1: Cancer Cell. 2005 Aug;8(2):155-67. 

Comment in:
    Cancer Cell. 2005 Aug;8(2):91-3.

Neuronal apoptosis linked to EglN3 prolyl hydroxylase and familial
pheochromocytoma genes: developmental culling and cancer.

Lee S, Nakamura E, Yang H, Wei W, Linggi MS, Sajan MP, Farese RV, Freeman RS,
Carter BD, Kaelin WG Jr, Schlisio S.

Department of Medical Oncology, Dana-Farber Cancer Institute and Brigham and
Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.

Germline NF1, c-RET, SDH, and VHL mutations cause familial pheochromocytoma.
Pheochromocytomas derive from sympathetic neuronal precursor cells. Many of
these cells undergo c-Jun-dependent apoptosis during normal development as NGF
becomes limiting. NF1 encodes a GAP for the NGF receptor TrkA, and NF1 mutations
promote survival after NGF withdrawal. We found that pheochromocytoma-associated
c-RET and VHL mutations lead to increased JunB, which blunts neuronal apoptosis
after NGF withdrawal. We also found that the prolyl hydroxylase EglN3 acts
downstream of c-Jun and is specifically required among the three EglN family
members for apoptosis in this setting. Moreover, EglN3 proapoptotic activity
requires SDH activity because EglN3 is feedback inhibited by succinate. These
studies suggest that failure of developmental apoptosis plays a role in
pheochromocytoma pathogenesis.

PMID: 16098468 [PubMed - indexed for MEDLINE]


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2: Gene. 2004 Oct 27;341:209-18. 

E2F sites in the Op18 promoter are required for high level of expression in the
human prostate carcinoma cell line PC-3-M.

Polzin RG, Benlhabib H, Trepel J, Herrera JE.

Department of Biochemistry, Molecular Biology, and Biophysics, University of
Minnesota, 321 Church St. S.E., 6-155 Jackson Hall, Minneapolis, MN 55455, USA.

Op18 (Oncoprotein 18, Stathmin) is an important mitotic regulator that is highly
expressed in many cancers including the metastatic prostate carcinoma cell line
PC-3-M. Recent studies indicate that antisense-mediated down-regulation of Op18
can inhibit cellular proliferation. However, the transcriptional mechanisms
responsible for its normal regulation and for its high level of expression in
proliferating cells remain poorly understood. In the study presented here, we
have characterized transcription factor binding sites that together account for
nearly 80% of the Op18 expression in PC-3-M cells. The 5' flanking region of the
Op18 gene contains four putative E2F sites located at -700 (site 1), -28 (site
2), -19 (site 3), and +720 (site 4) relative to the transcriptional start site.
E2F has been implicated in both the c-Jun-mediated up-regulation and the
doxorubicin-induced repression of Op18 expression. We have used
promoter-reporter assays and mobility shift assays to functionally examine each
of these E2F sites. Mutagenesis studies indicate that all sites contribute to
the basal expression of Op18. Mutagenesis of either site 1 or 4 reduced the
reporter activity by 40%, mutagenesis of site 2 reduced reporter activity by
20%, and mutations in site 3 had no effect on reporter activity. Combinatorial
mutagenesis indicates that site 1 and 4 function independently, whereas site 2
functions synergistically with either site 3 or 4. Mobility shift assays
indicate that all elements bind factors in the nuclear extracts of PC-3-M cells.
Characterization of the sites show that site 1, though a positive element, is
not E2F specific; sites 2 and 3 may contain an overlapping binding site for E2F
and NF1; and site 4, which resides in intron 1, is the only site shown to
interact exclusively with E2F. These studies suggest that the overexpression of
Op18 in PC-3-M cells is mediated predominantly through the E2F family of
transcription factors.

PMID: 15474303 [PubMed - indexed for MEDLINE]


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3: J Virol. 2004 Jul;78(14):7590-601. 

Identification and characterization of cis-acting elements residing in the
walleye dermal sarcoma virus promoter.

Hronek BW, Meagher A, Rovnak J, Quackenbush SL.

Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045,
USA.

