Cloning and Characterization of a 5′ Regulatory Region of the Prolactin Receptor-Associated Protein/17β Hydroxysteroid Dehydrogenase 7 Gene

Abstract

AbstractProlactin receptor-associated protein (PRAP) originally cloned in our laboratory was shown to be a novel, luteal isoform of 17β hydroxysteroid dehydrogenase 7 (17βHSD7). In this study, we cloned the promoter region of rat PRAP/17βHSD7 and investigated the mechanisms regulating both basal activity and LH-induced repression of this promoter. Truncated and site-specific mutants of PRAP/17βHSD7 promoter identified two enhancer regions that contained highly conserved Sp1 binding site and bound Sp1 from nuclear extracts of both corpora lutea and a rat luteal cell line. Repression of PRAP/17βHSD7 expression and promoter activity by human chorionic gonadotropin/forskolin was localized to a −52-bp proximal segment of the promoter. This region contained a conserved CCAAT site and bound nuclear factor Y; binding of this transcription factor was inhibited by human chorionic gonadotropin in vivo. Furthermore, mutation of the nuclear factor Y site in the −52-bp promoter-reporter construct abolished forskolin-mediated inhibition of the promoter in a rat luteal cell line. In summary, we have identified the promoter elements involved in the basal expression of PRAP/17βHSD7. We have also found that LH-mediated repression of this gene is at the level of transcription and involves inhibition of nuclear factor YA binding to the CCAAT site within the proximal promoter.