activated T cells) proteins that immediately neighbors a known binding site for the transcription factor Yin-Yang1 (YY1) [corrected]. Specific inactivation of the NFAT site reduced transgene expression in all cell types investigated and had a similar effect as the destruction of a neighboring SP1 motif. Combined destruction of individual motifs for NFAT, SP1, and E twenty-six transcription factors (ETS) resulted in a severe attenuation (by 40-60%) of the retroviral enhancer. These results provide novel clues for the manipulation of retrovirus replication and vector tropism. PMID: 12231166 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 22: J Virol. 2002 Aug;76(16):8225-35. Studies of the mechanism of transactivation of the adeno-associated virus p19 promoter by Rep protein. Lackner DF, Muzyczka N. Department of Molecular Genetics and Microbiology and University of Florida Gene Therapy Center, College of Medicine, University of Florida, Gainesville 32610, USA. During adeno-associated virus (AAV) type 2 productive infections, the p19 promoter of AAV is activated by the AAV Rep78 and Rep68 proteins. Rep-induced activation of p19 depends on the presence of one of several redundant Rep binding elements (RBEs) within the p5 promoter or within the terminal repeats (TR). In the absence of the TR, the p5 RBE and the p19 Sp1 site at position -50 are essential for p19 transactivation. To determine how a Rep complex bound at p5 induces transcription at p19, we made a series of p19 promoter chloramphenicol acetyltransferase constructs in which the p5 RBE was inserted at different locations upstream or downstream of the p19 mRNA start site. The RBE acted like a repressor element at most positions in the presence of both Rep and adenovirus (Ad), and the level of repression increased dramatically as the RBE was inserted closer to the p19 promoter. We concluded that the RBE by itself was not a conventional upstream activation signal and instead behaved like a repressor. To understand how the Rep-RBE complex within p5 activated p19, we considered the possibility that its role was to function as an architectural protein whose purpose was to bring other p5 transcriptional elements to the p19 promoter. In order to address this possibility, we replaced both the p5 RBE and the p19 Sp1 site with GAL4 binding sites. The modified GAL4-containing constructs were cotransfected with plasmids that expressed GAL4 fusion proteins capable of interacting through p53 and T-antigen (T-ag) protein domains. In the presence of Ad and the GAL4 fusion proteins, the p19 promoter exhibited strong transcriptional activation that was dependent on both the GAL4 fusion proteins and Ad infection. This suggested that the primary role of the p5 RBE and the p19 Sp1 sites was to act as a scaffold for bringing transcription complexes in the p5 promoter into close proximity with the p19 promoter. Since Rep and Sp1 themselves were not essential for transactivation, we tested mutants within the other p5 transcriptional elements in the context of GAL4-induced looping to determine which of the other p5 elements was necessary for p19 induction. Mutation of the p5 major late-transcription factor site reduced p19 activity but did not eliminate induction in the presence of the GAL4 fusion proteins. However, mutation of the p5 YY1 site at position -60 (YY1-60) eliminated GAL4-induced transactivation. This implicated the YY1-60 protein complexes in p19 induction by Rep. In addition, both basal p19 activity and activity in the presence of Ad increased when the YY1-60 site was mutated even in the absence of Rep or GAL4 fusion proteins. Therefore, there are likely to be alternative p5-p19 interactions that are Rep independent in which the YY1-60 complex inhibits p19 transcription. We concluded that transcriptional control of the p19 promoter was dependent on the formation of complexes between the p5 and p19 promoters and that activation of the p19 promoter depends largely on the ability of Rep and Sp1 to form a scaffold that positions the p5 YY1 complex near the p19 promoter. PMID: 12134028 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 23: Mol Cell Endocrinol. 2002 Mar 28;189(1-2):191-9. Characterization of promoter 1B in the human glucocorticoid receptor gene. Nunez BS, Vedeckis WV. Department of Biochemistry and Molecular Biology, Stanley S. Scott Cancer Center, Louisiana State University Health Sciences Center, 533 Bolivar Street, New Orleans, LA 70112, USA. nunez@utmsi.utexas.edu At least three different promoter regions (1A, 1B, and 1C) are involved in the expression of the human GR gene. Promoters 1B and 1C are found in a 2800 bp region of DNA immediately upstream of the exon 1C transcriptional initiation site. Transcripts containing either exon 1B or 1C are expressed in a wide variety of human tissues and cultured cells. Luciferase reporter constructs were created containing promoter 1B plus 1C (-2738 to +19), promoter 1B (-2738 to -1046) alone, or promoter 1C (-1045 to +19) alone. All three constructs were equally effective in driving luciferase expression in HeLa (human cervical carcinoma) cells. In Jurkat (human T-cell acute lymphoblastic leukemia) cells, constructs containing promoters 1B plus 1C or promoter 1B were equally active, but the promoters 1B plus 1C construct was 35% more active than the promoter 1C construct. However, in HepG2 (human hepatoma) cells, the promoter 1C construct was as effective as promoters 1B plus 1C and more than twice as effective as promoter 1B. Sequences that reside proximal to the exon 1B transcriptional start site included three Sp1 (FP2-FP4) sites. Another site (FP1) contains the sequence TGATAG, which strongly resembles the consensus binding sequence for the GATA family of transcription factors. However, oligonucleotide competition and supershift analysis of FP1 indicates that this site is not a binding site for GATA proteins. These four sites are in addition to three YY1 and one Sp1 sites previously reported in promoter 1B. In HeLa cells, deletion of the three YY1 sites results in only a 30% loss of activity and substantial loss of activity occurs only after deletion of all four Sp1 sites, indicating the critical importance of Sp1 in GR expression in these cells. In contrast, the elimination of the three YY1 sites results in a dramatic decrease in promoter strength in both HepG2 and Jurkat cells (64 and 77%, respectively), while subsequent deletions of promoter elements do not result in substantial changes in promoter activity in these cell lines. This study shows that both promoters 1B and 1C are important for the ubiquitous expression of the human GR gene. Differences in the utilization of these promoters in various cell types are likely a reflection of different promoter availability and/or the levels of specific transcription factors in the cell. This could contribute to tissue-specific expression of GR levels in different cell types. PMID: 12039077 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 24: Biochim Biophys Acta. 2002 Mar 19;1574(2):164-74. YY1 and Sp1 activate transcription of the human NDUFS8 gene encoding the mitochondrial complex I TYKY subunit. Lescuyer P, Martinez P, Lunardi J. Laboratoire BECP-EA2943 UJF/LRA6V CEA-DBMS, CEA Grenoble, 17 rue des Martyrs, 38054 Cedex 9, Grenoble, France. plescuyer@cea.fr Complex I is the most complicated of the multimeric enzymes that constitute the mitochondrial respiratory chain. It is encoded by both mitochondrial and nuclear genomes. We have previously characterized the human NDUFS8 gene that encodes the TYKY subunit. This essential subunit is thought to participate in the electron transfer and proton pumping activities of complex I. Here, we have analyzed the transcriptional regulation of the NDUFS8 gene. Using primer extension assays, we have identified two transcription start sites. The basal promoter was mapped to a 247 bp sequence upstream from the main transcription start site by reporter gene analysis in HeLa cells and in differentiated or non-differentiated C2C12 cells. Three Sp1 sites and one YY1 site were identified in this minimal promoter. Through gel shift analysis, all sites were shown to bind to their cognate transcription factors. Site-directed mutagenesis revealed that the YY1 site and two upstream adjacent Sp1 sites drive most of the promoter activity. This work represents the first promoter analysis for a complex I gene. Together with previous studies, our results indicate that YY1 and Sp1 control the expression of genes encoding proteins that are involved in almost all steps of the oxidative phosphorylation metabolism. PMID: 11955626 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 25: Gene. 2002 Mar 6;286(1):81-9. Transcriptional activators and coactivators in the nuclear control of mitochondrial function in mammalian cells. Scarpulla RC. Department of Cell and Molecular Biology, Northwestern Medical School, 303 East Chicago Avenue, Chicago, IL 60611, USA. rsc248@nwu.edu The biogenesis and function of mitochondria rely upon the regulated expression of nuclear genes. Recent evidence points to both transcriptional activators and coactivators as important mediators of mitochondrial maintenance and proliferation. Several sequence-specific activators including NRF-1, NRF-2, Sp1, YY1, CREB and MEF-2/E-box factors, among others, have been implicated in respiratory chain expression. Notably, recognition sites for NRF-1, NRF-2 and Sp1 are common to most nuclear genes encoding respiratory subunits, mitochondrial transcription and replication factors, as well as certain heme biosynthetic enzymes and components of the protein import machinery. Moreover, genetic evidence supports a role for NRF-1 in the maintenance of mtDNA during embryonic development. Despite these advances, the means by which multiple transcription factors are integrated into a program of mitochondrial biogenesis remains an open question. New insight into this problem came with the discovery of the transcriptional coactivator, PGC-1. This cofactor is cold inducible in brown fat and interacts with multiple transcription factors to orchestrate a program of adaptive thermogenesis. As part of this program, PGC-1 can up-regulate nuclear genes that are required for mitochondrial biogenesis in part through a direct interaction with NRF-1. Ectopic expression of PGC-1 induces the expression of respiratory subunit mRNAs and leads to mitochondrial proliferation in both cultured cells and transgenic mice. More recently, PRC was characterized as a novel coactivator that shares certain structural similarities with PGC-1 including an activation domain, an RS domain and an RNA recognition motif. However, unlike PGC-1, PRC is not induced significantly during thermogenesis but rather is cell-cycle regulated in cultured cells under conditions where PGC-1 is not expressed. PRC has a transcriptional specificity that is very similar to PGC-1, especially in its interaction with NRF-1 and in the activation of NRF-1 target genes. These regulated coactivators may provide a means for integrating sequence-specific activators in the biogenesis and function of mitochondria under diverse physiological conditions. PMID: 11943463 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 26: J Virol. 2002 Jan;76(1):313-26. Requirement of multiple cis-acting elements in the human cytomegalovirus major immediate-early distal enhancer for viral gene expression and replication. Meier JL, Keller MJ, McCoy JJ. Department of Internal Medicine and the Helen C. Levitt Center for Viral Pathogenesis and Disease, University of Iowa College of Medicine, Iowa City, Iowa, 52242, USA. jeffery-meier@uiowa.edu We have shown previously that the human cytomegalovirus (HCMV) major immediate-early (MIE) distal enhancer is needed for MIE promoter-dependent transcription and viral replication at low multiplicities of infection (MOI). To understand how this region works, we constructed and analyzed a series of HCMVs with various distal enhancer mutations. We show that the distal enhancer is composed of at least two parts that function independently to coordinately activate MIE promoter-dependent transcription and viral replication. One such part is contained in a 47-bp segment that has consensus binding sites for CREB/ATF, SP1, and YY1. At low MOI, these working parts likely function in cis to directly activate MIE gene expression, thus allowing viral replication to ensue. Three findings support the view that these working parts are likely cis-acting elements. (i) Deletion of either part of a bisegmented distal enhancer only slightly alters MIE gene transcription and viral replication. (ii) Reversing the distal enhancer's orientation largely preserves MIE gene transcription and viral replication. (iii) Placement of stop codons at -300 or -345 in all reading frames does not impair MIE gene transcription and viral replication. Lastly, we show that these working parts are dispensable at high MOI, partly because of compensatory stimulation of MIE promoter activity and viral replication that is induced by a virion-associated component(s) present at a high viral particle/cell ratio. We conclude that the distal enhancer is a complex multicomponent cis-acting region that is required to augment both MIE promoter-dependent transcription and HCMV replication. PMID: 11739696 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 27: J Clin Virol. 