HORMONAL-REGULATION OF THE CHOLESTEROL 7-ALPHA-HYDROXYLASE GENE (CYP7)

The transcriptional regulation of the rat cholesterol 7 alpha-hydroxyl ase gene (CYP7) by hormones and signal transduction pathways was studi ed by transient transfection assay of the promoter activity. HepG2 cel ls were transfected with deletion mutants of the CYP7 upstream region linked to the luciferase reporter gene. The transcription of CYP7/luci ferase chimeric genes was higher in confluent than in subconfluent cul tures of HepG2 cells. Glucocorticoid receptors, in the presence of dex amethasone, up-regulated the CYP7 gene through two regions located bet ween -3262 and -2803, and between -344 and -222, respectively. Thyroid hormones did not have any effect on the promoter activity. Insulin in hibited the promoter activity through sequences located between -344 a nd -222, and abolished the stimulation by dexamethasone. Hence, the in sulin effect was dominant over that of glucocorticoids. Treatment of t ransfected HepG2 cells with phorbol 12-myristate 13-acetate (PMA), a k nown activator of protein kinase C (PKC), resulted in a time-dependent inhibition of the CYP7 promoter activity. The negative phorbol ester- response sequences were mapped between -344 and -222, and between -200 and -161, respectively. The CYP7 promoter activity was induced nearly Ei-fold by all-trans-retinoic acid through sequences in the region fr om -200 to -129. Finally, cyclic AMP and protein kinase A (PKA) stimul ated the expression of the CYP7/luciferase gene through multiple seque nces in the distal and proximal regions, and both positive and negativ e response regions were mapped. Our results revealed that the -416 fra gment of the rat CYP7 gene confers the activation by glucocorticoids a nd retinoic acid, and inhibition by insulin, phorbol esters and cAMP. It appears that this proximal promoter may contain a pleiotropic domai n that regulates the effects of multiple signals.