Epidermal growth factor receptors (EGFR) play an important role in cell growth and development. Signaling from the EGFR has many effects including cell growth and division, proliferation and angiogenesis. EGFRs are inappropriately expressed or highly expressed in a variety of human tumors including breast, brain, prostate and others. Overexpression of the EGFR has been associated with advanced tumor stage, resistance to standard therapies and, in some tumors, with poor patient prognosis. The critical role of EGFR in both normal and abnormal cell growth and development necessitates a clear understanding of the regulation of its' expression. We hypothesize that changes in transcription factors interacting with the EGFR promoter leads to increased expression in the absence of gene amplification. Our research focuses on two projects seeking to understand the factors that regulate EGFR expression during normal cell growth and in cancer. We seek to determine the transcription factors involved in EGFR gene regulation and to examine their role in cancer. We further seek to understand how GC-binding factor 2 (GCF2), a repressor of EGFR expression, decreases the activity of cellular and viral promoters. We initially characterized the EGFR promoter region as a GC-rich, TATA-less regulatory region with multiple transcription initiation sites and specificity protein 1 (Sp1) binding sites. Many DNA-binding factors have been identified that interact with the promoter region including p53, activator protein 2 (AP2) and interferon regulated factor 1 (IRF-1). We have determined the roles of activator protein 1 (AP-1), p63 and early growth response gene 1 (egr-1) in regulating EGFR expression. We determined that AP-1 bound to at least seven sites in the promoter and partially mediated phorbol ester induced EGFR expression. We found that the p53 homologue, p63, repressed EGFR expression through protein-protein interactions with Sp1, an activator of EGFR expression. Egr-1 was determined to increase endogenous EGFR expression. Furthermore, we were able to show that upregulation of egr-1 during hypoxia leads to increased EGFR expression. Additionally, we determined that nuclear factor kappa B (NF-kB) bound to the EGFR promoter but did not transactivate. Expression of the EGFR is regulated by p53. The p53 homologue, p73, has a high degree of homology and possibly similar functions. Analysis of p73 in ovarian carcinoma cell lines has revealed that 71% of invasive tumors and 92% of borderline tumor tissues express elevated levels of p73 transcript and protein. This increased level of p73 expression and the association of EGFR with a more invasive and metastatic phenotype has prompted us to determine if p73 regulates EGFR expression in ovarian cancer cells. We hypothesize that increased p73 expression in ovarian cancer leads to increased EGFR expression. Using transient transfection experiments, we determined that increased p73-alpha expression resulted in a 4 - 5 fold increase in EGFR promoter activity. Consistent with this finding was a similar increase in the endogenous EGFR level. We further determined that p73-alpha binds to the previously defined EGFR p53 response element. Mutation of this element abrogates the p73 induction. We have cloned and characterized GCF2, a transcriptional repressor. We have determined the GCF2 binding site in the EGFR promoter and shown that GCF2 can partially inhibit AP2 activation of EGFR promoter activity. In addition, we have shown that GCF2 was able to bind to a highly structured RNA element TAR (transactivation response element) that is located at the 5' end of nascent HIV transcripts. GCF2 inhibits activation of the HIV-1 LTR by a small regulatory protein, Tat, which is required for efficient transcription of genes linked to the HIV-1 LTR. We further determined that the amino terminal region of GCF2 was required to repress the Tat-mediated enhancement of the HIV-1 LTR and that GCF2 can also decrease basal HIV-1 LTR activity. We have also identified two additional members of the GCF2 family, GCF2-SK (skeletal muscle) and GCF2-SM (smooth muscle). GCF2-SK is encoded by a 2.9 kilobase mRNA found predominantly in skeletal muscle and heart tissues. GCF2-SM is encoded by a 2.4 kilobase mRNA found in most tissues. GCF2-SK appears to repress expression from several promoters while GCF2-SM does not. Analysis of these gene products should aid in understanding the function of GCF2.

National Institute of Health (NIH)
Division of Basic Sciences - NCI (NCI)
Intramural Research (Z01)
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Basic Sciences
United States
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Nishi, Hirotaka; Neta, Gila; Nishi, Katsura H et al. (2003) Analysis of the epidermal growth factor receptor promoter: the effect of nuclear factor-kappaB. Int J Mol Med 11:49-55
Rikiyama, Toshiki; Curtis, Joseph; Oikawa, Masaya et al. (2003) GCF2: expression and molecular analysis of repression. Biochim Biophys Acta 1629:15-25
Nishi, H; Senoo, M; Nishi, K H et al. (2001) p53 Homologue p63 represses epidermal growth factor receptor expression. J Biol Chem 276:41717-24
Johnson, A C; Murphy, B A; Matelis, C M et al. (2000) Activator protein-1 mediates induced but not basal epidermal growth factor receptor gene expression. Mol Med 6:17-27
Liu, X W; Katagiri, Y; Jiang, H et al. (2000) Cloning and characterization of the promoter region of the rat epidermal growth factor receptor gene and its transcriptional regulation by nerve growth factor in PC12 cells. J Biol Chem 275:7280-8
Khachigian, L M; Santiago, F S; Rafty, L A et al. (1999) GC factor 2 represses platelet-derived growth factor A-chain gene transcription and is itself induced by arterial injury. Circ Res 84:1258-67