2001 Dec;23(1-2):65-77. Intratype HPV16 sequence variation within LCR of isolates from asymptomatic carriers and cervical cancers. Schmidt M, Kedzia W, Gozdzicka-Jozefiak A. Department of Molecular Virology, Institute of Molecular Biology and Biotechnology, Adam Mickiewicz University, Miedzychodzka 5, 60-371 Poznan, Poland. mschmidt@amu.edu.pl BACKGROUND: HPV16 is a predominant type of virus identified in genital lesions and strongly associated with the development of genital cancers. Infection with the virus is considered to be the main risk factor in the development of cervical cancer. Based on HPV16 DNA isolated from invasive cancers, a classification of intratype genetic variants was established and the strains were designated according to geographical regions. The HPV16 variants classification was based on isolates derived from cancers. OBJECTIVES: Analysis of HPV16 LCR variants isolated from asymptomatic carriers for comparison with cervical cancer isolates to examine whether a correlation can be found between cervical epithelium state and variant of HPV16 it carries. MATERIALS AND METHODS: The HPV16 LCR fragments were amplified by PCR using DNA isolated from cervical swabs and tissue sections then screened for nucleotide changes by SSCP. Polymorphic sites were analysed for regulatory protein binding properties by EMSA. RESULTS: Comparison of the two groups revealed that isolates from cervical cancers predominantly carry changes in sequences of YY1 binding sites (especially at nucleotide 7519), while variants from asymptomatic carriers contained nucleotide changes within or close to transcription binding sites for AP-1, Oct-1, NF1, Tef-1, Tef-2, Sp1, YY1 and viral E2. EMSA study showed that sequence changes in the segment alter binding and formation of transcriptional complexes in quantitative and/or qualitative manner and so they may inflict viral activity. CONCLUSION: The results of our study show that there might be HPV16 variants of decreased oncogenic potential therefore infection with such variants can recede. PMID: 11595585 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 28: Life Sci. 2001 Jun 1;69(2):201-12. Transcriptional regulation of galectin-10 (eosinophil Charcot-Leyden crystal protein): a GC box (-44 to -50) controls butyric acid induction of gene expression. Dyer KD, Rosenberg HF. Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA. Galectin-10 (gal-10, also known as Charcot-Leyden crystal protein) is a member of the galectin family of beta-galactoside binding proteins that is expressed uniquely in eosinophilic and basophilic leukocytes. To gain a better understanding of galectin gene expression, we present an analysis of the transcriptional regulation of the gene encoding gal-10. Analysis of the minimal promoter revealed nine consensus-binding sites for transcription factors, including several that are also found in the minimal promoters of galectins -1, -2, and -3. The decrease in gal-10 promoter activity after disruption of either the GC box (-44 to -50) or the Oct site (-255 to -261) suggests that these sites, along with the previously characterized GATA and EoTF sites, are necessary for full promoter activity. By supershift analysis, we demonstrate binding of the transcription factors Sp1 and Oct1 to the consensus GC box and the Oct site, respectively. Similar to gal-1, gal-10 expression is induced by butyric acid, an effect that is lost upon ablation of the GC box. Additionally, we demonstrate AML3 binding to the consensus AML site and YY1 binding to the Inr sequence, both elements functioning as silencers in the gal-10 promoter. PMID: 11441910 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 29: Arch Med Res. 2001 Jul-Aug;32(4):263-7. Expression of TCF, TPF/YY1, and the Sp family transcription factors in rabbit endometrium throughout pregnancy. Arias J, Hernandez A, Barron A, Castro I. Departamento de Bioquimica y Biologia Molecular, Instituto Nacional de Perinatologia, Mexico City, Mexico. BACKGROUND: TCF, TPF/YY1, and the Sp family are specific transcription factors that bind sequences found within the uteroglobin (UG) gene promoter region that are necessary for transcription. To date, UG gene expression and regulation in vivo are not fully understood. The purpose of this study was to assess the expression patterns of these factors in the rabbit endometrium throughout pregnancy. METHODS: Endometrial nuclear extracts were obtained from female rabbits on days 0, 3, 5, 7, 15, and 28 after mating. Transcription factor expression was assessed by DNA-protein binding assays using endometrial nuclear proteins and specific oligonucleotides. Band shifts were observed on 4% acrylamide gels and analyzed by densitometry. RESULTS: The expression patterns of the transcription factors analyzed here differed, as TPF/YY1 and Sp3/SpR-2 were expressed constitutively while TCF and Sp1 showed variable expression patterns throughout pregnancy. CONCLUSIONS: Our results suggest that UG gene expression in the intact pregnant rabbit is controlled by two constitutive and two regulated factors, and that the DNA-binding sites are located at the TATA box and the GT1 sites within the gene promoter. PMID: 11440780 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 30: J Gen Virol. 2001 Jun;82(Pt 6):1473-80. Cellular transcription factors that interact with p6 promoter elements of parvovirus B19. Raab U, Bauer B, Gigler A, Beckenlehner K, Wolf H, Modrow S. Institut fur Medizinische Mikrobiologie und Hygiene, Universitat Regensburg, Franz-Josef-Strauss-Allee 11, D-93053 Regensburg, Germany. All transcripts of the human parvovirus B19 identified so far are regulated by a single promoter at map unit 6 of the viral genome, the so-called p6 promoter. This promoter is active in a wide variety of different cells. In order to identify cellular transcription factors involved in regulating promoter activity, we performed gel-retardation and supershift assays using the parts of the p6 promoter sequence shown previously to be protected in footprint experiments. Thereby, binding was demonstrated of the Oct-1 protein to an octamer motif within the p6 promoter and of the transcription factor Sp1 to three GC boxes. A specific preferential interaction of the factor Sp3 with one of these boxes was observed, indicating that the ratio Sp1:Sp3 may be involved in the regulation of promoter activity. Consensus sites for the regulatory protein YY1 are located close to the GC boxes and the octamer motif, to which this factor binds efficiently. PMID: 11369893 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 31: J Biol Chem. 2001 Apr 27;276(17):13664-74. Epub 2001 Jan 26. The p38 mitogen-activated kinase pathway regulates the human interleukin-10 promoter via the activation of Sp1 transcription factor in lipopolysaccharide-stimulated human macrophages. Ma W, Lim W, Gee K, Aucoin S, Nandan D, Kozlowski M, Diaz-Mitoma F, Kumar A. Department of Pediatrics, University of Ottawa, Ottawa, Ontario K1H 8L1, Canada. Interleukin-10 (IL-10), a pleiotropic cytokine that inhibits inflammatory and cell-mediated immune responses, is produced by a wide variety of cell types including T and B cells and monocytes/macrophages. Regulation of pro- and anti-inflammatory cytokines has been suggested to involve distinct signaling pathways. In this study, we investigated the regulation of the human IL-10 (hIL-10) promoter in the human monocytic cell line THP-1 following activation with lipopolysaccharide (LPS). Analysis of hIL-10 promoter sequences revealed that DNA sequences located between base pairs -652 and -571 are necessary for IL-10 transcription. A computer analysis of the promoter sequence between base pairs -652 and -571 revealed the existence of consensus sequences for Sp1, PEA1, YY1, and Epstein-Barr virus-specific nuclear antigen-2 (EBNA-2)-like transcription factors. THP-1 cells transfected with a plasmid containing mutant Sp1 abrogated the promoter activity, whereas plasmids containing the sequences for PEA1, YY1, and EBNA-2-like transcription factors did not influence hIL-10 promoter activity. To understand the events upstream of Sp1 activation, we investigated the role of p38 and extracellular signal-regulated kinase mitogen-activated protein kinases by using their specific inhibitors. SB202190 and SB203580, the p38-specific inhibitors, inhibited LPS-induced IL-10 production. In contrast, PD98059, a specific inhibitor of extracellular signal-regulated kinase kinases, failed to modulate IL-10 production. Furthermore, SB203580 inhibited LPS-induced activation of Sp1, as well as the promoter activity in cells transfected with a plasmid containing the Sp1 consensus sequence. These results suggest that p38 mitogen-activated protein kinase regulates LPS-induced activation of Sp1, which in turn regulates transcription of the hIL-10 gene. PMID: 11278848 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 32: Gene. 2000 Dec 31;261(2):311-20. Isolation of a bi-directional promoter directing expression of the mouse GABPalpha and ATP synthase coupling factor 6 genes. Chinenov Y, Coombs C, Martin ME. Department of Biochemistry, University of Missouri at Columbia, MO, Columbia 65212, USA. The GA-binding protein (GABP) is a ubiquitous heteromeric transcription factor implicated in the regulation of several genes involved in mitochondrial energy metabolism including subunits of cytochrome c oxidase, ATP synthase, and mitochondrial transcription factor 1 (mtTF1). GABPalpha subunit binds the PEA3/Ets binding sites (EBS), while GABPbeta contains a transcription activation domain and mediates alphabeta dimer and alpha(2)beta(2) tetramer formation essential for activation of transcription. Here we report the cloning of 2449 bp of the mouse (m) GABPalpha promoter region including 201 bp of the 5' end of the published mGABPalpha cDNA sequence. Surprisingly, sequences homologous to the 5'UTR of mouse, rat and human mitochondrial ATP synthase coupling factor 6 (ATPsynCF6) cDNAs were found165-240 bp upstream of the mGABPalpha cDNA. A search of the non-redundant nucleotide database revealed a human genomic sequence derived from chromosome 21 (21q22) bearing significant homology to the mGABPalpha/ATPsynCF6 promoter region and encompassed the entire hGABPalpha and hATPsynCF6 genes. Primer extension analysis revealed multiple transcription start sites for both mGABPalpha and mATPsynCF6 mRNAs that mapped near the published cDNA 5' ends. Sequence analysis identified several binding sites upstream of the GABPalpha cDNA sequence including sites for GABP (-86, -104, -169, -257, and -994), YY1 (-57), Sp1 (-242 and -226), and NRF1 (-5). No 'TATA' motif was identified near either the GABPalpha or ATPsynCF6 transcription start sites. The human and mouse promoters retain significant sequence identity including binding sites for several tissue-specific transcription factors. Transient transfection assays using Luciferase reporter constructs containing the intergenic region and flanking sequences confirmed that this region of DNA promotes transcription in both directions. PMID: 11167019 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 33: Endocrinology. 