Walleye dermal sarcoma virus (WDSV) is a complex retrovirus found associated
with tumors that appear and regress on a seasonal basis. There are quantitative
and qualitative differences in the amount of virus expression between developing
and regressing tumors. To understand the role of host cell factors in WDSV
expression, DNase I footprint analysis, electrophoretic mobility shift assays
(EMSA), and reporter gene assays were employed. DNase I footprint analysis of
the U3 region of the WDSV long terminal repeat with nuclear extract prepared
from a walleye cell line revealed protection of an Oct1, AP1, Whn, and two E4BP4
sites. Additionally, three regions that contained no putative transcription
factor binding sites were protected. EMSA confirmed the specific binding of the
protected sites and revealed three additional sites, NF1, AP3, and LVa, not
protected in DNase I footprint analysis. Site-directed mutagenesis of the
individual sites, in the context of a luciferase reporter plasmid, revealed that
the NF1, Oct1, AP1, E4BP4#2, AP3, and LVa sites contributed to transcription
activation driven by the WDSV U3 region. Mutation of Novel#2 resulted in an
increase in luciferase activity, suggesting the Novel#2 site may function to
bind a negative regulator of transcription. Anti-Jun and anti-Fos antiserum
specifically inhibited protein-DNA complex formation, indicating the presence of
c-Jun and c-Fos in the walleye cell nuclear extracts and their participation in
binding to the AP1 site. Interestingly, degenerative 15-bp repeats found in the
U3 region are differentially protected in DNase I footprint analysis by the
walleye cell line nuclear extract and regressing-tumor nuclear extract. EMSA
utilizing the 15-bp repeat probe revealed that there are similarities of binding
with W12 cell and developing-tumor nuclear extracts and that the binding differs
from that observed with regressing-tumor nuclear extract.

PMID: 15220434 [PubMed - indexed for MEDLINE]


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4: J Biol Chem. 2002 Sep 20;277(38):35232-9. Epub 2002 Jul 1. 

Role of AP-1 in the coordinate induction of rat glutamate-cysteine ligase and
glutathione synthetase by tert-butylhydroquinone.

Yang H, Zeng Y, Lee TD, Yang Y, Ou X, Chen L, Haque M, Rippe R, Lu SC.

Division of Gastroenterology and Liver Diseases, University of Southern
California Liver Disease Research Center, Keck School of Medicine, University of
Southern California, Los Angeles, California 90033, USA.

GSH synthesis occurs via two enzymatic steps catalyzed by glutamate-cysteine
ligase (GCL, made up of two subunits) and GSH synthetase (GS). Recently, we
described coordinate induction of GCL subunits and GS. To study GS
transcriptional regulation, we have cloned and characterized a 2.2-kb
5'-flanking region of the rat GS (GenBank accession number AF333982). One
transcriptional start site is located at 51 nucleotides upstream of the
translational start site. The rat GS promoter drove efficiently luciferase
expression in H4IIE cells. Sequential deletion analysis revealed DNA regions
that are involved in positive and negative regulation. One repressor identified
was NF1. tert-Butylhydroquinone (TBH) exerted a dose- and time-dependent
increase in the mRNA level and promoter activity of both GCL subunits and GS.
TBH increased protein binding to several regions of the GS promoter, c-jun
expression, and activator protein 1 (AP-1) binding activity to several of the
putative AP-1-binding sites of the GS promoter. Blocking AP-1 binding with
dominant-negative c-jun led to decreased basal expression and significantly
blocked the TBH-induced increase in promoter activity and mRNA level of all
three genes. In conclusion, AP-1 is required for basal expression of GCL and GS;
while NF1 serves as a repressor of GS, increased AP-1 transactivation is the
predominant mechanism for coordinate induction of GCL and GS expression by TBH.

PMID: 12093805 [PubMed - indexed for MEDLINE]


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5: Eur J Biochem. 1998 Sep 15;256(3):541-9. 

Phorbol ester causes ligand-independent activation of the androgen receptor.

Darne C, Veyssiere G, Jean C.

Centre National de la Recherche Scientifique Unite Mixte de Recherche 6547,
Universite Blaise Pascal-Clermont-Ferrand II, Aubiere, France.

We have examined the human androgen receptor (hAR) for its ability to activate
AR-dependent transcription of a transgene in a ligand-independent manner. The
transcriptional activity was determined by analysis of chloramphenicol
acetyltransferase (CAT) activity in T47D cells cotransfected with a plasmid
expressing the hAR and a natural AR-regulated promoter (the MVDP
androgen-dependent enhancer) ligated to the reporter CAT gene. In this study,
the effects of the protein kinase C (PKC) activator 12-O-tetradecanoyphorbol-13
acetate (TPA) on AR activity were tested. We demonstrated that in the absence of
androgen, TPA enhanced AR-mediated transactivation by 10-12-fold. This effect
was specific of the PKC pathway since stimulation to the PKA pathway did not
activate the unliganded AR. This ligand-independent pathway can function through
another androgen-regulated promoter as shown by the use of the mouse mammary
tumor virus MMTV-CAT reporter. The human glucocorticoid receptor (hGR) and the
rabbit progesterone receptor (rPR) could not be activated by TPA, indicating
that the effects are not universal for steroid receptors. A reporter plasmid
containing the MVDP androgen response element (ARE) in front of the thymidine
kinase promoter ligated to the CAT gene was activated by DHT but not by TPA,
indicating that the context of the natural promoter is critical for
ligand-independent activation of the AR. Exogenous c-jun enhanced
transcriptional activation by the AR in a ligand-dependent manner, but had no
effect in the absence of DHT. Base pair substitutions in both AR-binding
(5'-TGTTCT-3' to 5'-TTTTTT-3') and NF1-binding (5'-GTGGCTG-3' to 5'-GTTTTTG-3')
sites resulted in a loss of TPA responsiveness. Our results suggest that
ligand-independent activation of the AR by TPA results from interaction of
unliganded AR with other proteins in the transcription machinery.

