title: AR suppresses transcription of the LHbeta subunit by interacting with steroidogenic factor-1.
abstract: Synthesis of LH is suppressed by feedback from gonadal steroids. Previously, we demonstrated that 779 bp of the bovine LHbeta promoter was sufficient to target expression of a chloramphenicol acetyltransferase reporter gene specifically to the pituitary in transgenic mice, and found that it was appropriately suppressed after administration of T or E2. In this study, we report that ligand-bound AR, but not ligand-bound ER, directly suppressed activity of the bovine LHbeta promoter when examined in a gonadotrope-derived cell line. Additional studies with mutated bovine LHbeta promoter constructs focused on the proximal 5'-flanking region because of the presence of several cis-acting elements that are highly conserved across all mammals. These include regulatory elements that bind steroidogenic factor 1 (SF-1), Egr-1, and Pitx1. When tested by cotransfection with AR, overexpression of Egr-1, Pitx1, and constitutively active steroidogenic factor 1 (SF-1DeltaLBD) each individually rescued androgen-mediated suppression of the bovine LHbeta promoter. This suggested a functional interaction between each of these transcription proteins and AR. In contrast, overexpression of full-length SF-1 was incapable of relieving the bovine LHbeta promoter from the suppressive effect imposed by AR. This suggested that the ligand-binding domain of SF-1 plays an important role in functional interactions that occur between this protein and AR. This notion was further supported by binding assays performed with glutathione-S-transferase-AR: these identified SF-1 as a key interactive partner and localized this interaction to the ligand-binding domain of the protein. Additional binding studies indicated that protein interactions between SF-1, Pitx1, and Egr-1 interfere with formation of a binary complex that contains AR and SF-1. Thus, we conclude that AR suppresses activity of the bovine LHbeta promoter through protein-protein interactions with SF-1 and that the degree of this interaction can be modified by the presence of Egr-1 and Pitx1.
authors.affiliation: Department of Pharmacology, Case Western Reserve University, Cleveland, Ohio 44106-4965, USA.
authors.forename: J S
authors.initials: JS
authors.lastname: Jorgensen
authors.forename: J H
authors.initials: JH
authors.lastname: Nilson
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D051766:Early Growth Response Protein 1; D051717:Fushi Tarazu Transcription Factors; D005786:Gene Expression Regulation / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D017874:Immediate-Early Proteins; D007986:Luteinizing Hormone / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D051761:Paired Box Transcription Factors; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D011944:Receptors, Androgen / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear; D011960:Receptors, Estrogen / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D054339:Steroidogenic Factor 1; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D017874:Immediate-Early Proteins; C515057:NR5A1 protein, human; D051761:Paired Box Transcription Factors; D021122:Protein Subunits; D011944:Receptors, Androgen; D018160:Receptors, Cytoplasmic and Nuclear; D011960:Receptors, Estrogen; D011993:Recombinant Fusion Proteins; D054339:Steroidogenic Factor 1; D014157:Transcription Factors; C101485:homeobox protein PITX1; C515058:steroidogenic factor 1, mouse; D007986:Luteinizing Hormone