title: KLF6 cooperates with NUR77 and SF1 to activate the human INSL3 promoter in mouse MA-10 leydig cells.
abstract: Insulin-like 3 (INSL3), a Leydig cell-specific hormone, is essential for testis descent during foetal life and bone metabolism in adults. Despite its essential roles in male reproductive and bone health, very little is known regarding its transcriptional regulation in Leydig cells. To date, few transcription factors have been shown to activate INSL3 promoter activity: the nuclear receptors AR, NUR77, COUP-TFII and SF1. To identify additional regulators, we have isolated and performed a detailed analysis of a 1.1 kb human INSL3 promoter fragment. Through 5' progressive deletions and site-directed mutagenesis, we have mapped a 10 bp element responsible for about 80% of INSL3 promoter activity in Leydig cells. This element is identical to the CPE element of the placental-specific glycoprotein-5 (PSG5) promoter that is recognized by the developmental regulator Krüppel-like factor 6 (KLF6). Using PCR and western blotting, we found that KLF6 is expressed in several Leydig and Sertoli cell lines. Furthermore, immunohistochemistry on adult mouse testis revealed the presence of KLF6 in the nuclei of both Leydig and Sertoli cells. KLF6 binds to the 10 bp KLF element at -108 bp and activates the -1.1 kb human, but not the mouse, INSL3 promoter. KLF6-mediated activation of the human INSL3 promoter required an intact KLF element as well as Leydig/Sertoli-enriched factors because KLF6 did not stimulate the human INSL3 promoter activity in CV-1 fibroblast cells. Consistent with this, we found that KLF6 transcriptionally cooperates with NUR77 and SF1. Collectively, our results identify KLF6 as a regulator of human INSL3 transcription.
authors.affiliation: ReproductionMother and Child Health, Centre de recherche du centre hospitalier universitaire de Québec, Québec City, Québec, Canada.
authors.forename: Maxime A
authors.initials: MA
authors.lastname: Tremblay
authors.affiliation: ReproductionMother and Child Health, Centre de recherche du centre hospitalier universitaire de Québec, Québec City, Québec, Canada.
authors.forename: Raifish E
authors.initials: RE
authors.lastname: Mendoza-Villarroel
authors.affiliation: ReproductionMother and Child Health, Centre de recherche du centre hospitalier universitaire de Québec, Québec City, Québec, Canada.
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Robert
authors.affiliation: ReproductionMother and Child Health, Centre de recherche du centre hospitalier universitaire de Québec, Québec City, Québec, Canada.
authors.forename: Francis
authors.initials: F
authors.lastname: Bergeron
authors.affiliation: ReproductionMother and Child Health, Centre de recherche du centre hospitalier universitaire de Québec, Québec City, Québec, Canada Centre for Research in ReproductionDevelopment and Intergenerational Health, Department of Obstetrics, Gynecology, and Reproduction, Faculty of Medicine, Université Laval, Québec City, Québec, Canada Jacques-J.Tremblay@crchudequebec.ulaval.ca.
authors.forename: Jacques J
authors.initials: JJ
authors.lastname: Tremblay
mesh_terms: D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002874:Chromosome Mapping; D005786:Gene Expression Regulation; D006801:Humans; D007328:Insulin / Q000235:genetics*; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism*; D007985:Leydig Cells / Q000378:metabolism*; D008297:Male; D051379:Mice; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1 / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011506:Proteins / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D000072260:RNA Splicing Factors / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid; D020218:Response Elements; D013737:Testis / Q000378:metabolism; D015533:Transcriptional Activation
chemical_list: D007328:Insulin; C491358:Klf6 protein, mouse; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors; C084797:Leydig insulin-like protein; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D011506:Proteins; D011518:Proto-Oncogene Proteins; D000072260:RNA Splicing Factors; C491514:Sf1 protein, mouse
keywords: KLF; cooperation; insulin-like 3; leydig cells; nuclear receptor; testis; transcription