Phosphorylation of serine 256 by protein kinase B disrupts transactivationby FKHR and mediates effects of insulin on insulin-like growth factor-binding protein-1 promoter activity through a conserved insulin response sequence

Insulin inhibits the expression of multiple genes in the liver containing a n insulin response sequence (IRS) (CAAAA(C/T)AA), and we have reported that protein kinase B (PKB) mediates this effect of insulin, Genetic studies in Caenorhabditis elegans indicate that daf-16, a forkhead/winged-helix trans cription factor, is a major target of the insulin receptor-PKB signaling pa thway. FKHR, a human homologue of daf-16, contains three PKB sites and is e xpressed in the liver. Reporter gene studies in HepG2 hepatoma cells show t hat FKHR stimulates insulin-like growth factor-binding protein-1 promoter a ctivity through an IRS, and introduction of IRSs confers this effect on a h eterologous promoter. Insulin disrupts IRS-dependent transactivation by FKH R, and phosphorylation of Ser-256 by PKB is necessary and sufficient to med iate this effect. Antisense studies indicate that FKHR contributes to basal promoter function and is required to mediate effects of insulin and PKB on promoter activity via an IRS, To our knowledge, these results provide the first report that FKHR stimulates promoter activity through an IRS and that phosphorylation of FKHR by PKB mediates effects of insulin on gene express ion. Signaling to FKHR-related forkhead proteins via PKB may provide an evo lutionarily conserved mechanism by which insulin and related factors regula te gene expression.