2001 Jan;142(1):49-58. Yin yang 1 protein negatively regulates high-density lipoprotein receptor gene transcription by disrupting binding of sterol regulatory element binding protein to the sterol regulatory element. Shea-Eaton W, Lopez D, McLean MP. Departments of Obstetrics and Gynecology and Biochemistry and Molecular Biology, University of South Florida, College of Medicine, Tampa, Florida 33606, USA. Because the high-density lipoprotein receptor (HDL-R) is a key element in cholesterol homeostasis and a potential therapeutic target for hypercholesterolemic drugs, an understanding of HDL-R regulation is essential. The sterol regulatory element (SRE) binding protein-1a (SREBP-1a) was shown to positively regulate HDL-R gene expression through two SREs. SREBP-1a requires the presence of a coactivator like simian-virus-40-protein-1 (Sp1) to promote maximum activation of the HDL-R promoter. Negative regulatory factors are also known to play a role in cholesterol homeostasis, and the ubiquitous Yin Yang-1 zinc finger transcription factor (YY1) has been shown to repress several sterol-responsive gene promoters. A search of the rat HDL-R promoter revealed two putative YY1 binding sites (distal, -1329 to -1321; proximal, -1211 to -1203). Upon removal of both YY1 binding sites, YY1 was unable to repress HDL-R activation under basal (unstimulated) promoter conditions. However, YY1 was still an efficient transcriptional repressor for SREBP-1a-induced activation. YY1 was able to attenuate the transcriptional synergy caused by the combined actions of SREBP-1a and Sp1. Two-hybrid studies confirmed that YY1 bound with high affinity to SREBP-1a, and mobility shift assays demonstrated that YY1 could disrupt SREBP-1a binding to both SREs. The molecular consequence of YY1 intervention seems to override any positive interactions between Sp-1 and SREBP-1a and results in the disruption of SREBP-1a binding to the SREs in the HDL-R promoter. PMID: 11145566 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 34: Biochim Biophys Acta. 2000 Dec 15;1517(1):119-27. Upstream stimulatory factor 2 stimulates transcription through an initiator element in the mouse cytochrome c oxidase subunit Vb promoter. Breen GA, Jordan EM. Department of Molecular and Cell Biology, The University of Texas at Dallas, P.O. Box 830688, Richardson, TX 75083-0688, USA. breen@utdallas.edu Upstream stimulatory factor (USF) is a basic helix-loop-helix-leucine zipper transcription factor that plays an important role in transcriptional activation and cell proliferation. In this article, we demonstrate that the mouse cytochrome c oxidase subunit Vb gene (Cox5b) can be transactivated by ectopic expression of USF2 through an initiator (Inr) element in the core promoter. Importantly, using a dominant-negative mutant of USF2, we demonstrate the role of endogenous USF2 proteins in the transcriptional activation of the Cox5b Inr. Domains of USF2 encoded by exon 4, exon 5 and the USF-specific region are important for maximum activation of the Cox5b Inr. Using the adenovirus E1A oncoprotein, we show that p300/CBP acts as a coactivator in the USF2-dependent activation of the Cox5b Inr. We also demonstrate that although expression of multifunctional regulatory factor, Yin Yang 1 (YY1), can stimulate transcription of the Cox5b Inr to a modest extent, expression of YY1 together with USF2 greatly reduces the level of activation of the Cox5b Inr. Furthermore, we show that the transcription factor, Sp1, represses both the YY1- and the USF2-dependent activation of the Cox5b Inr, indicating competition among Sp1, YY1, and USF2. PMID: 11118624 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 35: Virology. 2000 Jun 20;272(1):191-203. Two leaky-late HSV-1 promoters differ significantly in structural architecture. Lieu PT, Wagner EK. Program in Animal Virology, University of California, Irvine, California 92697-3900, USA. The HSV-1 VP5 and VP16 transcripts are expressed with leaky-late (gamma1) kinetics and reach maximal levels after viral DNA replication. While the minimal VP5 promoter includes only an Sp1 site at -48, a TATA box at -30, and an initiator (Inr) element at the cap site, here we show that elements upstream of -48 can functionally compensate for the mutational loss of the critical Sp1 site at -48. To determine whether this is a general feature of leaky-late promoters, we have carried out a detailed analysis of the VP16 promoter in the context of the viral genome at the gC locus. Sequence analysis suggests a great deal of similarity between the two. Despite this, however, mutational analysis revealed that the 5' boundary of the VP16 promoter extends to ca. -90. This region includes an Sp1 binding site at -46, CAAT box homology at -77, and "E box" (CACGTG) at -85. Mutational and deletional analyses demonstrate that the proximal Sp1 site plays little or no role in promoter strength; despite this it can be shown to bind Sp1 protein using DNA mobility shift assays. Like the VP5 promoter, the VP16 promoter also requires an initiator element at the cap site. The VP16 Inr element differs in sequence from that of the VP5 promoter, and its deletion or mutation has a significantly smaller effect on promoter strength. The difference between these two Inr elements was confirmed by our finding that the VP16 initiator element binds to the 65-kDa YY1 transcription factor, and the VP5 Inr element competes poorly for the binding between the VP16 element and infected cell proteins in comparative bandshift assays. While the VP16 Inr sequence is identical to that of several murine TATA-less promoters, the VP16 Inr requires a TATA box for measurable activity. Copyright 2000 Academic Press. PMID: 10873762 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 36: Virus Res. 1999 Jul;62(1):77-88. Influence of different cellular transcription factors on the regulation of varicella-zoster virus glycoproteins E (gE) and I (gI) UTR's activity. Rahaus M, Wolff MH. Institute of Microbiology and Virology, University of Witten Herdecke, Germany. Varicella-zoster virus (VZV) glycoproteins E (ORF 68) and I (ORF 67) are members of late genes. They belong to the major components of the virion envelope and can be found on the host cell surface as well. To get further insights in the regulation of gE and gI expression, which are known to be activated by IE4 and IE62, we analysed the intergenic regions of ORF 66/67 and ORF 67/68, containing the putative promoters of gI and gE. We have mapped the transcriptional start site of gE and have identified an extensive set of eucaryotic cis-elements: typical TATA- and CAAT-motifs and further regulatory sequences to facilitate interaction with eucaryotic transcription factors. Reporter constructs have been made using the intergenic regions of ORF 66/67 and ORF 67/68 as promoter elements. In cis-trans interaction studies, an influence on the regulation of transcription and reporter gene expression of overexpressed transfactors, LAP/LIP, Sp1, YY1 and NF-E2 has become measurable. In addition, protein-DNA binding assays using both gE- and gI-intergenic regions and cellular extracts from different VZV-permissive cells have suggested a binding of a 32 and 18 kD protein. In conclusion, these data indicate an involvement of common cellular transcription factors in the regulation of VZV late gene expression. PMID: 10513289 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 37: Eur J Biochem. 1999 Oct;265(2):501-23. Serum amyloid A, the major vertebrate acute-phase reactant. Uhlar CM, Whitehead AS. Department of Pharmacology and Center for Pharmacogenetics, University of Pennsylvania School of Medicine, Philadelphia 19104-6084, USA. The serum amyloid A (SAA) family comprises a number of differentially expressed apolipoproteins, acute-phase SAAs (A-SAAs) and constitutive SAAs (C-SAAs). A-SAAs are major acute-phase reactants, the in vivo concentrations of which increase by as much as 1000-fold during inflammation. A-SAA mRNAs or proteins have been identified in all vertebrates investigated to date and are highly conserved. In contrast, C-SAAs are induced minimally, if at all, during the acute-phase response and have only been found in human and mouse. Although the liver is the primary site of synthesis of both A-SAA and C-SAA, extrahepatic production has been reported for most family members in most of the mammalian species studied. In vitro, the dramatic induction of A-SAA mRNA in response to pro-inflammatory stimuli is due largely to the synergistic effects of cytokine signaling pathways, principally those of the interleukin-1 and interleukin-6 type cytokines. This induction can be enhanced by glucocorticoids. Studies of the A-SAA promoters in several mammalian species have identified a range of transcription factors that are variously involved in defining both cytokine responsiveness and cell specificity. These include NF-kappaB, C/EBP, YY1, AP-2, SAF and Sp1. A-SAA is also post-transcriptionally regulated. Although the precise role of A-SAA in host defense during inflammation has not been defined, many potential clinically important functions have been proposed for individual SAA family members. These include involvement in lipid metabolism/transport, induction of extracellular-matrix-degrading enzymes, and chemotactic recruitment of inflammatory cells to sites of inflammation. A-SAA is potentially involved in the pathogenesis of several chronic inflammatory diseases: it is the precursor of the amyloid A protein deposited in amyloid A amyloidosis, and it has also been implicated in the pathogenesis of atheroscelerosis and rheumatoid arthritis. Publication Types: Review Review, Tutorial PMID: 10504381 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 38: J Gen Virol. 1999 Aug;80 ( Pt 8):2097-101. Overlapping YY1- and aberrant SP1-binding sites proximal to the early promoter of human papillomavirus type 16. Dong XP, Pfister H. Institute of Virology, Universitat zu Koln, Germany. Transcription of oncogenes E6 and E7 of human papillomavirus type 16 (HPV-16) from the P97 promoter is regulated by viral and cellular proteins. The transcription factor YY1 represses transcription through binding to cognate sequences in the long control region (LCR). In HPV-16 DNA from cervical carcinomas, mutations of YY1-binding sites have been identified that increase P97 activity 3-6-fold. A second, SP1-binding site has now been identified in the HPV-16 LCR (nt 7842-7847), which overlaps the YY1-binding site at positions 7840-7848. A point mutation within this YY1 site in viral DNA from a cervical cancer, previously shown to prevent YY1 binding, was shown to increase SP1 binding and P97 activity 4.7-fold. An engineered mutant eliminating SP1 binding showed only 1- to 1.6-fold increased P97 activity. It is concluded that competition between SP1 and YY1 for DNA binding plays a major role in YY1 repression mediated by the binding site at positions 7840-7848. PMID: 10466808 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 39: Gene. 1999 Aug 20;236(2):197-208. Unlocking the mechanisms of transcription factor YY1: are chromatin modifying enzymes the key? Thomas MJ, Seto E. Molecular Oncology Program, H. Lee Moffitt Cancer Center & Research Institute, University of South Florida, Tampa, FL 33612, USA. The transcription factor YY1 is a complex protein that is involved in repressing and activating a diverse number of promoters. Numerous studies have attempted to understand how this one factor can act both as a repressor and an activator in such a wide set of different contexts. The fact that YY1 interacts with a number of key regulatory proteins (e.g. TBP, TFIIB, TAFII55, Sp1, and E1A) has suggested that these interactions are important for determining which particular function of YY1 is displayed at a specific promoter. Two groups of proteins, previously known to function as corepressors and coactivators, that now seem likely to modulate YY1's functions, are the histone deacetylases (HDAC) and histone acetyltransferases (HAT). These two groups of enzymes modify histones, and this modification is proposed to alter chromatin structure. Acetylated histones are typically localized to active chromatin while deacetylated histones colocalize with transcriptionally inactive chromatin. When these enzymes are directed to a promoter through a DNA binding factor such as YY1, that promoter can be activated or repressed. This review will discuss the recent work dealing with the different proteins that interact with YY1, with particular emphasis on ones that modify chromatin, and how they could be involved in regulating YY1's activities. Publication Types: Review Review, Tutorial PMID: 10452940 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 40: Gene. 1999 Jul 8;234(2):337-44. Characterization of the murine gene encoding 1-Cys peroxiredoxin and identification of highly homologous genes. Lee TH, Yu SL, Kim SU, Kim YM, Choi I, Kang SW, Rhee SG, Yu DY. Korea