title: Proximal cis-acting elements, including steroidogenic factor 1, mediate the efficiency of a distal enhancer in the promoter of the rat gonadotropin-releasing hormone receptor gene.
abstract: The gonadotrope-specific and regulated expression of the GnRH receptor (GnRH-R) gene is dependent on multiple transcription factors that interact with the noncanonical GnRH-R activating sequence (GRAS), the activator protein-1 (AP-1) element, and the steroidogenic factor-1 (SF-1) binding site. However, these three elements are not sufficient to mediate the complete cell-specific expression of the rat GnRH-R gene. In the present study, we demonstrate, by transient transfection in gonadotrope-derived alphaT3-1 and LssT2 cell lines, the existence of a distal enhancer [GnRH-R- specific enhancer (GnSE)] that is highly active in the context of the GnRH-R gene promoter. We show that the GnSE activity (-1,135/-753) is mediated through a functional interaction with a proximal region (-275/-226) that includes the SF-1 response element. Regions of similar length containing either the AP-1 or GRAS elements are less active or inactive. Transfection assays using an artificial promoter containing two SF-1 elements fused to a minimal PRL promoter indicate that SF-1 is crucial in this interaction. In addition, by altering the promoter with deletion and block- replacement mutations, we have identified the active elements of GnSE within two distinct sequences at positions -983/-962 and -871/-862. Sequence analysis and electrophoretic mobility shift experiments suggest that GnSE response elements interact, in these two regions, with GATA- and LIM-related factors, respectively. Altogether, these data establish the importance of the GnSE in the GnRH-R gene expression and reveal a novel role for SF-1 as a mediator of enhancer activity, a mechanism that might regulate other SF-1 target genes.
authors.affiliation: Endocrinologie Cellulaire et Moléculaire de la Reproduction, Université Pierre et Marie Curie, Centre National de la Recherche Scientifique, ESA 7080, Case 244, Paris cedex 05, France.
authors.forename: H
authors.initials: H
authors.lastname: Pincas
authors.forename: K
authors.initials: K
authors.lastname: Amoyel
authors.forename: R
authors.initials: R
authors.lastname: Counis
authors.forename: J N
authors.initials: JN
authors.lastname: Laverrière
mesh_terms: D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000494:pharmacology / Q000502:physiology*; D004742:Enhancer Elements, Genetic / Q000235:genetics; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins / Q000235:genetics; D016296:Mutagenesis; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear; D011966:Receptors, LHRH / Q000235:genetics*; D020218:Response Elements / Q000235:genetics; D017422:Sequence Analysis, DNA; D054339:Steroidogenic Factor 1; D013329:Structure-Activity Relationship; D018808:Transcription Factor AP-1 / Q000494:pharmacology; D014157:Transcription Factors / Q000494:pharmacology / Q000502:physiology*; D014162:Transfection
chemical_list: D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D015345:Oligonucleotide Probes; D018160:Receptors, Cytoplasmic and Nuclear; D011966:Receptors, LHRH; D054339:Steroidogenic Factor 1; D018808:Transcription Factor AP-1; D014157:Transcription Factors; C515060:steroidogenic factor 1, rat; D004247:DNA