title: Direct binding of AP-1 (Fos/Jun) proteins to a SMAD binding element facilitates both gonadotropin-releasing hormone (GnRH)- and activin-mediated transcriptional activation of the mouse GnRH receptor gene.
abstract: The response of pituitary gonadotropes to gonadotropin-releasing hormone (GnRH) correlates directly with the concentration of GnRH receptors (GnRHR) on the cell surface, which is mediated in part at the level of gene expression. Several factors are known to affect expression of the mouse GnRHR (mGnRHR) gene, including GnRH and activin. We have previously shown that activin augments GnRH-mediated transcriptional activation of mGnRHR gene, and that region -387/-308 appears to be necessary to mediate this effect. This region contains two overlapping cis-regulatory elements of interest: GnRHR activating sequence (GRAS) and a putative SMAD-binding element (SBE). This study investigates the role of these elements and their cognate transcription factors in transactivation of the mGnRHR gene. Transfection studies confirm the presence of GnRH- and activin-response elements within -387/-308 of mGnRHR gene promoter. Competition electrophoretic mobility shift assay experiments using -335/-312 as probe and alphaT3-1 nuclear extract or SMAD, Jun, and Fos proteins demonstrate direct binding of AP-1 (Fos/Jun) protein complexes to -327/-322 and SMAD proteins to -329/-328. Further transfection studies using mutant constructs of these cis-regulatory elements confirm that both are functionally important. These data define a novel cis-regulatory element comprised of an overlapping SBE and newly characterized non-consensus AP-1 binding sequence that integrates the stimulatory transcriptional effects of both GnRH and activin on the mGnRHR gene.
authors.affiliation: Department of Obstetrics, Gynecology and Reproductive Biology, Brigham & Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA. enorwitz@partners.org
authors.forename: Errol R
authors.initials: ER
authors.lastname: Norwitz
authors.forename: Shuyun
authors.initials: S
authors.lastname: Xu
authors.forename: Jian
authors.initials: J
authors.lastname: Xu
authors.forename: Lisa B
authors.initials: LB
authors.lastname: Spiryda
authors.forename: Joong Shin
authors.initials: JS
authors.lastname: Park
authors.forename: Kyeong-Hoon
authors.initials: KH
authors.lastname: Jeong
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: McGee
authors.forename: Ursula B
authors.initials: UB
authors.lastname: Kaiser
mesh_terms: D016475:3T3 Cells; D028341:Activins / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology*; D007700:Kinetics; D051379:Mice; D010902:Pituitary Gland / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic; D016760:Proto-Oncogene Proteins c-fos / Q000737:chemistry / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000737:chemistry / Q000378:metabolism; D011966:Receptors, LHRH / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D051785:Smad Proteins; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects / Q000502:physiology*; D014162:Transfection
chemical_list: D004268:DNA-Binding Proteins; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011966:Receptors, LHRH; D011993:Recombinant Fusion Proteins; D051785:Smad Proteins; D015534:Trans-Activators; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D028341:Activins; D007987:Gonadotropin-Releasing Hormone