Abstract
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/EBPalpha and -beta increased StAR promoter activity in COS-1 and HepG2 cells. Cotransfection of C/EBPalpha or -beta 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/EBPalpha and -beta 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/EBPbeta, but not C/EBPalpha, 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/EBPalpha and C/EBPbeta in human granulosa-lutein cell nuclear extracts. C/EBPbeta levels were up-regulated 3-fold by 8-Br-cAMP treatment. Our studies demonstrate a role for C/EBPbeta 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/EBPbeta participates in the cAMP regulation of StAR gene transcription.
Highlights
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
Recent observations revealed that ovarian CCAAT/enhancer-binding proteins (C/EBPs), basic leucine zipper transcription factors implicated in the regulation of a variety of genes involved in energy metabolism and cell differentiation pathways [15], are modulated by luteinizing hormone, which acts on ovarian cells through the intermediacy of cAMP, and that steroidogenic cell function is altered in mice upon loss of C/EBP␣ or -
The overall level of promoter activity in response to 8-Br-cAMP was reduced when the C/EBP binding sites were mutated, there was no change in the fold increase in promoter activity in response to 8-Br-cAMP stimulation, demonstrating that C/EBPs, and in particular C/EBP, and the cognate response elements regulate the basal level of StAR gene expression and boost the overall response to 8-Br-cAMP
Summary
StAR, steroidogenic acute regulatory protein; SF-1, steroidogenic factor 1; C/EBP, CCAAT/enhancer-binding protein; 8-Br-cAMP, 8-bromoadenosine 3Ј:5Ј-cyclic monophosphate; SREBP, sterol regulatory element-binding protein; DMEM, Dulbecco’s modified Eagle’s medium. The human StAR promoter was shown to have at least three SF-1 binding sites that are functionally important for basal as well as cAMP-stimulated transcription [8]. Sequence analysis indicated the presence of a variety of other transcription factor binding sites including multiple putative Sp-1 binding sites, several of which were recently implicated in transcriptional control of StAR gene expression [12]. Recent observations revealed that ovarian CCAAT/enhancer-binding proteins (C/EBPs), basic leucine zipper transcription factors implicated in the regulation of a variety of genes involved in energy metabolism and cell differentiation pathways [15], are modulated by luteinizing hormone, which acts on ovarian cells through the intermediacy of cAMP, and that steroidogenic cell function is altered in mice upon loss of C/EBP␣ or -. The focus of the present studies was to determine if C/EBPs modulate basal and cAMP-stimulated StAR gene expression
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
