Insulin suppresses transactivation by CAAT/enhancer-binding proteins beta (C/EBP beta) - Signaling to p300/CREB-binding protein by protein kinase B disrupts interaction with the major activation domain of C/EBP beta

CAAT/enhancer-binding proteins (C/EBPs) play an important role in the regul ation of gene expression in insulin-responsive tissues. We have found that a complex containing C/EBP beta interacts with an insulin response sequence in the insulin-like growth factor-binding protein-1 (IGFBP-1) gene and tha t a C/EBP-binding site can mediate effects of insulin on promoter activity. Here, we examined mechanisms mediating this effect of insulin. The ability of insulin to suppress promoter activity via a C/EBP-binding site is block ed by LY294002, a phosphatidylinositol 3-kinase inhibitor, but not by rapam ycin, which blocks activation of p70(S6 kinase). Dominant negative phosphat idylinositol 3-kinase and protein kinase B (PKB) block the effect of insuli n, while activated PKB suppresses promoter function via a C/EBP-binding sit e, mimicking the effect of insulin, Coexpression studies indicate that insu lin and PKB suppress transactivation by C/EBP beta, but not C/EBP alpha, an d that N-terminal transactivation domains in C/EBP beta are required. Studi es with Gal4 fusion proteins reveal that insulin and PKB suppress transacti vation by the major activation domain in C/EBP beta (AD II), located betwee n amino acids 31 and 83, Studies with E1A protein indicate that interaction with p300/CBP is required for transactivation by AD II and the effect of i nsulin and PKB, Based on a consensus sequence, we identified a PKB phosphor ylation site (Ser(1834)) within the region of p300/CBP known to bind C/EBP beta, Mammalian two-hybrid studies indicate that insulin and PKB disrupt in teractions between this region of p300 and AD II and that Ser(1834) is crit ical for this effect. Signaling by PKB and phosphorylation of Ser(1834) may play an important role in modulating interactions between p300/CBP and tra nscription factors and mediate effects of insulin and related growth factor s on gene expression.