Phosphorylation of CtBP1 by cAMP-dependent Protein Kinase Modulates Induction of CYP17 by Stimulating Partnering of CtBP1 and 2

Abstract

In the human adrenal cortex, the peptide hormone adrenocorticotropin (ACTH) directs cortisol and adrenal androgen biosynthesis by activating a cAMP/cAMP-dependent protein kinase (PKA) pathway. Carboxyl-terminal binding protein 1 (CtBP1) is a corepressor that regulates transcription of the CYP17 gene by periodically interacting with steroidogenic factor-1 in response to ACTH signaling. Given that CtBP1 function is regulated by NADH binding, we hypothesized that ACTHstimulated changes in cellular pyridine nucleotide concentrations modulate the ability of CtBP1 to repress CYP17 transcription. Further, we postulated that PKA evokes changes in the phosphorylation status of CtBP1 that control the ability of the protein to bind to steroidogenic factor-1 and the coactivator GCN5 (general control nonderepressed 5) and repress CYP17 gene expression. We show that ACTH alters pyridine nucleotide redox state and identify amino acid residues in CtBP1 that are targeted by PKA and PAK6. Both ACTH/cAMP signaling and NADH/NAD ؉ ratio stimulate nuclear-cytoplasmic oscillation of both CtBP proteins. We provide evidence that PKA 1) induces metabolic changes in the adrenal cortex and 2) phosphorylates CtBP proteins, particularly CtBP1 at T144, resulting in CtBP protein partnering and ACTH-dependent CYP17 transcription. CtBP12 was originally identified as a repressor of the adenoviral early antigen 1A oncogenic transcription activator (1) but has subsequently been shown to have 2-hydroxy acid dehydrogenase activity and a conserved lactate dehydrogenase structure (2, 3), although the endogenous substrates targeted by this activity are probably different (4). Perhaps via a preference for binding of reduced pyridine nucleotide (5, 6) or innate dehydrogenase activity (7), CtBP1 and 2 are able to regulate numerous genes with dependence on NADH concentration and thus the energy and redox state of the cell. Intriguingly, CtBP1 transcriptionally represses the SirT1 histone deacetylase when NADH is elevated (8), possibly regulating SirT1 energy-restricted longevity in some species. E-cadherin is repressed by CtBP (9 -11) during hypoxia-induced tumor metastasis, where hypoxia significantly increases cellular NADH (9). Of note, there is a report that hypoxia stimulates adrenal output of corticosterone secretion and steroidogenic acute regulatory protein and peripheral benzodiazepine receptor in rat pups (12).Although CtBP represses transcription of nuclear receptor target genes (13-19), CtBP1 has only been shown to interact with SF-1 (20). Bartholin et al. (13) showed that the functional CtBP interaction motif of a retinoid X receptor-interacting corepressor is required for acute repression of a retinoic acidinducible reporter and retinoic acid-induced teratogenesis. CtBP1 and 2 reversibly interact with and reinforce the corepressor function of RIP140 (140-kDa receptor interacting protein) (14,15,17,18), which in a ligand-dependent manner interacts with nuclear receptors (21, 22), including SF-1 (23), via one or more of several NR boxes (LXX(L/...

Involvement of miRNAs in ovarian follicular and luteal development

“…This conclusion is in contrast with the view that miRNAs act primarily as translational repressors rather than activators. CTBP1 can regulate CYP17A1 expression by interacting with NR5A1 in adrenal cells (Dammer & Sewer 2008). However, LNA-mediated inhibition of miR-132 and miR-212 had no effects on oestradiol or progesterone production by granulosa cells and therefore Fiedler's results did not clarify whether miR-132 and miR-212 are involved in regulating steroidogenesis (or any other function) in granulosa cells.…”

Hormonal Regulation of MicroRNA Expression in Periovulatory Mouse Mural Granulosa Cells1

“…Interestingly, CTBP1 was recently shown to interact with steroidogenic factor-1 [46] in adrenal cells to modulate its ability to stimulate promoter activity. Our studies indicate that CTBP1 is posttranscriptionally regulated, as evidenced by the lack of change in its mRNA levels in response to in vivo hCG treatment combined with the pronounced loss in the protein when miRNA function-blocking LNA oligonucleotides were administered.…”

Effects of monocrotophos pesticide on steroidogenesis and transcription of steroidogenic enzymes in rainbow trout RTG-2 cells involving the protein kinase A signal pathway

“…These results are consistent with previous findings that showed transcriptional up-regulation of the cyp11a1, cyp17, and cyp19a genes via the cAMP/PKA pathway in different organisms. Thus, increased cyp11a1 and cyp17 mRNA levels were induced by cAMP in mouse Leydig cells (Payne and Sha, 1991), whereas cyp17 gene expression was stimulated in H295R adrenocortical cells by PKA via phosphorylation of carboxyl-terminal binding protein 1 (Dammer and Sewer, 2008). In fishes, a CRE has been identified in the promoter of the cyp19a gene in several teleost species, including rainbow trout (Tanaka et al, 1992), zebrafish (Kazeto et al, 2001), and gilthead seabream (Sparus aurata) (Wong et al, 2006), indicating PKA-mediated transcriptional regulation via phosphorylated CREB.…”