PMID: 9780230 [PubMed - indexed for MEDLINE]


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6: Proc Natl Acad Sci U S A. 1998 Apr 28;95(9):5139-44. 

A role for Pak protein kinases in Schwann cell transformation.

Tang Y, Marwaha S, Rutkowski JL, Tennekoon GI, Phillips PC, Field J.

Department of Pharmacology, University of Pennsylvania School of Medicine,
Philadelphia, PA 19104, USA.

Neurofibromatosis type 1 (NF1), a common autosomal dominant disorder caused by
loss of the NF1 gene, is characterized clinically by neurofibromas and more
rarely by neurofibrosarcomas. Neurofibromin, the protein encoded by NF1,
possesses an intrinsic GTPase accelerating activity for the Ras proto-oncogene.
Through this activity, it is a negative regulator of Ras. The Pak protein kinase
is a candidate for a downstream signaling protein that may mediate Ras signals
because it is activated by Rac and Cdc42, two small G proteins required for Ras
signaling. Here, we use Pak mutants to explore the role of Pak in Ras signaling
in Schwann cells, the cells affected in NF1. Whereas an activated Pak mutant
does not transform cells, dominant negative Pak mutants are potent inhibitors of
Ras transformation of rat Schwann cells and of a neurofibrosarcoma cell line
from an NF1 patient. Although activated Pak stimulated jun-N-terminal kinase,
inhibition of Ras transformation by dominant negative Pak did not require
inhibition of jun-N-terminal kinase. Instead, the Pak mutants appeared to
inhibit transformation by preventing Ras activation of the ERK/mitogen-activated
protein kinase cascade. These results have implications for our understanding of
NF1 because a neurofibrosarcoma cell line derived from a patient with NF1 was
reverted by stable expression of the Pak dominant negative mutants.

PMID: 9560242 [PubMed - indexed for MEDLINE]


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7: Oncogene. 1998 Mar 26;16(12):1525-31. 

The OMgp gene, a second growth suppressor within the NF1 gene.

Habib AA, Gulcher JR, Hognason T, Zheng L, Stefansson K.

Department of Neurology, Beth Israel Deaconess Medical Center, Boston,
Massachusetts 02115, USA.

The Oligodendrocyte-Myelin glycoprotein gene (OMgp) is placed within an intron
of the NF1 gene. Neurofibromin, the product of NF1, acts as a RasGAP and
suppresses growth; inactivating mutations in NF1 lead to neurofibromatosis type
1. We report that OMgp also has growth suppressive effects and downregulates
mitogenic signaling pathways closely related to those influenced by
neurofibromin. Overexpression of OMgp alters mitogenic signaling in NIH3T3
fibroblasts. Cells overexpressing OMgp grow more slowly in serum compared to
controls and show a partial G1 block upon cell cycle analysis. PDGF is the
primary mitogen for fibroblasts in serum. Overexpression of OMgp alters PDGF
signaling in fibroblasts which results in a block of mitogenic signaling. PDGF
induced activation of c-Src is blocked, as is the induction of c-Myc and c-Fos,
while tyrosine phosphorylation of the PDGFbeta receptor, PLCgamma1 and induction
of c-Jun are intact. Although a number of genes embedded within other genes have
been described, the biological significance of this arrangement remains unknown.
We demonstrate here that structurally unrelated products of two such genes may
exercise closely related functions. Our data also raise the possibility of a
role for OMgp in disorders of cell proliferation such as NF1.

PMID: 9569019 [PubMed - indexed for MEDLINE]


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8: Yakugaku Zasshi. 1996 Jul;116(7):505-18. 

[Regulation mechanism of specific expression of tumor marker gene during
carcinogenesis]

[Article in Japanese]

Imagawa M.

Faculty of Pharmaceutical Sciences, Osaka University, Japan.