Research Institute of Bioscience and Biotechnology, PO Box 115, Yusong, Taejon 305-600, South Korea. A new type of peroxiredoxin, named 1-Cys peroxiredoxin (1-Cys Prx), reduces hydrogen peroxide with the use of electrons from unidentified electron donor(s). We have isolated the mouse gene encoding 1-Cys Prx (CP-3) and shown that it is comprised of five exons and four introns. Analysis of 5' flanking regions revealed binding sequences of several putative transcription factors such as Sp1, Pit-1a, c-Jun, c-Myc and YY1. It is noticeable that several potential Sp1 binding sites assigned the -60 through -96bp from putative transcription initiation site. The gel shift assays showed that Sp1 and Pit-1a bind specifically to each binding site in 1-Cys Prx promoter. We also isolated two highly related genes such as CP-2 and CP-5. These genes are encoded by single exons, and show 85% of nucleotide sequence homology with the CP-3. The structural features of these genes suggest that they might be intronless genes derived from the CP-3 by the mechanism involving retrotransposition. In addition, our data suggest that they are inserted to a specific site of the mouse L1 repetitive element. The 1-Cys Prx was actively transcribed in a variety of adult tissues as well as in the developing embryos. These results suggest that only the 1-Cys Prx gene might be relevant for studying the function of the 1-Cys Prx in the murine system. PMID: 10395907 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 41: J Biol Chem. 1999 May 7;274(19):13025-32. Co-stimulation of promoter for low density lipoprotein receptor gene by sterol regulatory element-binding protein and Sp1 is specifically disrupted by the yin yang 1 protein. Bennett MK, Ngo TT, Athanikar JN, Rosenfeld JM, Osborne TF. Department of Molecular Biology and Biochemistry, University of California, Irvine, California 92697, USA. Sterol regulation of gene expression in mammalian cells is mediated by an interaction between the cholesterol-sensitive sterol regulatory element-binding proteins (SREBPs) and promoter-specific but generic co-regulatory transcription factors such as Sp1 and NF-Y/CBF. Thus, sterol-regulated promoters that require different co-regulatory factors could be regulated independently through targeting the specific interaction between the SREBPs and the individual co-regulatory proteins. In the present studies we demonstrate that transiently expressed yin yang 1 protein (YY1) inhibits the SREBP-mediated activation of the low density lipoprotein (LDL) receptor in a sensitive and dose-dependent manner. The inhibition is independent of YY1 binding directly to the LDL receptor promoter, and we show that the same region of YY1 that interacts in solution with Sp1 also interacts with SREBP. Furthermore, other SREBP-regulated genes that are not co-regulated by Sp1 are either not affected at all or are not as sensitive to the repression. Thus, the specific interaction that occurs between SREBPs and Sp1 to stimulate the LDL receptor promoter is a specific target for inhibition by the YY1 protein, and we provide evidence that the mechanism can be at least partially explained by the ability of YY1 to inhibit the interaction between SREBP and Sp1 in solution in vitro. The LDL receptor is the key gene of cholesterol uptake, and the rate-controlling genes of cholesterol synthesis are stimulated by the concerted action of SREBPs along with coregulators that are distinct from Sp1. Therefore, repression of gene expression through specifically targeting the interaction between SREBP and Sp1 would provide a molecular mechanism to explain how cholesterol uptake can be regulated independently from cholesterol biosynthesis in mammalian cells. PMID: 10224053 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 42: Mol Cell Biol. 1999 Apr;19(4):2724-33. Human SWI-SNF component BRG1 represses transcription of the c-fos gene. Murphy DJ, Hardy S, Engel DA. Department of Microbiology and Cancer Center, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA. Yeast and mammalian SWI-SNF complexes regulate transcription through active modification of chromatin structure. Human SW-13 adenocarcinoma cells lack BRG1 protein, a component of SWI-SNF that has a DNA-dependent ATPase activity essential for SWI-SNF function. Expression of BRG1 in SW-13 cells potentiated transcriptional activation by the glucocorticoid receptor, which is known to require SWI-SNF function. BRG1 also specifically repressed transcription from a transfected c-fos promoter and correspondingly blocked transcriptional activation of the endogenous c-fos gene. Mutation of lysine residue 798 in the DNA-dependent ATPase domain of BRG1 significantly reduced its ability to repress c-fos transcription. Repression by BRG1 required the cyclic AMP response element of the c-fos promoter but not nearby binding sites for Sp1, YY1, or TFII-I. Using human C33A cervical carcinoma cells, which lack BRG1 and also express a nonfunctional Rb protein, transcriptional repression by BRG1 was weak unless wild-type Rb was also supplied. Interestingly, Rb-dependent repression by BRG1 was found to take place through a pathway that is independent of transcription factor E2F. PMID: 10082538 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 43: J Steroid Biochem Mol Biol. 1998 Dec;67(5-6):369-81. The human glucocorticoid receptor promoter upstream sequences contain binding sites for the ubiquitous transcription factor, Yin Yang 1. Breslin MB, Vedeckis WV. Department of Biochemistry and Molecular Biology and Stanley S. Scott Cancer Center, Louisiana State University Medical Center, New Orleans 70112, USA. Studies on the human glucocorticoid receptor (GR) promoter were carried out so as to understand the regulation of GR expression. A -2738 to +19 fragment of the human GR promoter was used to identify important regulatory elements involved in the control in GR expression in NIH 3T3 cells (mouse fibroblasts) and HeLa cells (human cervical carcinoma cells). Important regulatory domains in the distal region of the human GR promoter were identified by sequential 5' end deletion analysis. A region between -2490 and -2025 contributed 50% of the measured transcriptional activity to the promoter. Using DNase I footprint analysis, four sites in this region were identified: -2362 to -2339 (mouse footprint, mFP); -2301 to -2293 (distal YY1, dYY1); -2130 to -2122 (middle YY1, mYY1); and, -2086 to -2078 (proximal YY1, pYY1). Three sites contained an identical core sequence, CCAAGATGG and were identified as Yin Yang 1 (YY1) binding sites. The site located at -2362 to -2339 was footprinted in NIH 3T3 cells only. The sequence of this site is a direct repeat with a 2-nucleotide spacer region, and it does not share homology with any known transcription factor binding sites. Computer analysis of the entire promoter sequence revealed an additional YY1 site located at -260 to -249 (initiator YY1, iYY1) with the sequence CTCCTCCATTTTG. Electrophoretic mobility supershift assays, with an anti-YY1 antibody, were used to confirm YY1 binding to these four putative YY1 binding sites. Site-directed deletion of all three upstream YY1 sites but not the iYY1 site, or the iYY1 site alone, showed a approximately 60% decrease in transcriptional activity of the hGR promoter in HeLa cells but had no effect in NIH 3T3 cells. A similar (50%) decrease in the expression of a full-length hGR/luciferase reporter gene was obtained when HeLa cells were cotransfected with a full-length antisense YY1 expression plasmid. Additionally, a region between -1841 and -1689 contributed to hGR promoter activity in both cell types tested. An Sp1 binding site was identified in this region (-1748 to -1733) by DNase I footprint and mobility supershift analyses. The presence of four YY1 binding sites in the human GR promoter suggests that these sites play a critical role in GR gene regulation. PMID: 10030686 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 44: J Biol Chem. 1999 Jan 29;274(5):3076-93. Characterization of the human endothelial nitric-oxide synthase promoter. Karantzoulis-Fegaras F, Antoniou H, Lai SL, Kulkarni G, D'Abreo C, Wong GK, Miller TL, Chan Y, Atkins J, Wang Y, Marsden PA. Renal Division and Department of Medicine, St. Michael's Hospital and University of Toronto, Toronto, Ontario M5S 1A8, Canada. Understanding transcription initiation of the endothelial nitric-oxide synthase (eNOS) gene appears pivotal to gaining a comprehensive view of NO biology in the blood vessel wall. The present study therefore focused upon a detailed dissection of the functionally important cis-DNA elements and the multiprotein complexes implicated in the cooperative control of constitutive expression of the human eNOS gene in vascular endothelium. Two tightly clustered cis-regulatory regions were identified in the proximal enhancer of the TATA-less eNOS promoter using deletion analysis and linker-scanning mutagenesis: positive regulatory domains I (-104/-95 relative to transcription initiation) and II (-144/-115). Analysis of trans-factor binding and functional expression studies revealed a surprising degree of cooperativity and complexity. The nucleoprotein complexes that form upon these regions in endothelial cells contained Ets family members, Sp1, variants of Sp3, MAZ, and YY1. Functional domain studies in Drosophila Schneider cells and endothelial cells revealed examples of positive and negative protein-protein cooperativity involving Sp1, variants of Sp3, Ets-1, Elf-1, and MAZ. Therefore, multiprotein complexes are formed on the activator recognition sites within this 50-base pair region of the human eNOS promoter in vascular endothelium. PMID: 9915847 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 45: Arch Biochem Biophys. 1999 Jan 1;361(1):7-16. Identification of two repressor elements in the mouse alpha 2(I) collagen promoter. Miao K, Potter JJ, Anania FA, Rennie-Tankersley L, Mezey E. Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, 21205-2195, USA. We recently identified three areas of Sp1 binding located between -568 and -453 of the 5' flanking region of the murine alpha2(I) collagen promoter which are necessary for optimal activity. We now identify two additional regions of Sp1 binding located at -371 to -351 (region 4) and at -690 to -613 (region 5), which when mutated increased promoter activity in transfected rat hepatic stellate cells indicating they contain negative regulatory elements. AP-2 bound to region 4 while YY1 bound most strongly to region 5. AP-2 decreased Sp1 binding to region 4 and had a dual effect on Sp1 binding to region 5 decreasing and increasing Sp1 binding at low and high concentrations of AP-2, respectively. YY1 enhanced Sp1 binding to both regions. AP-2 inhibited or enhanced the stimulatory effect of a transfected Sp1 expression vector on the alpha2(I) collagen promoter in Drosophila cells at low or high AP-2 expression, respectively. YY1 enhanced or inhibited the activation of the promoter by low or high Sp1 expression, respectively. This study identifies two negative regulatory elements in the murine alpha2(I) collagen promoter and shows that AP-2 and YY1 interact with Sp1 at these sites and can inhibit the activating action of Sp1. Copyright 1999 Academic Press. PMID: 9882423 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 46: Am J Respir Crit Care Med. 1998 Dec;158(6):1958-62. Interleukin-10 and transforming growth factor-beta promoter polymorphisms in allergies and asthma. Hobbs K, Negri J, Klinnert M, Rosenwasser LJ, Borish L. Departments of Medicine and Pediatrics, National Jewish Medical and Research Center, University of Colorado Health Sciences Center, Denver, Colorado, USA. Interleukin-10 (IL-10) and transforming growth factor beta (TGF-beta) are inhibitory for B and T cells, IgE production, and mast cell proliferation, and they induce apoptosis in eosinophils. These cytokines are therefore candidate genes which could contribute to the development of asthma or allergies. We investigated the hypothesis that polymorphic nucleotides within the IL-10 and TGF-beta gene promoters would link to the expression of allergies and asthma. DNA taken from families with an asthmatic proband was examined for base exchanges by single-stranded conformational polymorphism (SSCP). We demonstrated the presence of a polymorphism in the promoter region of the IL-10 gene and four in the TGF-beta gene promoters (3 in TGF-beta1 and 1 in TGF-beta2). The IL-10 gene polymorphism was a