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Regulation of P450c17 Expression in the Early Embryo Depends on GATA Factors

Department of Obstetrics, Gynecology, and Reproductive Science, The Center for Reproductive Sciences, University of California, San Francisco, San Francisco, California 94143

Address all correspondence and requests for reprints to: Synthia H. Mellon, Ph.D., Department of Obstetrics, Gynecology, and Reproductive Science, University of California San Francisco, 513 Parnassus Avenue, Box 0556, San Francisco, California 94143-0556. E-mail: mellon{at}cgl.ucsf.edu.

The enzyme P450c17 is required for glucocorticoid, sex steroid, and some neurosteroid biosynthesis. Defective human P450c17 causes sexual infantilism and 46,XY sex reversal but is compatible with life, whereas ablation of the corresponding mouse gene causes embryonic lethality at around E7. Normal mouse embryos express P450c17 protein and activity in the embryonic endoderm at E7. Adult adrenal and gonadal steroidogenesis requires steroidogenic factor-1 (SF-1), but SF-1 is not expressed in the early mouse embryo. We show that P450c17 is expressed in differentiated mouse parietal and visceral endoderm lineages, in cultured mouse F9 embryonic carcinoma stem cells, in mouse embryonic stem cells, and in cultured mouse P19 stem cells. Bases –110 to –55 (which contain an SF-1 site and two potential GATA sites) of the rat cyp17 gene confer promoter activity in F9 cells. Overexpression of SF-1 has no effect, whereas overexpression of GATA4 in F9 cells increases transcription from –110/–55 fused to a reporter and increases endogenous P450c17 mRNA. Chromatin immunoprecipitation assays show that GATA4 binds to –215/+55 of mouse cyp17. Stimulating F9 cells with retinoic acid and cAMP differentiates them into visceral and parietal endoderm. Commensurate with cell differentiation, quantitative PCR showed increased GATA4 and GATA6 mRNAs, temporally followed by increased P450c17 mRNA. Small interfering RNA inhibition of GATA4 or GATA6 in undifferentiated or differentiated F9 cells diminished endogenous cyp17 expression. Thus, P450c17 is expressed in mouse embryonic stem cells, its expression increases upon differentiation to an early embryonic endoderm lineage, and GATA4/6 are responsible for activation of P450c17 gene expression at this early stage of embryonic development.