Stat5-mediated regulation of the human type II 3 beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4) isomerase gene: Activation by prolactin

Altered PRL levels are associated with infertility in women. Molecular targ ets at which PRL elicits these effects have yet to be determined. These stu dies demonstrate transcriptional regulation by PRL of the gene encoding the final enzymatic step in progesterone biosynthesis: 3 beta-hydroxysteroid d ehydrogenase/Delta(5)-Delta(4) isomerase (3 beta-HSD). A 9/9 match with the consensus State response element was identified at -110 to -118 in the hum an Type II 3 beta-HSD promoter. 3 beta-HSD chloramphenicol acetyltransferas e (CAT) reporter constructs containing either an intact or mutated State el ement were tested for PRL activation. Expression vectors for State and the PRL receptor were cotransfected with a -300 --> +45 3 beta-HSD CAT reporter construct into HeLa cells, which resulted in a 21-fold increase in reporte r activity in the presence of PRL. Promoter activity showed an increased re sponse with a stepwise elevation of transfected State expression or by trea tment with increasing concentrations of PRL (max, 250 ng/ml). This effect w as dramatically reduced when the putative State response element was remove d by 5'-deletion of the promoter or by the introduction of a 3-bp mutation into critical nucleotides in the element. Furthermore, P-32-labeled promote r fragments containing the State element were shifted in electrophoretic mo bility shift assay experiments using nuclear extracts from cells treated wi th PRL, and this complex was supershifted with antibodies to State. These r esults demonstrate that PRL has the ability to regulate expression of a key human enzyme gene (type II 3 beta-HSD) in the progesterone biosynthetic pa thway, which is essential for maintaining pregnancy.