Transcriptional regulation of human 11 beta-hydroxylase (hCYP11B1)

Steroid 11 beta-hydroxylase is a mitochondrial enzyme that catalyzes the co nversion of deoxycortisol to cortisol. The gene encoding human 11 beta-hydr oxylase (hCYP11B1) is expressed in the adrenal cortex under the control of circulating levels of ACTH. The current study was undertaken to define the cis-regulatory elements and transacting factors that regulate hCYP11B1 tran scription. The hCYP11B1 5'-flanking DNA was studied using transient transfe ction of luciferase reporter constructs in NCI-H295R human adrenocortical c ells. A cAMP analogue ((Bu)(2)cAMP) increased expression of a construct con taining -1102 bp of hCYP11B1 5'-flanking DNA (pB1-1102). An element at posi tion -71/-64 (TGACGTGA, previously termed Ad1) resembling a consensus cAMP response element (CRE) was required for maximal induction by cAMP. The Adl element bound several transcriptional factors in electrophoretic mobility s hift assays, including CRE-binding protein, activating transcription factor -1 (ATF-1), and ATF-2, but only the ATF-2 complex migrated similarly to a c omplex seen using H295R nuclear extract. In addition, Western analysis of H 295R and adrenal lysates demonstrated expression of high levels of ATF-2 an d ATF-1. CRE-binding protein levels varied among the strains of H295R cells tested. Transcription of CYP11B1 also appeared to be regulated by steroido genic factor-1 (SF-1). Luciferase reporter gene activity was increased afte r cotransfection with expression vectors containing SF-l. An element in hCY P11B1 at positions -242/-234 (CCAAGGCTC), previously termed Ad4, was requir ed for maximal induction by SF-1 and was found to bind SF-1 in electrophore tic mobility shift assays. The key role for SF-1 in hCYP11B1 transcription is in contrast to its lack of an effect on expression of the hCYP11B2 (aldo sterone synthase) isozyme. The differential effects of SF-l on transcriptio n of hCYP11B1 and hCYP11B2 may be one of the mechanisms controlling differe ntial expression of these isozymes within the zonae fasciculata and glomeru losa of the human adrenal cortex.