CCAAT/enhancer-binding proteins regulate expression of the human steroidogenic acute regulatory protein (StAR) gene

Two putative CCAAT/enhancer-binding protein (C/EBP) response elements were identified in the proximal promoter of the human steroidogenic acute regulatory protein (STAR) gene, which encodes a key protein-regulating steroid hormone synthesis. Expression of C/EBPα and -β increased StAR promoter activity in COS-1 and HepG2 cells. Cotransfection of C/EBPα or -β and steroidogenic factor 1, a transcription factor required for cAMP regulation of StAR expression, into COS-1 augmented 8-bromoadenosine 3':5'- cyclic monophosphate (8-Br-cAMP)-stimulated promoter activity. When the putative C/EBP response elements were mutated, individually or together, a pronounced decline in basal StAR promoter activity in human granulosa-lutein cells resulted, but the fold stimulation of promoter activity by 8-Br-cAMP was unaffected. Recombinant C/EBPα and -β bound to the two identified sequences but not the mutated elements. Human granulosa-lutein cell nuclear extracts also bound these elements but not the mutated sequences. An antibody to C/EBPβ, but not C/EBPα, supershifted the nuclear protein complex associated with the more distal element. The complex formed by nuclear extracts with the proximal element was not supershifted by either antibody. Western blot analysis revealed the presence of C/EBPα and C/EBPβ in human granulosa-lutein cell nuclear extracts. C/EBPβ levels were up-regulated 3- fold by 8-Br-cAMP treatment. Our studies demonstrate a role for C/EBPβ as well as yet to be identified proteins, which can bind to C/EBP response elements, in the regulation of StAR gene expression and suggest a mechanism by which C/EBPβ participates in the cAMP regulation of StAR gene transcription.