Rat glutathione transferase P (GST-P) is expressed at low levels in the normal
liver but becomes highly expressed in hyperplastic nodules and in hepatocellular
carcinomas during chemical hepatocarcinogenesis. To understand the regulation
mechanisms of this gene, we have characterized the 5'-flanking region and have
found that GST-P gene is regulated by at least two elements: one is a strong
enhancer and the other is a silencer. GST-P enhancer I (GPEI), located at -2.5
Kb, consists of two TPA-responsive element (TRE)-like sequences that are
palindromically oriented with 3 bp in between. It is well known that TRE is
activated by two nuclear oncogenes, c-Jun and c-Fos. Although GPEI is
trans-activated by these oncogenes, it is also active in F9 embryonal carcinoma
cells that lack c-Jun protein, suggesting that it can function with some
trans-activator other than AP-1 (c-Jun/c-Fos heterodimer). Indeed, another
protein is identified from the F9 nuclear extract. We have also identified a
silencer element at 300 bp upstream from the cap site. There are several
cis-elements in this region and at least three trans-acting factors bind to
these elements. We purified SF-A (silencer factor A) which binds to several
regions in this silencer, and determined the partial amino acid sequence.
Interestingly, SF-A seemed to be a related protein to NF1 (nuclear factor 1)
which is an activator for the transcription and DNA replication. Another factor
SF-B (silencer factor B) has been cloned and found to be the same as LIP (liver
inhibitory protein) which is a competitor for LAP (liver activator protein),
both are from the same gene designated as C/EBP beta. By transfection analysis
using GAL4 DNA binding domain we found LIP is not only a competitor but a direct
repressor. In the normal liver, another C/EBP family member, C/EBP alpha also
acts as a negative regulator, and this expression decreases during
hepatocarcinogenesis, resulting in the loss of silencer function. We carried out
the carcinogenesis experiments using transgenic rats harboring a chloramphenicol
acetyltransferase (CAT) reporter gene with -2900 to + 59 of the GST-P gene.
Liver foci and nodules produced by chemical carcinogens were found to express
high levels CAT activity by both CAT assay and immunohistochemical study, while
normal liver cells did not express any CAT activity. These results demonstrate
that the GST-P gene is trans-activated locus-independently during rat
hepatocarcinogenesis. Moreover, the similar results were obtained using
transgenic rats carrying GPEI-CAT, indicating that GPEI is an important
cis-element for activation of GST-P gene during hepatocarcinogenesis.

Publication Types:
    Review
    Review, Tutorial

PMID: 8831256 [PubMed - indexed for MEDLINE]


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9: Virus Genes. 1996;13(1):45-52. 

Induction of the HPV16 enhancer activity by Jun-B and c-Fos through cooperation
of the promoter-proximal AP-1 site and the epithelial cell type--specific
regulatory element in fibroblasts.

Kikuchi K, Taniguchi A, Yasumoto S.

Laboratory of Molecular and Cellular Biology, Kanagawa Cancer Center Research
Institute, Yokohama, Japan.

The epithelial cell type-specific enhancer of the human papillomavirus (HPV)
type 16 termed the long control region (LCR) carries three AP-1 binding sites.
We investigated the roles of the AP-1 sites for transactivation of the LCR by
Jun-B that may be a cell type specific-transactivator for the HPVs in human
fibroblasts in which expression of the endogenous Jun-B gene is low. Transient
expression of Jun-B alone poorly activated transcriptional activity of the LCR.
However, when combined with c-Fos, Jun-B activated the LCR. The
promoter-proximal AP-1 site was required for transactivation of the LCR by
Jun-B:c-Fos, but the site itself was not sufficient for the maximal response.
The proposed cell type specific--regulatory element that harbors binding sites
for NF1 as well as TEF-1 and PEA3 motifs, but neither other AP-1 sites nor the
proximal enhancer region, could augment the transcriptional response of the
promoter-proximal AP-1 site to Jun-B:c-Fos.

PMID: 8938978 [PubMed - indexed for MEDLINE]


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10: Virology. 1995 Aug 1;211(1):184-97. 

Transcriptional activity of human papillomavirus type 31b enhancer is regulated
through synergistic interaction of AP1 with two novel cellular factors.

Kyo S, Tam A, Laimins LA.

Department of Microbiology-Immunology, Northwestern University, Chicago,
Illinois 60611, USA.

