Phorbol ester inhibition of rat gonadotropin-releasing hormone promoter activity: role of Fos and Jun in the repression of transcription.
Treatment of GT1-7 neuronal cells with the phorbol ester, 12-O-tetradecanoyl phorbol 13-acetate (TPA), inhibits GnRH gene transcription. The present studies investigated the role of AP-1 (Fos and Jun) in this repression. Treatment of cells with TPA increased c-fos mRNA 20-fold with only a 2-fold increase in c-jun mRNA levels. In transient transfection studies, a luciferase expression vector containing fragments of the 5'-flanking DNA of the rat GnRH (rGnRH) promoter was cotransfected with Fos and Jun expression vectors to mimic the effects of TPA. A dose-dependent decrease in reporter activity was noted with increasing amounts of Fos but not with Jun overexpression. Deletion analysis mapped the region that mediates repression by AP-1 to the area between -126 and -73 base pairs (bp) of the rGnRH 5'-flanking region: the same area that mediates TPA-induced repression and contains an imperfect TPA response element sequence at -99. Gel retardation assays, however, showed that a DNA fragment from -111 to -73 of the rGnRH promoter does not directly interact with Fos in GT1-7 extracts. Coexpression of Fos proteins with mutations in the DNA-binding region, the dimerization domain, or carboxy terminus partially blocked inhibition of rGnRH promoter activity. These data support a novel mechanism of AP-1 repression of GnRH transcription that is mediated by Fos interaction with other protein(s) that directly bind to the proximal rGnRH promoter.
Bruder
J M
JM
Department of Medicine, University of Colorado Health Sciences Center, Denver 80220, USA.
Spaulding
A J
AJ
Wierman
M E
ME
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism; D015870:Gene Expression; D016762:Genes, fos / Q000235:genetics*; D016758:Genes, jun / Q000235:genetics*; D007987:Gonadotropin-Releasing Hormone / Q000235:genetics*; D016350:Leucine Zippers; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D018808:Transcription Factor AP-1 / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D007987:Gonadotropin-Releasing Hormone; D004247:DNA; D008156:Luciferases; D013755:Tetradecanoylphorbol Acetate