Insulin stimulates phosphorylation of the forkhead transcription factor FKHR on serine 253 through a Wortmannin-sensitive pathway.
In the nematode Caenorhabditis elegans, mutations of the insulin/insulin-like growth factor-1 receptor homologue Daf-2 gene cause developmental arrest at the dauer stage. The effect of Daf-2 mutations is counteracted by mutations in the Daf-16 gene, suggesting that Daf-16 is required for signaling by Daf-2. Daf-16 encodes a forkhead transcription factor. Based on sequence similarity, the FKHR genes are the likeliest mammalian Daf-16 homologues. FKHR proteins contain potential sites for phosphorylation by the serine/threonine kinase Akt. Because Akt is phosphorylated in response to insulin and has been implicated in a variety of insulin effects, we investigated whether insulin affects phosphorylation of FKHR. Insulin stimulated phosphorylation of endogenous FKHR and of a recombinant c-Myc/FKHR fusion protein transiently expressed in murine SV40-transformed hepatocytes. The effect of insulin was inhibited by wortmannin treatment, suggesting that PI 3-kinase activity is required for FKHR phosphorylation. Mutation of serine 253, located in a consensus Akt phosphorylation site at the carboxyl-terminal end of the forkhead domain, abolished the effect of insulin on FKHR phosphorylation. In contrast, mutation of two additional Akt phosphorylation sites, at amino acids threonine 24 or serine 316, did not abolish insulin-induced phosphorylation. These data indicate that FKHR may represent a distal effector of insulin action.
Nakae
J
J
Developmental Endocrinology Branch, NICHD, National Institutes of Health, Bethesda, Maryland 20892, USA.
Park
B C
BC
Accili
D
D
D019943:Amino Acid Substitution; D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D001798:Blood Proteins / Q000378:metabolism*; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D016384:Consensus Sequence; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D051858:Forkhead Transcription Factors; D005786:Gene Expression Regulation; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine / Q000378:metabolism; D013539:Simian virus 40; D013329:Structure-Activity Relationship; D013912:Threonine / Q000378:metabolism; D014157:Transcription Factors; D000077191:Wortmannin
D000730:Androstadienes; D001798:Blood Proteins; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D051858:Forkhead Transcription Factors; D007328:Insulin; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine; D013912:Threonine; D014157:Transcription Factors; D000077191:Wortmannin; C108803:FOXO4 protein, human; C494918:AKT1 protein, human