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The Nuclear Receptor COUP-TFII Regulates Amhr2 Gene Transcription via a GC-Rich Promoter Element in Mouse Leydig Cells.

ABSTRACT: The nuclear receptor chicken ovalbumin upstream promoter-transcription factor type II (COUP-TFII)/NR2F2 is expressed in adult Leydig cells, and conditional deletion of the Coup-tfii/Nr2f2 gene impedes their differentiation. Steroid production is also reduced in COUP-TFII-depleted Leydig cells, supporting an additional role in steroidogenesis for this transcription factor. COUP-TFII action in Leydig cells remains to be fully characterized. In the present work, we report that COUP-TFII is an essential regulator of the gene encoding the anti-Müllerian hormone receptor type 2 (Amhr2), which participates in Leydig cell differentiation and steroidogenesis. We found that Amhr2 mRNA levels are reduced in COUP-TFII-depleted MA-10 Leydig cells. Consistent with this, COUP-TFII directly activates a -1486 bp fragment of the mouse Amhr2 promoter in transient transfection assays. The COUP-TFII responsive region was localized between -67 and -34 bp. Chromatin immunoprecipitation assay confirmed COUP-TFII recruitment to the proximal Amhr2 promoter whereas DNA precipitation assay revealed that COUP-TFII associates with the -67/-34 bp region in vitro. Even though the -67/-34 bp region contains an imperfect nuclear receptor element, COUP-TFII-mediated activation of the Amhr2 promoter requires a GC-rich sequence at -39 bp known to bind the specificity protein (SP)1 transcription factor. COUP-TFII transcriptionally cooperates with SP1 on the Amhr2 promoter. Mutations that altered the GCGGGGCGG sequence at -39 bp abolished COUP-TFII-mediated activation, COUP-TFII/SP1 cooperation, and reduced COUP-TFII binding to the proximal Amhr2 promoter. Our data provide a better understanding of the mechanism of COUP-TFII action in Leydig cells through the identification and regulation of the Amhr2 promoter as a novel target.

SUBMITTER: Mehanovic S

PROVIDER: S-EPMC6839530 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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The Nuclear Receptor COUP-TFII Regulates Amhr2 Gene Transcription via a GC-Rich Promoter Element in Mouse Leydig Cells.

Mehanovic Samir S Mendoza-Villarroel Raifish E RE Viger Robert S RS Tremblay Jacques J JJ

Journal of the Endocrine Society 20191001 12

The nuclear receptor chicken ovalbumin upstream promoter-transcription factor type II (COUP-TFII)/NR2F2 is expressed in adult Leydig cells, and conditional deletion of the Coup-tfii/Nr2f2 gene impedes their differentiation. Steroid production is also reduced in COUP-TFII-depleted Leydig cells, supporting an additional role in steroidogenesis for this transcription factor. COUP-TFII action in Leydig cells remains to be fully characterized. In the present work, we report that COUP-TF ...[more]

PMID: 31723721

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Project description:In males, Leydig cells are the main producers of testosterone and insulin-like 3 (INSL3), two hormones essential for sex differentiation and reproductive functions. Chicken ovalbumin upstream promoter-transcription factors I (COUP-TFI/NR2F1) and COUP-TFII (NR2F2) belong to the steroid/thyroid hormone nuclear receptor superfamily of transcription factors. In the testis, COUP-TFII is expressed and plays a role in the differentiation of cells committed to give rise to fully functional steroidogenic adult Leydig cells. Steroid production has also been shown to be diminished in COUP-TFII-depleted Leydig cells, indicating an important functional role in steroidogenesis. Until now, only a handful of target genes have been identified for COUP-TFII in Leydig cells. To provide new information into the mechanism of action of COUP-TFII in Leydig cells, we performed microarray analyses of COUP-TFII-depleted MA-10 Leydig cells. We identified 262 differentially expressed genes in COUP-TFII-depleted MA-10 cells. Many of the differentially expressed genes are known to be involved in lipid biosynthesis, lipid metabolism, male gonad development, and steroidogenesis. We validated the microarray data for a subset of the modulated genes by RT-qPCR. Downregulated genes included hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 1 (Hsd3b1), cytochrome P450, family 11, subfamily a, polypeptide 1 (Cyp11a1), prolactin receptor (Prlr), nuclear receptor subfamily 0, group B, member 2 (Shp/Nr0b2), ferredoxin 1 (Fdx1), scavenger receptor class B, member 1 (Scarb1), inhibin alpha (Inha), and glutathione S-transferase, alpha 3 (Gsta3). Finally, analysis of the Gsta3 and Inha gene promoters showed that at least two of the downregulated genes are potentially new direct targets for COUP-TFII. These data provide new evidence that further strengthens the important nature of COUP-TFII in steroidogenesis, androgen homeostasis, cellular defense, and differentiation in mouse Leydig cells.

Dataset Information

The Nuclear Receptor COUP-TFII Regulates Amhr2 Gene Transcription via a GC-Rich Promoter Element in Mouse Leydig Cells.

Publications

The Nuclear Receptor COUP-TFII Regulates <i>Amhr2</i> Gene Transcription via a GC-Rich Promoter Element in Mouse Leydig Cells.

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