C-to-A exchange 571 base pairs upstream from the translation start site and was present between consensus binding sequences for Sp1 and elevated total serum. This polymorphism was associated with elevated total serum IgE in subjects heterozygotic or homozygotic for this base exchange (p < 0.009). The base exchange at -509 (from the transcription initiation site) in the TGF-beta promoter also linked to elevated total IgE (p < 0.01). This polymorphism represented a C-to-T base exchange which induced a YY1 consensus sequence and is present in a region of the promoter associated with negative transcription regulation. PMID: 9847292 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 47: Am J Respir Crit Care Med. 1998 Nov;158(5 Pt 3):S100-8. Transcriptional regulation of smooth muscle contractile apparatus expression. Solway J, Forsythe SM, Halayko AJ, Vieira JE, Hershenson MB, Camoretti-Mercado B. Section of Pulmonary and Critical Care Medicine, Department of Medicine, Section of Pulmonary Biology, Critical Care, Department of Pediatrics, University of Chicago, Chicago, Illinois, USA. The transcriptional regulatory mechanisms that control gene expression during differentiation and contractile protein accumulation are becoming well understood in skeletal and cardiac muscle lineages. Current understanding of smooth muscle-specific gene transcription is much more limited, though recent studies have begun to shed light on this topic. In this review, we summarize some of the themes emerging from these studies and identify transcriptional regulatory elements common to several smooth muscle genes. These include potential binding sites for serum response factor, Sp1, AP2, Mhox, and YY1, as well as a potential transforming growth factor-beta control element. We speculate that it may be possible to manipulate smooth muscle-specific gene expression in asthma or pulmonary arterial hypertension as an eventual therapy. Publication Types: Review Review, Tutorial PMID: 9817732 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 48: Prog Nucleic Acid Res Mol Biol. 1998;61:309-44. Structural organization and transcription regulation of nuclear genes encoding the mammalian cytochrome c oxidase complex. Lenka N, Vijayasarathy C, Mullick J, Avadhani NG. Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia 19104, USA. Cytochrome c Oxidase (COX) is the terminal component of the bacterial as well as the mitochondrial respiratory chain complex that catalyzes the conversion of redox energy to ATP. In eukaryotes, the oligomeric enzyme is bound to mitochondrial innermembrane with subunits ranging from 7 to 13. Thus, its biosynthesis involves a coordinate interplay between nuclear and mitochondrial genomes. The largest subunits, I, II, and III, which represent the catalytic core of the enzyme, are encoded by the mitochondrial DNA and are synthesized within the mitochondria. The rest of the smaller subunits implicated in the regulatory function are encoded on the nuclear DNA and imported into mitochondria following their synthesis in the cytosol. Some of the nuclear coded subunits are expressed in tissue and developmental specific isologs. The ubiquitous subunits IV, Va, Vb, VIb, VIc, VIIb, VIIc, and VIII (L) are detected in all the tissues, although the mRNA levels for the individual subunits vary in different tissues. The tissue specific isologs VIa (H), VIIa (H), and VIII (H) are exclusive to heart and skeletal muscle. cDNA sequence analysis of nuclear coded subunits reveals 60 to 90% conservation among species both at the amino acid and nucleotide level, with the exception of subunit VIII, which exhibits 40 to 80% interspecies homology. Functional genes for COX subunits IV, Vb, VIa 'L' & 'H', VIIa 'L' & 'H', VIIc and VIII (H) from different mammalian species and their 5' flanking putative promoter regions have been sequenced and extensively characterized. The size of the genes range from 2 to 10 kb in length. Although the number of introns and exons are identical between different species for a given gene, the size varies across the species. A majority of COX genes investigated, with the exception of muscle-specific COXVIII(H) gene, lack the canonical 'TATAA' sequence and contain GC-rich sequences at the immediate upstream region of transcription start site(s). In this respect, the promoter structure of COX genes resemble those of many house-keeping genes. The ubiquitous COX genes show extensive 5' heterogeneity with multiple transcription initiation sites that bind to both general as well as specialized transcription factors such as YY1 and GABP (NRF2/ets). The transcription activity of the promoter in most of the ubiquitous genes is regulated by factors binding to the 5' upstream Sp1, NRF1, GABP (NRF2), and YY1 sites. Additionally, the murine COXVb promoter contains a negative regulatory region that encompasses the binding motifs with partial or full consensus to YY1, GTG, CArG, and ets. Interestingly, the muscle-specific COX genes contain a number of striated muscle-specific regulatory motifs such as E box, CArG, and MEF2 at the proximal promoter regions. While the regulation of COXVIa (H) gene involves factors binding to both MEF2 and E box in a skeletal muscle-specific fashion, the COXVIII (H) gene is regulated by factors binding to two tandomly duplicated E boxes in both skeletal and cardiac myocytes. The cardiac-specific factor has been suggested to be a novel bHLH protein. Mammalian COX genes provide a valuable system to study mechanisms of coordinated regulation of nuclear and mitochondrial genes. The presence of conserved sequence motifs common to several of the nuclear genes, which encode mitochondrial proteins, suggest a possible regulatory function by common physiological factors like heme/O2/carbon source. Thus, a well-orchestrated regulatory control and cross talks between the nuclear and mitochondrial genomes in response to changes in the mitochondrial metabolic conditions are key factors in the overall regulation of mitochondrial biogenesis. Publication Types: Review PMID: 9752724 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 49: Nucleic Acids Res. 1998 Aug 15;26(16):3776-83. Cloning, chromosomal localization and promoter analysis of the human transcription factor YY1. Yao YL, Dupont BR, Ghosh S, Fang Y, Leach RJ, Seto E. H. Lee Moffitt Cancer Center and Research Institute, Department of Microbiology and Immunology, College of Medicine, University of South Florida, 12902 Magnolia Drive, Tampa, FL 33612, USA. Yin Yang 1 (YY1) is a protein that activates and represses transcription of a large number of cellular and viral genes. In addition, studies suggest that YY1 may play an important role in development and differentiation. Here, we report the isolation and analysis of a YY1 genomic clone from a lambda human liver library. Fluorescence in situ hybridization with the YY1 clone has localized the YY1 gene to chromosome 14 band q32. A major YY1 gene transcription initiation site has been mapped to 478 bp upstream of the ATG translation start site. The proximal promoter contains multiple Sp1 transcription factor binding sites but lacks a consensus TATA or CCAAT box. Transient transfections and detailed deletion analyses localized the promoter to no more than 277 bp upstream from the major transcription start site. Finally, we have found that overexpression of the adenovirus E1A protein represses expression of a reporter gene directed by the YY1 promoter. PMID: 9685495 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 50: J Gen Virol. 1998 Jul;79 ( Pt 7):1659-63. Point mutations in SP1 motifs in the upstream regulatory region of human papillomavirus type 18 isolates from cervical cancers increase promoter activity. Rose B, Steger G, Dong XP, Thompson C, Cossart Y, Tattersall M, Pfister H. Department of Infectious Diseases, Blackburn Building (DO6), The University of Sydney, NSW, Australia. b_rose@infdis.usyd.edu.au Evidence of the functional significance of two naturally occurring mutations at nt 40 or 41 in the Sp1 motif in the promoter proximal segment of the upstream regulatory region (URR) of human papillomavirus (HPV) type 18 is presented. In electrophoretic mobility shift assays, Sp1 protein bound more efficiently to the Sp1 mutant motifs than to the prototype; while in both HeLa and HT3 cells, luciferase activity controlled by the mutant URRs was upregulated 2- and 3-fold, or 4- and 6-fold, in comparison with the prototype URR or HeLa cell-derived URR respectively. The HeLa URR represents a more appropriate baseline for promoter activity, containing a series of point mutations representative of most HPV-18 cancer isolates, including one in the Yin Yang 1 (YY1) site at the P105 promoter. The effect of the Sp1 mutations was found to be largely maintained in the context of the HeLa URR containing the prototype YY1 site. PMID: 9680128 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 51: J Biol Chem. 1998 Apr 3;273(14):8287-93. Regulation of human B19 parvovirus promoter expression by hGABP (E4TF1) transcription factor. Vassias I, Hazan U, Michel Y, Sawa C, Handa H, Gouya L, Morinet F. Service de Microbiologie, Hopital Saint-Louis, 1 avenue Claude Vellefaux, 75475 Paris CEDEX 10, France. The genetic expression of human B19 parvovirus is only dependent on one promoter in vivo and in vitro. This is the P6 promoter, which is located on the left side of the genome and is a single-stranded DNA molecule. This led us to investigate the regulation of the P6 promoter and the possible resulting variability of the nucleotide sequence. After analysis of the promoter region of 17 B19 strains, only 1.5% variability was found. More exciting was the finding of mutations that were clustered around the TATA box and defined a highly conserved region (nucleotides 113-210) in the proximal part of the P6 promoter. HeLa and UT7/Epo cell extracts were found to protect this region, which contained a core motif for Ets family proteins, with YY1 and Sp1 binding sites on either side. Gel mobility shift assays performed with nuclear proteins from HeLa and UT7/Epo cells identified DNA-binding proteins specific for these sites. By supershift analysis, we demonstrated the binding of the hGABP (also named E4TF1) protein to the Ets binding site and the fixation of Sp1 and YY1 proteins on their respective motifs. In Drosophila SL2 cells, hGABPalpha and -beta stimulated P6 promoter activity, and hGABPalpha/hGABPbeta and Sp1 exerted synergistic stimulation of this activity, an effect diminished by YY1. PMID: 9525935 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 52: Genomics. 1998 Feb 15;48(1):111-6. Isolation and functional characterization of the mouse p75 TNF receptor promoter. Seitz C, Mannel DN, Hehlgans T. Institute of Pathology/Tumorimmunology, University of Regensburg, Germany. Tumor necrosis factor (TNF) is a pleiotropic cytokine that plays an important role in immunological and inflammatory responses. It exerts its biological effects via two distinct membrane receptors of apparent molecular weight of 55 (p55TNFR) and 75 kDa (p75TNFR), respectively. Most cell lines and primary tissues express both receptor types. While the p55TNFR gene is constitutively expressed at rather low levels, the transcription of p75TNFR is strongly modulated by a number of stimulatory agents. To characterize the mouse p75TNFR gene expression on a molecular level, we screened a mouse genomic library using the 5' end of the p75TNFR cDNA as a probe. A 6.3kb genomic clone containing about 6 kb of 5' flanking region and 300 bp of 3' sequence including the translational start site and the first exon was isolated and subcloned. Primer extension analysis revealed three transcriptional start sites located at -35, -39, and -564 bp upstream of the ATG-containing first exon. To determine whether the 5' flanking region exerts functional promoter activity, we generated deletion mutants fused to the luciferase reporter gene. Transfection of mouse fibroblasts (NIH3T3) with these constructs showed functional promoter activity of the isolated 5' region. By further sequence analysis of the 5' flanking region a number of putative DNA-binding sites for transcription factors, e.g., Sp1, CREB, Yi, YY1, and IFN gamma-responsive element, were identified. PMID: 9503023 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 53: Biochem Biophys Res Commun. 