Transcription of human papillomaviruses (HPV) is regulated by enhancer sequences
located in the upstream regulatory region. The factors regulating expression of
one of the high risk genital HPV types, HPV 31b, were investigated using
transient expression and protein binding assays. A region of 262 base pairs in
length was identified as the minimal functional enhancer and a series of five
protected binding sites were observed by footprint analyses. Electrophoretic
mobility shift assays demonstrated that AP1, Oct-1, as well as three novel
factors bound these sequences. Mutational analyses indicated that AP1
synergistically activated the HPV31b enhancer together with either of two novel
factors. One of these novel factors bound a sequence similar to an NF1 site but
was distinct from NF-1. The second factor bound sequences bearing similarity to
KRF-1 binding sites which have previously been characterized in HPV 18.
Competition binding assays demonstrated that this factor was not identical to
KRF-1. Additional studies implicated Oct-1 as a negative regulator of HPV 31b
expression as mutation of Oct-1 binding sequences resulted in an increase in
viral expression. None of the factors observed to be important for HPV 31b
enhancer activity was found exclusively in epithelial cells and instead were
detected in a variety of cell types. Of these factors, AP1 binding correlated
most strongly with enhancer function in a variety of cell types, implicating it
as a principal regulator of HPV expression. Variations in the constituents of
the AP1 complex that bind the HPV 31b enhancer were also observed in different
cell types, suggesting that changes in the distribution of jun proteins may play
a significant role in determining the tropism of HPV. These results indicate
that AP1 may be a common regulator for various HPV types and that it contributes
to enhancer specificity. In addition, a set of novel factors, which may be
specific for each HPV type, act synergistically with AP1 for full activation of
the enhancer.

PMID: 7645210 [PubMed - indexed for MEDLINE]


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11: Proc Natl Acad Sci U S A. 1995 May 9;92(10):4497-501. 

Inhibition of AP-1 binding and transcription by gold and selenium involving
conserved cysteine residues in Jun and Fos.

Handel ML, Watts CK, deFazio A, Day RO, Sutherland RL.

Cancer Biology Division, Garvan Institute of Medical Research, Sydney, New South
Wales, Australia.

Gold(I) salts and selenite, which have diverse therapeutic and biological
effects, are noted for their reactivity with thiols. Since the binding of
Jun-Jun and Jun-Fos dimers to the AP-1 DNA binding site is regulated in vitro by
a redox process involving conserved cysteine residues, we hypothesized that some
of the biological actions of gold and selenium are mediated via these residues.
In electrophoretic mobility-shift analyses, AP-1 DNA binding was inhibited by
gold(I) thiolates and selenite, with 50% inhibition occurring at approximately 5
microM and 1 microM, respectively. Thiomalic acid had no effect in the absence
of gold(I), and other metal ions inhibited at higher concentrations, in a rank
order correlating with their thiol binding affinities. Cysteine-to-serine
mutants demonstrated that these effects of gold(I) and selenite require Cys272
and Cys154 in the DNA-binding domains of Jun and Fos, respectively. Gold(I)
thiolates and selenite did not inhibit nonspecific protein binding to the AP-1
site and were at least an order of magnitude less potent as inhibitors of
sequence-specific binding to the AP-2, TFIID, or NF1 sites compared with the
AP-1 site. In addition, 10 microM gold(I) or 10 microM selenite inhibited
expression of an AP-1-dependent reporter gene, but not an AP-2-dependent
reporter gene. These data suggest a mechanism regulating transcription factor
activity by inorganic ions which may contribute to the known antiarthritic
action of gold and cancer chemoprevention by selenium.

PMID: 7753832 [PubMed - indexed for MEDLINE]


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12: J Biol Chem. 1994 Sep 30;269(39):24321-7. 

Activation of nuclear factor kappa B and oncogene expression by
12(R)-hydroxyeicosatrienoic acid, an angiogenic factor in microvessel
endothelial cells.

Laniado-Schwartzman M, Lavrovsky Y, Stoltz RA, Conners MS, Falck JR, Chauhan K,
Abraham NG.

Department of Pharmacology, New York Medical College, Valhalla 10595.

12(R)-Hydroxy-5,8,14(Z,Z,Z)-eicosatrienoic acid (12(R)-HETrE) is an arachidonic
acid metabolite formed by the corneal epithelium of several species, porcine
leukocytes, and human and rat epidermal cells. It is a potent, stereospecific
proinflammatory and angiogenic factor and its synthesis is increased manyfold in
inflamed tissues, e.g. cornea and skin. It is possible that the angiogenic
activity of 12(R)-HETrE is due to a direct mitogenic effect on microvessel
endothelial cells via yet to be elucidated cellular and molecular mechanisms. In
the present study, we demonstrated the ability of 12(R)-HETrE to stimulate the
growth of quiescent endothelial cells in a time- and concentration-dependent
manner with a maximal effect at 0.1 nM. This effect was highly stereospecific
since its enantiomer, 12(S)-HETrE, had no effect within the same concentration
range. Northern blot analysis and transient transfection experiments with
chloramphenicol acetyltransferase constructs of oncogene promoter regions
demonstrated significant increases over control (0.5% fetal calf serum) in
c-myc-, c-jun, and c-fos mRNA levels and expression in cells treated with 0.1 nM
12(R)-HETrE. Electrophoretic mobility shift assay of nuclear protein extracts
from cells treated with 12(R)-HETrE with specific radiolabeled oligonucleotides
corresponding to known transcriptional binding sites, including AP-1, AP-2, SP1,
TRE, NF kappa B, TFIID, OKT1, CREB, CTF/NF1, and GRE demonstrated a markedly
rapid and specific increase in the binding activity of NF kappa B and to a
lesser extent, AP-1. No significant increase was observed in the binding of
other transcription factors assayed as compared to control (untreated) cells.
Since the protooncogenes (c-fos, c-jun, and c-myc) are immediate early response
genes that are implicated in the process of cell proliferation and
differentiation, and activation of certain transcription factors, in particular
NF kappa B, is associated with the immediate response of the cell to an injury,
we propose that 12(R)HETrE's mitogenic and angiogenic activities are mediated,
in part, via the activation of NF kappa B and expression of these
protooncogenes.

