Abstract
Protein kinase B lies "downstream" of phosphatidylinositide (PtdIns) 3-kinase and is thought to mediate many of the intracellular actions of insulin and other growth factors. Here we show that FKHR, a human homologue of the DAF16 transcription factor in Caenorhabditis elegans, is rapidly phosphorylated by human protein kinase Balpha (PKBalpha) at Thr-24, Ser-256, and Ser-319 in vitro and at a much faster rate than BAD, which is thought to be a physiological substrate for PKB. The same three sites, which all lie in the canonical PKB consensus sequences (Arg-Xaa-Arg-Xaa-Xaa-(Ser/Thr)), became phosphorylated when FKHR was cotransfected with either PKB or PDK1 (an upstream activator of PKB). All three residues became phosphorylated when 293 cells were stimulated with insulin-like growth factor 1 (IGF-1). The IGF-1-induced phosphorylation was abolished by the PtdIns 3-kinase inhibitor wortmannin but not by PD 98059 (an inhibitor of the mitogen-activated protein kinase cascade) or by rapamycin. These results indicate that FKHR is a physiological substrate of PKB and that it may mediate some of the physiological effects of PKB on gene expression. DAF16 is known to be a component of a signaling pathway that has been partially dissected genetically and includes homologues of the insulin/IGF-1 receptor, PtdIns 3-kinase and PKB. The conservation of Thr-24, Ser-256, and Ser-319 and the sequences surrounding them in DAF16 therefore suggests that DAF16 is also a direct substrate for PKB in C. elegans.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3-Phosphoinositide-Dependent Protein Kinases
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Amino Acid Sequence
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Androstadienes / pharmacology
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Antibody Specificity
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Caenorhabditis elegans Proteins
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Carrier Proteins / metabolism
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Consensus Sequence
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / immunology
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DNA-Binding Proteins / metabolism*
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Enzyme Activation
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Forkhead Box Protein O1
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Forkhead Transcription Factors
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Humans
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Insulin-Like Growth Factor I / pharmacology
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Molecular Sequence Data
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Phosphoinositide-3 Kinase Inhibitors
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Phosphorylation
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Phosphoserine / immunology
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Phosphothreonine / immunology
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Protein Processing, Post-Translational
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism*
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Proto-Oncogene Proteins c-akt
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Proto-Oncogene Proteins*
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Recombinant Proteins / metabolism
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Serine / metabolism
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Signal Transduction
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Sirolimus / pharmacology
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Threonine / metabolism
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Transcription Factors / genetics
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Transcription Factors / immunology
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Transcription Factors / metabolism*
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Wortmannin
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bcl-Associated Death Protein
Substances
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Androstadienes
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BAD protein, human
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Caenorhabditis elegans Proteins
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Carrier Proteins
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DNA-Binding Proteins
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FOXO1 protein, human
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Forkhead Box Protein O1
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Forkhead Transcription Factors
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Phosphoinositide-3 Kinase Inhibitors
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Proto-Oncogene Proteins
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Recombinant Proteins
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Transcription Factors
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bcl-Associated Death Protein
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Phosphothreonine
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Phosphoserine
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Threonine
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Serine
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Insulin-Like Growth Factor I
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3-Phosphoinositide-Dependent Protein Kinases
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PDPK1 protein, human
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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akt-1 protein, C elegans
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Sirolimus
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Wortmannin