1998 Feb 4;243(1):233-40. Identification of 5' flanking sequence of RH50 gene and the core region for erythroid-specific expression. Iwamoto S, Omi T, Yamasaki M, Okuda H, Kawano M, Kajii E. Department of Legal Medicine and Human Genetics, Jichi Medical School, Tochigi, Japan. siwamoto@ms.jichi.ac.jp The Rh blood group antigens are carried by two distinct but homologous membrane proteins encoded by two closely related genes, RCHE and RHD. Rh50 glyco-protein is the membrane protein tightly associated with Rh polypeptides and is critical for expression of Rh antigens. The amino acid sequence and predicted membrane topology of Rh50 glycoprotein are significantly homologous with those of the Rh proteins. Northern blot analysis of leukemic cell lines showed that expression of RH50 gene is restricted to cells with erythroid features. HEL and K562 cells showed a transcription levels ratio of 1 to 9.9 for Rh50, and 12.3 to 1 for Rh. The nucleotide sequence of 5' flanking region of RH50 gene and functional promoter assays also supported the erythroid-specific regulation of the gene, whereas the sequence had lower homology with the promoter sequence of RH genes. Seven GATAs, nine E-boxes, two CACCCs, one YY1, and one October motif were identified in the 1868bp 5' flanking sequence. The core promoter of RH50 gene was located within 68bp length from the translation start position, which included an inverse GATA motif, although obvious motifs for Sp1 or erythroid Kruppel-like factor were lacking. The inverse GATA motif was the target sequence of GATA-1 protein, and disruption of the motif abolished the transactivating activity of erythroid cells. These studies confirm the erythroid-specific expression of Rh antigens, but suggest distinct regulatory mechanisms for RH vs RH50 genes. PMID: 9473510 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 54: Biochem Mol Med. 1997 Dec;62(2):159-64. Analysis of the 5' flanking region of the human galactocerebrosidase (GALC) gene. Luzi P, Victoria T, Rafi MA, Wenger DA. Department of Medicine (Medical Genetics), Jefferson Medical College, Philadelphia, Pennsylvania 19107, USA. Galactocerebrosidase (GALC) is the lysosomal enzyme deficient in human and certain animal species with globoid cell leukodystrophy (GLD) or Krabbe disease. It catalyzes the hydrolysis of specific galactolipids including galactosylceramide and psychosine. The GALC protein is found in very low amounts in all tissues, which delayed its purification and the subsequent cloning of its cDNA and gene. We previously published the exon-intron organization of the human gene, but did not functionally analyze the 5' flanking region. We now provide a description of this GC-rich region which includes one potential YY1 element and one potential SP1 binding site. There are 13 GGC trinucleotides within the first 150 bp preceding the initiation codon. The 5' end of intron 1 contains six potential Sp1 binding sites, one AP1 binding site, and eight AP2 binding sites. A construct containing nucleotides -176 to -24 had the strongest promoter activity using a vector containing the chloramphenicol acetyltransferase reporter gene. We also provide evidence for the presence of inhibitory sequences located immediately upstream of the promoter region, and within the first 234 nucleotides of intron 1. These elements together with a suboptimal nucleotide at position +4 may explain the low level of GALC protein in all cell types. PMID: 9441867 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 55: Nucleic Acids Res. 1997 Sep 15;25(18):3705-11. A functional YY1 binding site is necessary and sufficient to activate Surf-1 promoter activity in response to serum growth factors. Cole EG, Gaston K. Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK. The human Surf-1 and Surf-2 housekeeping genes are divergently transcribed and share a bi-directional, TATA-less promoter. Housekeeping promoters typically contain complex arrays of transcription factor binding sites and several studies have suggested that many of these sites might be functionally redundant. The Surf-1/Surf-2 promoter region contains four factor binding sites; members of the ETS family of transcription factors bind to two of these sites whilst YY1 binds to a third site immediately downstream of the major Surf-1 transcription start point. Here we show that Sp1 binds to the fourth transcription factor binding site. Although YY1 and Sp1 have previously been shown to interact both in vitro and in vivo, these proteins function independently at the Surf-1/Surf-2 promoter. The binding of Sp1 alone is sufficient to bring about full promoter activity in the Surf-2 direction. In contrast, both Sp1 and ETS proteins are required to bring about full promoter activity in the Surf-1 direction. The YY1 binding site is not required for basal transcription in either direction. The YY1 binding site is, however, both necessary and sufficient to confer growth factor inducibility on transcription in the Surf-1 direction. Our data suggest that functionally redundant transcription factor binding sites might not be a general feature of housekeeping promoters. PMID: 9278494 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 56: J Cell Biochem. 1997 Jul 1;66(1):123-32. Subcellular partitioning of transcription factors during osteoblast differentiation: developmental association of the AML/CBF alpha/PEBP2 alpha-related transcription factor-NMP-2 with the nuclear matrix. Lindenmuth DM, van Wijnen AJ, Hiebert S, Stein JL, Lian JB, Stein GS. Department of Cell Biology, University of Massachusetts Medical School and Cancer Center, Worcester, 01655, USA. The subnuclear location of transcription factors may functionally contribute to the regulation of gene expression. Several classes of gene regulators associate with the nuclear matrix in a cell type, cell growth, or cell cycle related-manner. To understand control of nuclear matrix-transcription factor interactions during tissue development, we systematically analyzed the subnuclear partitioning of a panel of transcription factors (including NMP-1/YY-1, NMP-2/AML, AP-1, and SP-1) during osteoblast differentiation using biochemical fractionation and gel shift analyses. We show that nuclear matrix association of the tissue-specific AML transcription factor NMP-2, but not the ubiquitous transcription factor YY1, is developmentally upregulated during osteoblast differentiation. Moreover, we show that there are multiple AML isoforms in mature osteoblasts, consistent with the multiplicity of AML factors that are derived from different genes and alternatively spliced cDNAs. These AML isoforms include proteins derived from the AML-3 gene and partition between distinct subcellular compartments. We conclude that the selective partitioning of the YY1 and AML transcription factors with the nuclear matrix involves a discriminatory mechanism that targets different classes and specific isoforms of gene regulatory factors to the nuclear matrix at distinct developmental stages. Our results are consistent with a role for the nuclear matrix in regulating the expression of bone-tissue specific genes during development of the mature osteocytic phenotype. PMID: 9215534 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 57: Mol Cell Biol. 1997 Jul;17(7):3723-32. Multiple mechanisms of transcriptional repression by YY1. Galvin KM, Shi Y. Department of Pathology, Harvard Cancer Center, Harvard Medical School, Boston, Massachusetts 02115, USA. The four C-terminal GLI-Kruppel type zinc fingers of YY1 have been identified as a transcriptional repression domain. Previous reports have proposed DNA-bending and activator-quenching mechanisms for this zinc finger-mediated repression. In addition, previous work indicated that p300 and CBP might be involved in YY1-mediated repression. We have analyzed these possible models for the zinc finger-mediated repression. The role of each zinc finger in the repression and DNA-binding functions was determined by using a structure-and-function approach. We show that zinc finger 2 of YY1 plays a central role in both DNA binding and transcriptional repression. However, a survey of a panel of YY1 mutants indicates that these two functions can be separated, which argues against the DNA-bending model for repression. We show that the physical interaction between YY1 and p300, a coactivator for CREB, is not sufficient for repression of CREB-mediated transcription. Our studies indicate that YY1 functions as an activator-specific repressor. Repression of CTF-1-directed transcription may be accomplished through direct physical interaction between YY1 and this activator. In contrast, physical interaction is not necessary for YY1 to repress Sp1- and CREB-mediated transcription. Rather, the repression likely reflects an ability of YY1 to interfere with communication between these activators and their targets within the general transcription machinery. Taken together, our results suggest that YY1 employs multiple mechanisms to achieve activator-specific repression. PMID: 9199306 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 58: Nucleic Acids Res. 1997 Jul 1;25(13):2661-71. Identification and characterization of the human XIST gene promoter: implications for models of X chromosome inactivation. Hendrich BD, Plenge RM, Willard HF. Department of Genetics and Center for Human Genetics, Case Western Reserve University School of Medicine, Cleveland, OH 44106-4955, USA. The XIST gene in both humans and mice is expressed exclusively from the inactive X chromosome and is required for X chromosome inactivation to occur early in development. In order to understand transcriptional regulation of the XIST gene, we have identified and characterized the human XIST promoter and two repeated DNA elements that modulate promoter activity. As determined by reporter gene constructs, the XIST minimal promoter is constitutively active at high levels in human male and female cell lines and in transgenic mice. We demonstrate that this promoter activity is dependent in vitro upon binding of the common transcription factors SP1, YY1 and TBP. We further identify two cis -acting repeated DNA sequences that influence reporter gene activity. First, DNA fragments containing a set of highly conserved repeats located within the 5'-end of XIST stimulate reporter activity 3-fold in transiently transfected cell lines. Second, a 450 bp alternating purine-pyrimidine repeat located 25 kb upstream of the XIST promoter partially suppresses promoter activity by approximately 70% in transient transfection assays. These results indicate that the XIST promoter is constitutively active and that critical steps in the X inactivation process must involve silencing of XIST on the active X chromosome by factors that interact with and/or recognize sequences located outside the minimal promoter. PMID: 9185579 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 59: Mol Cell Biol. 1997 Jun;17(6):2973-84. Accurate positioning of RNA polymerase II on a natural TATA-less promoter is independent of TATA-binding-protein-associated factors and initiator-binding proteins. Weis L, Reinberg D. Department of Biochemistry, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway 08854-5635, USA. Two promoter elements, the TATA element and initiator (Inr), are capable of directing specific transcription initiation of protein-encoding genes by RNA polymerase II (RNAPII). Although binding to the TATA element by the TATA-binding protein (TBP) has been shown to be the initial recognition step in transcription complex formation in vitro, the mechanism through which the basal machinery assembles into a functional complex on TATA-less promoters is controversial. Evidence supporting numerous models of Inr-mediated transcription complex formation exists, including the nucleation of a complex by Inr-binding proteins, a component of the TFIID complex, or a specific upstream activator common to many TATA-less promoters, Sp1. Using various techniques, we have undertaken a systematic analysis of the natural TATA-less human DNA polymerase beta (beta-pol) gene promoter. Although the beta-pol promoter contains upstream Sp1 elements and a functional Inr that binds YY1, neither of these factors is essential for Inr-mediated transcription complex formation. A complex containing TBP, TFIIB, TFIIF, and RNAPII (DBPolF complex) is capable of forming on the promoter in an Inr-dependent manner. A single point mutation within the Inr that affects DBPolF complex formation diminishes beta-pol transcriptional activity. PMID: 9154795 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 60: J Immunol. 