title: GATA4 knockdown in MA-10 Leydig cells identifies multiple target genes in the steroidogenic pathway.
abstract: GATA4 is an essential transcription factor required for the initiation of genital ridge formation, for normal testicular and ovarian differentiation at the time of sex determination, and for male and female fertility in adulthood. In spite of its crucial roles, the genes and/or gene networks that are ultimately regulated by GATA4 in gonadal tissues remain to be fully understood. This is particularly true for the steroidogenic lineages such as Leydig cells of the testis where many in vitro (promoter) studies have provided good circumstantial evidence that GATA4 is a key regulator of Leydig cell gene expression and steroidogenesis, but formal proof is still lacking. We therefore performed a microarray screening analysis of MA-10 Leydig cells in which Gata4 expression was knocked down using an siRNA strategy. Analysis identified several GATA4-regulated pathways including cholesterol synthesis, cholesterol transport, and especially steroidogenesis. A decrease in GATA4 protein was associated with decreased expression of steroidogenic genes previously suspected to be GATA4 targets such as Cyp11a1 and Star. Gata4 knockdown also led to an important decrease in other novel steroidogenic targets including Srd5a1, Gsta3, Hsd3b1, and Hsd3b6, as well as genes known to participate in cholesterol metabolism such as Scarb1, Ldlr, Soat1, Scap, and Cyp51. Consistent with the decreased expression of these genes, a reduction in GATA4 protein compromised the ability of MA-10 cells to produce steroids both basally and under hormone stimulation. These data therefore provide strong evidence that GATA4 is an essential transcription factor that sits atop of the Leydig cell steroidogenic program.
authors.affiliation: ReproductionMother and Child Health, Room T3-67, Centre de Recherche du Centre Hospitalier Universitaire (CHU) de Québec, Centre de Recherche en Biologie de la Reproduction (CRBR), 2705 Laurier Boulevard, Quebec City, Quebec, Canada G1V 4G2Department of Obstetrics and GynecologyFaculty of Medicine, Laval University, Quebec City, Quebec, Canada G1K 7P4.
authors.forename: Francis
authors.initials: F
authors.lastname: Bergeron
authors.affiliation: ReproductionMother and Child Health, Room T3-67, Centre de Recherche du Centre Hospitalier Universitaire (CHU) de Québec, Centre de Recherche en Biologie de la Reproduction (CRBR), 2705 Laurier Boulevard, Quebec City, Quebec, Canada G1V 4G2Department of Obstetrics and GynecologyFaculty of Medicine, Laval University, Quebec City, Quebec, Canada G1K 7P4.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Nadeau
authors.affiliation: ReproductionMother and Child Health, Room T3-67, Centre de Recherche du Centre Hospitalier Universitaire (CHU) de Québec, Centre de Recherche en Biologie de la Reproduction (CRBR), 2705 Laurier Boulevard, Quebec City, Quebec, Canada G1V 4G2Department of Obstetrics and GynecologyFaculty of Medicine, Laval University, Quebec City, Quebec, Canada G1K 7P4 ReproductionMother and Child Health, Room T3-67, Centre de Recherche du Centre Hospitalier Universitaire (CHU) de Québec, Centre de Recherche en Biologie de la Reproduction (CRBR), 2705 Laurier Boulevard, Quebec City, Quebec, Canada G1V 4G2Department of Obstetrics and GynecologyFaculty of Medicine, Laval University, Quebec City, Quebec, Canada G1K 7P4 robert.viger@crchul.ulaval.ca.
authors.forename: Robert S
authors.initials: RS
authors.lastname: Viger
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002784:Cholesterol / Q000378:metabolism; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics / Q000378:metabolism; D050991:GATA4 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D012739:Gonadal Steroid Hormones / Q000096:biosynthesis*; D007985:Leydig Cells / Q000378:metabolism*; D008297:Male; D051379:Mice; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering; D000098682:Steroidogenic Acute Regulatory Protein
chemical_list: D050991:GATA4 Transcription Factor; C484171:Gata4 protein, mouse; D012739:Gonadal Steroid Hormones; D010750:Phosphoproteins; D034741:RNA, Small Interfering; D000098682:Steroidogenic Acute Regulatory Protein; D002784:Cholesterol; D002786:Cholesterol Side-Chain Cleavage Enzyme