Proximal GATA-binding sites are essential for human HSD3B1 gene transcription in the placenta

Abstract

Abstract The enzyme 3β-hydroxysteroid dehydrogenase/isomerase (3β-HSD) is involved in the synthesis of active steroid hormones. Two human 3β-HSD isoforms are expressed in a tissue-specific pattern. HSD3B1 (type I) expression is essential to produce progesterone for pregnancy maintenance. To understand the mechanisms of human HSD3B1 activation in the placenta, 2.2 kb of 5′-flanking sequence and 5′-deletions were fused to the luciferase reporter gene and transfected into human JEG-3 cells. The proximal −238/+337 sequence had the highest promoter activity. Two GATA elements were identified at −106/−99 and −52/−45. Mutations of either sites greatly reduced promoter activity in JEG-3 cells, demonstrating the importance of GATA sites. EMSA revealed the specific binding of GATA2 and GATA3 to the GATA sequences at −106/−99 and −52/−45. ChIP assays demonstrated the association of GATA2 but not GATA3 with the GATA-binding regions of the HSD3B1 promoter in JEG-3 cells. GATA2 knockdown significantly reduced HSD3B1 expression in JEG-3 cells; however, GATA3 knockdown increased HSD3B1 expression. Western blot analysis revealed high levels of GATA2 but not GATA3 in human placental tissues. This study identified GATA motifs as essential control elements for HSD3B1 transcription and GATA2 as a novel transcriptional regulator of HSD3B1 expression in the human placenta.