1996 Dec 15;157(12):5411-21. Cloning of the human platelet endothelial cell adhesion molecule-1 promoter and its tissue-specific expression. Structural and functional characterization. Almendro N, Bellon T, Rius C, Lastres P, Langa C, Corbi A, Bernabeu C. Department of Immunology, Center of Biological Investigations, High Council of Scientific Investigations, Madrid, Spain. Platelet endothelial cell adhesion molecule-1 (PECAM-1; CD31) is a cell adhesion molecule involved in transendothelial migration and expressed by hemopoietic and endothelial cells. To understand the mechanisms underlying its regulated expression, a genomic clone containing 1555 bp of the 5'-flanking region and the first exon of the human PECAM-1 gene has been isolated. The 5'-flanking region of the PECAM-1 gene lacks a consensus TATA box, but contains consensus motifs for Sp1, EGR1, ets, helix-loop-helix (HLH) box, GATA, AP-2, C/EBP, YY1, CCACC, LyF-1, imperfect octamer, heptamer, high mobility group proteins (HMG) box, and nuclear factor-kappaB, as well as shear stress-, retinoic acid-, glucocorticoid-, and acute phase-responsive elements, and an Alu sequence. Successive 5' to 3' or 3' to 5' deletions revealed tissue-specific promoter activity within the two contiguous 0.22-kb NheI/BglII and 0.44-kb BglII/PstI fragments. The transcriptional activity displayed by the 0.22-kb NheI/BglII fragment was specific for the myeloid lineage, whereas the promoter activity of the 0.44-kb BglII/PstI fragment was apparently restricted to endothelial cells. The transcriptional activity of the 0.22-kb NheI/BglII fragment was confirmed by 5' RACE (rapid amplification of 5' cDNA ends) and S1 nuclease protection experiments that revealed previously unidentified transcription start sites. The 0.22-kb NheI/BglII promoter exhibited PMA inducibility in myeloid cells and contained a PMA-responsive element recognized by Sp1 and EGR-1 transcription factors. Isolation and characterization of the human PECAM-1 promoter represent an initial step in elucidating the controlled expression of the PECAM-1 gene. PMID: 8955189 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 61: Genomics. 1996 May 15;34(1):85-91. The complete sequence of the host cell factor 1 (HCFC1) gene and its promoter: a role for YY1 transcription factor in the regulation of its expression. Zoppe M, Frattini A, Faranda S, Vezzoni P. Istituto di Tecnologie Biomediche Avanzate (ITBA), Consiglio Nazionale delle Ricerche, Milan, 20131, Italy. We report here the complete sequence of the Host Cell Factor (HCFC1) gene, including two kilobases of the 5'-flanking region and 5.9 kb of the first intron. The upstream and 5'-untranslated regions contain several putative transcriptional factor binding sites and a 17-nt-long repeated element (SiSa element) present in six regularly spaced copies, of which five are perfectly identical, while the sixth has a transition substitution (CT for TC) at nucleotides 13 and 14. Four copies are contained in the flanking region, the fifth is at the beginning of the mRNA (position +9), and the sixth is at position 195 of the mRNA. This 17-bp element contains at its 5' side an octamer sequence known to bind the Yin/Yang 1 (YY1) transcription factor; another YY1 binding octamer is present at the end of the first intron. The promoter also contains several Sp1 binding sites, some of which are located very close to SiSa elements. We demonstrate that YY1 binds to the 5' half of the SiSa element, whose 3' region binds in gel shift experiments an additional, as yet unidentified nuclear factor. Therefore the YY1 binding site in HCFC1 overlaps the site of a second factor, as has been described in several YY1-site-containing promoters. This suggests that HCFC1 expression might be regulated by the reciprocal interaction of several transcription factors. PMID: 8661027 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 62: J Biol Chem. 1996 May 3;271(18):10827-33. Serum response factor mediates AP-1-dependent induction of the skeletal alpha-actin promoter in ventricular myocytes. Paradis P, MacLellan WR, Belaguli NS, Schwartz RJ, Schneider MD. Department of Medicine, Baylor College of Medicine, Houston, Texas 77030, USA. "Fetal" gene transcription, including activation of the skeletal alpha-actin (SkA) promoter, is provoked in cardiac myocytes by mechanical stress and trophic ligands. Induction of the promoter by transforming growth factor beta or norepinephrine requires serum response factor (SRF) and TEF-1; expression is inhibited by YY1. We and others postulated that immediate-early transcription factors might couple trophic signals to this fetal program. However, multiple Fos/Jun proteins exist, and the exact relationship between control by Fos/Jun versus SRF, TEF-1, and YY1 is unexplained. We therefore cotransfected ventricular myocytes with Fos, Jun, or JunB, and SkA reporter genes. SkA transcription was augmented by Jun, Fos/Jun, Fos/JunB, and Jun/JunB; Fos and JunB alone were neutral or inhibitory. Mutation of the SRF site, SRE1, impaired activation by Jun; YY1, TEF-1, and Sp1 sites were dispensable. SRE1 conferred Jun activation to a heterologous promoter, as did the c-fos SRE. Deletions of DNA binding, dimerization, or trans-activation domains of Jun and SRF abolished activation by Jun and synergy with SRF. Neither direct binding of Fos/Jun to SREs, nor physical interaction between Fos/Jun and SRF, was detected in mobility-shift assays. Thus, AP-1 factors activate a hypertrophy-associated gene via SRF, without detectable binding to the promoter or to SRF. PMID: 8631897 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 63: Gene Expr. 1996;6(3):151-68. Specific binding sites for a pol III transcriptional repressor and pol II transcription factor YY1 within the internucleosomal spacer region in primate Alu repetitive elements. Humphrey GW, Englander EW, Howard BH. Laboratory of Molecular Growth Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA. Alu interspersed repetitive elements possess internal RNA polymerase III promoters that are transcribed in vitro and in transfected mouse cells but are nearly silent in human HeLa cells. Transcriptional repression of these elements is to some extent reversible, as pol III-dependent Alu expression can be induced with herpes simplex or adenovirus. To assess whether sequence-specific DNA binding proteins might contribute to Alu transcriptional silencing, we examined the internucleosomal spacer region surrounding the B box of the Alu pol III promoter in HeLa cell nuclei for evidence of proteins bound at specific sites in vivo. We identified a DNase I-hypersensitive site 5' to the B box and a DNase I-resistant region 3' to the B box in nuclei. An Alu-specific repressor binds to a 5-bp inverted repeat motif overlapping the 5' end of the TFIIIC binding site and may inhibit pol III transcription through competitive displacement. The level of Alu-specific pol III repressor activity is significantly reduced in adenovirus-infected HeLa cells, suggesting that the repressor may contribute to Alu transcriptional silencing in vivo. The 3' DNase I-resistant region coincided with a binding site for the pol II transcription factor YY1 in vitro. YY1 is one of the major proteins in HeLa cells having binding specificity for Alu elements. YY1 bound to tandem arrays of genomic Alu elements may play a role in chromatin organization and silencing. PMID: 9041122 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 64: J Gerontol A Biol Sci Med Sci. 1996 Jan;51(1):B66-75. YY1 and Sp1 transcription factors bind the human transferrin gene in an age-related manner. Adrian GS, Seto E, Fischbach KS, Rivera EV, Adrian EK, Herbert DC, Walter CA, Weaker FJ, Bowman BH. Department of Cellular and Structural Biology, University of Texas Health Science Center, San Antonio, USA. The iron-binding protein transferrin has major roles in transporting, delivering, and sequestering ferric ions acquired by body tissues. Yet, during aging, serum transferrin levels decrease in humans. Likewise, in transgenic mice carrying chimeric human transferrin transgenes, liver expression of transferrin transgenes decreases with age. The aging regulation is due to decreased gene transcription. Electrophoretic mobility shift assays and antibody-recognition have revealed the binding of 5' regulatory elements of the human transferrin gene by three YY1 proteins, called YY1, YY1-a, and YY1-b, and an Sp1-a transcription factor. An age-related increase in YY1-a and YY1-b binding activities and a decrease in Sp1-like binding activity were shown. Since Sp1 is a positive transcription factor and YY1 can be a negative transcription factor, the alterations in their binding with age could cause the decreased transcription of the human transferrin transgene, and also the age-related decreased serum transferrin levels in humans. PMID: 8548503 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 65: Mol Cell Biol. 1996 Jan;16(1):157-67. Characterization of the human granulocyte-macrophage colony-stimulating factor gene promoter: an AP1 complex and an Sp1-related complex transactivate the promoter activity that is suppressed by a YY1 complex. Ye J, Zhang X, Dong Z. Laboratory of Experimental Immunology, DCT, National Cancer Institute-Frederick Cancer Research and Development Center, Maryland 21702, USA. It is well documented that a repeated CATT element in the human granulocyte-macrophage colony-stimulating factor (GM-CSF) gene promoter is required for promoter activity. However, the transcription factors that are able to transactivate this enhancer element remain unidentified. Recently, we have found that nuclear factor YY1 can interact with the enhancer element. Here, we report that in addition to YY1, two other nuclear factors have been identified in the DNA-protein complexes formed by the CATT oligonucleotide and the Jurkat T-cell nuclear protein. One of these factors is AP1, and the other one is an Sp1-related protein. Results from transient transfection of Jurkat T cells have revealed that formation of both AP1 and the Sp1-related complex is required for the full enhancer activity of the CATT element. This result is supported by cotransfection of a c-jun expression vector and mutational analysis of the AP1 site or the Sp1-related protein binding site. In contrast, formation of the YY1 complex suppresses enhancer activity, since deletion of the YY1 complex induces an augmentation of the enhancer activity and overexpression of YY1 results in an attenuation of the enhancer activity. Results from the mechanism study have revealed that YY1 is able to inhibit transactivation mediated by either AP1 or the Sp1-related protein, and YY1 suppressive activity is DNA binding dependent. Taken together, these data support the ideas that AP1 and the Sp1-related nuclear protein are required for transactivation of the human GM-CSF gene promoter and that YY1 can suppress transactivation of the promoter even under inducible conditions. PMID: 8524292 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 66: Nucleic Acids Res. 1995 Sep 11;23(17):3473-80. TBP binding and the rate of transcription initiation from the human beta-globin gene. Antoniou M, de Boer E, Spanopoulou E, Imam A, Grosveld F. Laboratory of Gene Structure and Expression, National Institute for Medical Research, London, UK. DNA-protein interaction studies in vitro revealed several factors binding over the TATA box and the region of transcription initiation (cap) site of the human beta-globin promoter; TATA binding protein TBP at -30, Sp1 at -19, GATA-1 at -12 and +5, YY1 at -9 and a novel factor C1 over the site of initiation (-4 to +7). Point mutants which specifically abolish the binding of each of these proteins were tested in a beta-globin locus control region (LCR) construct which allows quantitative comparisons at physiological levels of transcription. Only mutants which drastically affect the binding of TBP resulted in decreased levels of transcription. A threshold value of TBP binding of 15-30% of wild type was sufficient to give normal levels of transcription. This indicates that the association of TF IID with the TATA box is not limiting in the rate of initiation of transcription. PMID: 7567458 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 67: Science. 