PMID: 7523372 [PubMed - indexed for MEDLINE]


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13: Bioessays. 1994 Jul;16(7):489-96. 

Regulation of the Ras signalling network.

Maruta H, Burgess AW.

Ludwig Institute for Cancer Research, Melbourne, Australia.

The mitogenic action of cytokines such as epidermal growth factor (EGF) or
platelet derived growth factor (PDGF) involves the stimulation of a signal
cascade controlled by a small G protein called Ras. Mutations of Ras can cause
its constitutive activation and, as a consequence, bypass the regulation of cell
growth by cytokines. Both growth factor-induced and oncogenic activation of Ras
involve the conversion of Ras from the GDP-bound (D-Ras) to the GTP-bound
(T-Ras) forms. T-Ras activates a network of protein kinases including c-Mos,
c-Raf-1 and MAP kinase. Eventually the activation of MAP kinase leads to the
activation of the elongation factor 4E and several transcription factors such as
c-Jun, c-Myc and c-Fos. There are several modulators of Ras activity, such as
the GTPase activating proteins (GAP1 and NF1), which stimulate the conversion of
T-Ras to D-Ras. A series of small NF1 fragments, which bind T-Ras, as well as
truncated forms of derivatives of c-Raf-1, c-Jun and c-Myc, are capable of
blocking the T-Ras-activated mitogenesis in a competitive manner. These agents
offer a unique opportunity to control the proliferation of T-Ras-associated
tumors, which represent more than 30% of total human carcinomas.

Publication Types:
    Review
    Review, Tutorial

PMID: 7945277 [PubMed - indexed for MEDLINE]


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14: J Biol Chem. 1994 Jun 10;269(23):16512-7. 

Sphingosine 1-phosphate, a novel signaling molecule, stimulates DNA binding
activity of AP-1 in quiescent Swiss 3T3 fibroblasts.

Su Y, Rosenthal D, Smulson M, Spiegel S.

Department of Biochemistry and Molecular Biology, Georgetown University Medical
Center, Washington, District of Columbia 20007.

Sphingosine and sphingosine 1-phosphate, metabolites of sphingolipids, stimulate
cell proliferation in quiescent Swiss 3T3 fibroblasts and induce transient
increases in intracellular free calcium (Zhang, H., Desai, N. N., Olivera, A.,
Seki, T., Brooker, G., and Spiegel, S. (1991) J. Cell Biol. 114, 155-167).
However, little is yet known of the nuclear events that follow the early
responses induced by sphingolipid metabolites. Using a gel retardation assay, we
found that specific DNA binding activity of activator protein-1 (AP-1) was
markedly increased after treatment of quiescent Swiss 3T3 fibroblasts with
sphingosine 1-phosphate and sphingosine. The DNA binding specificity of AP-1 was
confirmed with competing probes containing consensus sequences of AP-1, AP-2,
AP-3, SP-1, and NF1/CTF. The c-fos gene product was detected in the AP-1 complex
using anti-c-Fos antibody. The dose response for stimulation of DNA binding
activity of AP-1 by sphingosine 1-phosphate correlated closely with its effect
on DNA synthesis. Furthermore, an inhibitor of sphingosine kinase,
DL-threo-dihydrosphingosine, which inhibits sphingosine-induced DNA synthesis
and the formation of sphingosine 1-phosphate, also inhibited
sphingosine-stimulated AP-1 DNA binding activity. This result further supports
our proposal that sphingosine 1-phosphate mediates the mitogenic effect of
sphingosine. Our results indicate that sphingosine 1-phosphate-induced DNA
synthesis and cell division may result from activation of AP-1 protein, linking
signal transduction by sphingolipid metabolites to gene expression.

PMID: 8206962 [PubMed - indexed for MEDLINE]


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15: Mol Cell Biol. 1993 Apr;13(4):2497-503. 

Differential regulation of cellular activities by GTPase-activating protein and
NF1.

al-Alawi N, Xu G, White R, Clark R, McCormick F, Feramisco JR.

Department of Medicine and Pharmacology, UCSD Cancer Center, La Jolla 92093.