1995 Jan 27;267(5197):531-6. Cloning of an intrinsic human TFIID subunit that interacts with multiple transcriptional activators. Chiang CM, Roeder RG. Laboratory of Biochemistry and Molecular Biology, Rockefeller University, New York, NY 10021. TFIID is a multisubunit protein complex comprised of the TATA-binding protein (TBP) and multiple TBP-associated factors (TAFs). The TAFs in TFIID are essential for activator-dependent transcription. The cloning of a complementary DNA encoding a human TFIID TAF, TAFII55, that has no known homolog in Drosophila TFIID is now described. TAFII55 is shown to interact with the largest subunit (TAFII230) of human TFIID through its central region and with multiple activators--including Sp1, YY1, USF, CTF, adenoviral E1A, and human immunodeficiency virus-type 1 Tat proteins--through a distinct amino-terminal domain. The TAFII55-interacting region of Sp1 was localized to its DNA-binding domain, which is distinct from the glutamine-rich activation domains previously shown to interact with Drosophila TAFII110. Thus, this human TFIID TAF may be a co-activator that mediates a response to multiple activators through a distinct mechanism. PMID: 7824954 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 68: Mol Reprod Dev. 1994 Sep;39(1):112-7. Control of cardiac gene transcription by fibroblast growth factors. Schneider MD, Kirshenbaum LA, Brand T, MacLellan WR. Molecular Cardiology Unit, Baylor College of Medicine, Houston, Texas 77030. Skeletal alpha-actin (SkA) is representative of the cardiac genes that are expressed at high levels in embryonic myocardium, downregulated after birth, and reactivated by tropic signals including basic fibroblast growth factor (FGF-2) and type beta transforming growth factors (TGF beta). To investigate the molecular basis for cardiac-restricted and growth factor-induced SkA transcription, we have undertaken a mutational analysis of the SkA promoter in neonatal ventricular myocytes, with emphasis on the role of three nominal serum response elements. Serum response factor (SRF) and the bifunctional factor YY1 are the predominant cardiac proteins contacting the proximal SRE (SRE1). Mutations of SRE1 that prevent recognition by SRF and YY1. or SRF alone, virtually abolish SkA transcription; mutation of distal SREs was ineffective. A mutation which selectively abrogates YY1 binding increases expression, substantiating the predicted role of YY1 as an inhibitor of SRF effects. SkA transcription requires combinational action of SRE1 with consensus sites for Sp1 and the SV40 enhancer binding protein, TEF-1. As an isolated motif, SRE1 can confer responsiveness to both FGF-2 and TGF beta to a heterologous promoter. Whether TEF-1 binding sites likewise can function as FGF response elements is unknown. Molecular dissection of mechanisms that govern the differentiated cardiac phenotype has largely been undertaken to date in neonatal ventricular myocytes, as the adult ventricular myocyte has been refractory to conventional procedures for gene transfer. To circumvent expected limitations of other methods, we have used replication-deficient adenovirus to achieve efficient gene transfer to adult cardiac cells in culture.(ABSTRACT TRUNCATED AT 250 WORDS) Publication Types: Review Review, Tutorial PMID: 7528025 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 69: J Biol Chem. 1994 Jun 17;269(24):16754-60. Transforming growth factor-beta response elements of the skeletal alpha-actin gene. Combinatorial action of serum response factor, YY1, and the SV40 enhancer-binding protein, TEF-1. MacLellan WR, Lee TC, Schwartz RJ, Schneider MD. Department of Medicine, Baylor College of Medicine, Houston, Texas 77030. Skeletal alpha-actin (SkA) is representative of the cardiac genes that are expressed at high levels in embryonic myocardium, down-regulated after birth, and reactivated by trophic signals including type beta-transforming growth factors (TGF beta). To investigate the molecular basis for cardiac-restricted and TGF beta-induced SkA transcription, we have undertaken a mutational analysis of the SkA promoter in ventricular myocytes, with emphasis on the role of three nominal serum response elements. Serum response factor (SRF) and the bifunctional factor YY1 are the predominant cardiac proteins contacting the proximal SRE (SRE1). Mutations of SRE1 that prevent recognition by SRF and YY1, or SRF alone, virtually abolish SkA transcription in both TGF beta- and vehicle-treated cells; mutation of distal SREs was ineffective. A mutation which selectively abrogates YY1 binding increases both basal and TGF beta-dependent expression, substantiating the predicted role of YY1 as an inhibitor of SRF effects. However, efficient SkA transcription requires combinatorial action of SRE1 with consensus sites for Sp1 and the SV40 enhancer-binding protein, TEF-1. As isolated motifs, either SRE1- or TEF-1-binding sites function as TGF beta response elements. Induction of the SkA promoter by TGF beta required SRF and TEF-1 in concert, unlike other pathways for TGF beta-dependent gene expression. PMID: 8206998 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 70: Nature. 1993 Sep 30;365(6445):462-4. Interaction between transcription factors Sp1 and YY1. Seto E, Lewis B, Shenk T. Department of Cellular & Structural Biology, University of Texas Health Science Center, San Antonio 78245-3207. A basal level of transcription is usually observed when all but a small region of DNA has been deleted from a eukaryotic gene promoter. These promoter elements, which are necessary and sufficient for specific transcription initiation, are referred to as minimal or core promoter elements. One element that is commonly present in a core promoter is the initiator. It has been demonstrated that the presence of Sp1 binding sites can greatly enhance the level of transcription initiation at initiator elements. A binding site for the YY1 transcription factor, located at the initiation site of the adeno-associated virus P5 promoter, functions as an initiator element; a synergistic enhancement of its activity is observed in vitro when upstream Sp1 binding sites are present. Here we report that this synergistic activation probably occurs through protein-protein interactions. PMID: 8003102 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 71: Mol Cell Biol. 1993 Sep;13(9):5439-49. Nuclear protein-binding sites in a transcriptional control region of the rabbit alpha-globin gene. Yost SE, Shewchuk B, Hardison R. Department of Molecular and Cell Biology, Pennsylvania State University, University Park 16802. The 5'-flanking and internal regions of the rabbit alpha-globin gene, which constitute a CpG island, are required for enhancer-independent expression in transfected cells. In this study, electrophoretic mobility shift assays revealed that a battery of nuclear proteins from both erythroid and nonerythroid cells bind specifically to these regulatory regions. Assays based on exonuclease III digestion, methylation interference, and DNase I footprinting identified sequences bound by proteins in crude nuclear extracts and by purified transcription factor Sp1. In the 5' flank, recognition sites for the transcription factors alpha-IRP (positions -53 to -44 relative to the cap site), CP1 (-73 to -69), and Sp1 (-95 to -90) are bound by proteins in K562 cell nuclear extracts, as are three extended upstream regions. Two recognition sites for Sp1 in intron 1 are also bound both by proteins in crude nuclear extracts and by purified Sp1. The sequences CCAC in intron 2 and C5 in the 3'-untranslated region also bind proteins. A major binding site found in exon 1, TATGGCGC, matches in sequence and methylation interference pattern the binding site for nuclear protein YY1, and binding is inhibited through competition by YY1-specific oligonucleotides. The protein-binding sites flanking and internal to the rabbit alpha-globin gene may form an extended promoter. PMID: 8355692 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 72: Proc Natl Acad Sci U S A. 1993 Jul 1;90(13):6145-9. Evidence for physical interaction between the zinc-finger transcription factors YY1 and Sp1. Lee JS, Galvin KM, Shi Y. Committee on Virology, Harvard Medical School, Boston, MA 02115. Two promoter elements are important for basal-level transcription, the TATA motif typically located 30 nucleotides upstream of the transcription initiation site and the initiator (Inr) element encompassing the start site. The mechanism of how Inr elements work is poorly understood, partly because very few proteins that bind to Inr elements have been identified and isolated. The recently cloned YY1 is such an Inr-binding protein. YY1 is able to direct transcription upon binding to its recognition sequence in vitro. The ability of YY1 to initiate transcription is augmented by the presence of a TATA motif or binding sites for transcription factor Sp1. To study the mechanism underlying the apparent functional cooperation between YY1 and Sp1, we explored the possibility of protein-protein interactions between these two transcription factors. We found that YY1 and Sp1 can form a physical complex. In addition, we identified domains within YY1 and Sp1 that mediate their interactions with each other. The physical interaction between YY1 and Sp1 may thus form the basis for the functional interplay observed previously. PMID: 8327494 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 73: Proc Natl Acad Sci U S A. 1993 Jun 15;90(12):5559-63. Characterization of the mouse gene that encodes the delta/YY1/NF-E1/UCRBP transcription factor. Safrany G, Perry RP. Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, PA 19111. The mouse gene that encodes the delta transcription factor has been cloned and characterized. This gene spans 23 kb and is composed of five exons and four introns. The first exon consists of a long (431 bp), (C+G)-rich, untranslated segment and a 679-bp coding segment, which specifies the unusual tracts of consecutive acidic residues and histidines and the long alanine-glycine stretches. The sequence that encodes the four zinc-finger motifs of this protein is interrupted by two introns. Nuclease protection experiments revealed a major transcriptional start point and several additional start points distributed over a 28-bp segment. Transfection experiments with 5' and 3' deletion mutants localized the promoter to a (C+G)-rich region that is < 700 bp upstream and no more than 32 bp downstream of the major start point. An especially critical promoter element lies between -58 and -18 and contains a high-affinity Sp1 binding site, as demonstrated by electrophoretic mobility-shift experiments with nuclear extract and recombinant Sp1 proteins. Several striking similarities between the delta gene and genes encoding other transcription factors and regulatory proteins are noted and discussed with respect to their possible biological significance. PMID: 8516301 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 74: Crit Rev Eukaryot Gene Expr. 1993;3(4):229-54. Transcription from TATA-less promoters: dihydrofolate reductase as a model. Azizkhan JC, Jensen DE, Pierce AJ, Wade M. Roswell Park Cancer Institute, Buffalo, NY 24623. The promoters of many of the genes encoding the so-called "housekeeping" enzymes, oncogenes, growth factors and their receptors, and transcription factors, do not contain a canonical TATA box, which is known to direct transcription initiation. The mechanisms through which TATA-less promoters are regulated, and their transcription start sites selected, have begun to yield to investigation. Using the dihydrofolate reductase (DHFR) gene as a model, recent work on this group of genes has been reviewed. Control of transcription initiation and the role of "initiator" sequences, as well as their binding factors, in particular YY1 and E2F, are addressed. In the DHFR gene, neither the E2F site at the major initiation region nor the upstream Sp1 sites can alone produce wild-type initiation, despite the fact that each of these sites has certain properties of initiators. Many TATA-less genes are growth regulated, that is, transcription is increased in response to stimulation of cell proliferation. Although both Sp1 and E2F have been implicated in growth regulation, our recent studies suggest that Sp1 sites alone can confer a growth-dependent increase in transcription in the late G1 and early S phases of the cell cycle. The regulatory role of E2F, which binds to many TATA-less promoters and mediates viral stimulation of transcription, is also reviewed. Publication Types: Review PMID: 8286846 [PubMed - indexed for MEDLINE] ---------------------------------------------------------------