The regulation of the GTPase activity of the Ras proteins is thought to be a key
element of signal transduction. Ras proteins have intrinsic GTPase activity and
are active in signal transduction when bound to GTP but not following hydrolysis
of GTP to GDP. Three cellular Ras GTPase-activating proteins (Ras-gaps) which
increase the GTPase activity of wild-type (wt) Ras but not activated Ras in
vitro have been identified: type I and type II GAP and type I NF1. Mutations of
wt Ras resulting in lowered intrinsic GTPase activity or loss of response to
cellular Ras-gap proteins are thought to be the primary reason for the
transforming properties of the Ras proteins. In vitro assays show type I and
type II GAP and the GAP-related domain of type I NF1 to have similar biochemical
properties with respect to activation of the wt Ras GTPase, and it appears as
though both type I GAP and NF1 can modulate the GTPase function of Ras in cells.
Here we report the assembling of a full-length coding clone for type I NF1 and
the biological effects of microinjection of Ras and Ras-gap proteins into
fibroblasts. We have found that type I GAP, type II GAP, and type I NF1 show
markedly different biological activities in vivo. Coinjection of type I GAP or
type I NF1, but not type II GAP, with wt Ras abolished the ability of wt Ras to
induce expression from an AP-1-controlled reporter gene. We also found that
serum-stimulated DNA synthesis was reduced by prior injection of cells with type
I GAP but not type II GAP or type I NF1. These results suggest that type I GAP,
type II GAP, and type I NF1 may have different activities in vivo and support
the hypothesis that while type I forms of GAP and NF1 may act as negative
regulators of wt Ras, they may do so with differential efficiencies.

PMID: 8455625 [PubMed - indexed for MEDLINE]


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16: Oncogene. 1991 Aug;6(8):1417-22. 

Interactions between adenovirus E1A and members of the AP-1 family of cellular
transcription factors.

Maguire K, Shi XP, Horikoshi N, Rappaport J, Rosenberg M, Weinmann R.

Wistar Institute, Philadelphia, Pennsylvania 19104.

We have studied interactions between bacterially produced E1A linked to
Sepharose and the various DNA-binding proteins present in HeLa cell nuclear
extracts (NE). DNA-binding activities and cross-reactive polypeptides
recognizing the cAMP-responsive element (CRE) and the activator protein 1 (AP1)
sites were bound to the E1A column, whereas nuclear factor 1 (NF1) and the
activator protein 2 (AP2) DNA-binding activities were not retained by E1A. The
binding activities that were retained belonged to the CRE and JUN protein
family, as judged by Western blot analysis. Authentic CRE-BP1, c-Jun and c-Fos
proteins produced by in-vitro translation also bound to the E1A column. However,
efficient binding of in-vitro-translated CRE-BP1 and c-Fos proteins to E1A
required preincubation with NE. We show here that immobilized E1A sequesters
several cellular upstream transcription activators, and suggest a role for
members of the AP1 family of transcription factors in E1A-mediated gene
regulation.

PMID: 1832215 [PubMed - indexed for MEDLINE]


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17: Oncogene. 1990 Sep;5(9):1285-90. 

Protein-binding elements in the promoter region of the mouse p53 gene.

Ginsberg D, Oren M, Yaniv M, Piette J.

Department of Chemical Immunology, Weizmann Institute of Science, Rehovot,
Israel.

p53 is a cellular protein whose expression plays a crucial role in the
regulation of cell proliferation and of neoplastic processes. p53 mRNA levels in
mouse fibroblasts can be elevated in response to TPA and to serum stimulation.
The promoter region of the p53 gene contains a conserved element which is highly
homologous to the consensus AP1 binding site (7/8 matching bases). This AP1-like
site, denoted the PF1 site, confers upon a heterologous promoter ability to
respond to elevated expression of c-jun. Furthermore, the PF1 site binds
protein(s) in a specific and serum-induced manner. Unexpectedly, this factor is
most probably not AP1, as evident from the inability of an authentic AP1 site to
compete the binding efficiently, as well as from the failure of purified AP1 to
bind to the PF1 site. Hence, PF1 may be a novel AP1-related transcription
factor. In addition, the 5' region of the p53 gene also contains an NF1 binding
site, whose location suggests a possible regulatory role.

PMID: 2216454 [PubMed - indexed for MEDLINE]


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18: Biochem Biophys Res Commun. 1990 Jul 16;170(1):140-6. 

Transcription factor levels in medullary thyroid carcinoma cells differentiated
by Harvey ras oncogene: c-jun is increased.

Nelkin BD, Borges M, Mabry M, Baylin SB.

Oncology Center, Johns Hopkins University School of Medicine, Baltimore, MD
21231.

In the TT cell line of human medullary thyroid carcinoma, the viral Harvey ras
(v-rasH) oncogene induces differentiation, marked by morphological changes,
diminution of growth, and increased expression of the calcitonin gene. Here, we
show that the transcriptional factor c-jun is increased during v-rasH induced
differentiation of TT cells both at the mRNA and functional protein levels. In
contrast, nuclear proteins with binding activities related to AP2, AP3, NF1/CTF,
and Sp1 were unchanged in v-rasH differentiated TT cells.

PMID: 2115330 [PubMed - indexed for MEDLINE]


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19: J Virol. 1990 May;64(5):2004-13. 

Adenovirus early region 3 promoter regulation by E1A/E1B is independent of
alterations in DNA binding and gene activation of CREB/ATF and AP1.

Kornuc M, Kliewer S, Garcia J, Harrich D, Li C, Gaynor R.

Department of Medicine, University of California, Los Angeles School of Medicine
90024.

Transcription of the adenovirus early region 3 promoter is strongly induced by
the adenovirus E1A protein. Previous DNase I footprinting has indicated that
four regions in this promoter serve as binding sites for HeLa nuclear proteins.
These include binding sites for NF-1 (site IV), AP1 (site III), CREB/activating
transcription factor (ATF) (site II), and TATA (site I). To determine the
relative importance of these sites in both the in vivo and in vitro
transcriptional regulation of the E3 promoter, oligonucleotide-directed
mutagenesis of these sites was performed. Each of these constructs was assayed
by transfection onto HeLa cells in the presence of either dl434, an E1A/E1B
deletion mutant, or wild-type adenovirus. Mutations of either the ATF- or
AP1-binding sites but not the TATA- and NF1-binding sites resulted in severe
decreases in both basal and E1A/E1B-induced transcriptional levels. These
constructs were also assayed in in vitro transcription assays with cellular
extracts prepared from dl434-infected or wild-type-adenovirus-infected HeLa
cells. The wild-type E3 promoter was transcribed approximately 30 times more
efficiently in extracts containing the E1A/E1B proteins compared with extracts
lacking these proteins. Mutations of either the TATA element, the ATF site, or
the AP1-binding site decreased both basal and E1A/E1B-induced transcriptional
levels. Gel retardation analysis using these extracts indicated that the binding
to ATF, AP1, or NF1 oligonucleotides was not altered in the presence of the
E1A/E1B proteins compared with extracts lacking these proteins. Northern (RNA)
blot analysis of c-jun and CREB RNA prepared from wild-type adenovirus and
dl434-infected cells indicated that the levels of these RNAs were not altered by
the E1A/E1B proteins. Immunoprecipitation of AP1 and CREB from both dl434- and
wild-type-adenovirus-infected cells indicated that the amounts of these proteins
were not significantly altered. These results suggest that E1A/E1B-induced
activation of the E3 promoter does not involve activation of transcription
factor genes nor a change in the DNA binding activity of important
promoter-binding components. Our results are consistent with a model in which
the E1A/E1B proteins either directly or indirectly alter the interactions of
factors that bind to the basal E3 promoter transcription complex, thereby
inducing transcription.

PMID: 2139139 [PubMed - indexed for MEDLINE]


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20: Nucleic Acids Res. 1990 Feb 11;18(3):465-70. 

Transcriptional activation of human papillomavirus 16 by nuclear factor I, AP1,
steroid receptors and a possibly novel transcription factor, PVF: a model for
the composition of genital papillomavirus enhancers.

Chong T, Chan WK, Bernard HU.

Institute of Molecular and Cell Biology, National University of Singapore, Kent
Ridge.

Human papillomavirus 16 (HPV-16), which is involved in genital carcinogenesis,
contains an enhancer of transcription that is activated by cellular factors
rather than by the viral E2 proteins. The activity resides on a 232 bp segment
with 5 binding sites for nuclear factor 1 (NF1), 2 for AP1, and 1 for steroid
receptors. Deletions and point mutations show that the constitutive enhancer and
the steroid response depend on NF1 sites located 5' or 3' of a 65 bp fragment
with AP1 sites that by itself shows little activity. Enhancement through a
fragment with AP1 and NF1 sites is strongly reduced by mutation of the AP1
sites, or by mutation of the sequence AGGCACATAT. Sequence comparison and
footprint analysis make it likely that this sequence binds a novel transcription
factor which we call PVF. Fragments with one or several binding sites only for
NF1, or AP1, or PVF exhibit little enhancement by themselves, suggesting the
functional dependence of the HPV-16 enhancer on the cooperation of these
factors. A comparison of our findings with the genomes and transcription factor
binding sites of HPV-6, 11, 18, 31 and 33 lead us to propose a model of the
composition of enhancers of genital papillomaviruses.

PMID: 2155400 [PubMed - indexed for MEDLINE]


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