title: Phosphorylation of insulin receptor substrate-1 (IRS-1) by protein kinase B positively regulates IRS-1 function.
abstract: Incubation of cells with insulin leads to a transient rise in Tyr phosphorylation of insulin receptor substrate (IRS) proteins, accompanied by elevation in their Ser(P)/Thr(P) content and their dissociation from the insulin receptor (IR). Wortmannin, a phosphatidylinositol 3-kinase inhibitor, selectively prevented the increase in Ser(P)/Thr(P) content of IRS-1, its dissociation from IR, and the decrease in its Tyr(P) content following 60 min of insulin treatment. Four conserved phosphorylation sites within the phosphotyrosine binding/SAIN domains of IRS-1 and IRS-2 served as in vitro substrates for protein kinase B (PKB), a Ser/Thr kinase downstream of phosphatidylinositol 3-kinase. Furthermore, PKB and IRS-1 formed stable complexes in vivo, and overexpression of PKB enhanced Ser phosphorylation of IRS-1. Overexpression of PKB did not affect the acute Tyr phosphorylation of IRS-1; however, it significantly attenuated its rate of Tyr dephosphorylation following 60 min of treatment with insulin. Accordingly, overexpression of IRS-1(4A), lacking the four potential PKB phosphorylation sites, markedly enhanced the rate of Tyr dephosphorylation of IRS-1, while inclusion of vanadate reversed this effect. These results implicate a wortmannin-sensitive Ser/Thr kinase, different from PKB, as the kinase that phosphorylates IRS-1 and acts as the feedback control regulator that turns off insulin signals by inducting the dissociation of IRS proteins from IR. In contrast, insulin-stimulated PKB-mediated phosphorylation of Ser residues within the phosphotyrosine binding/SAIN domain of IRS-1 protects IRS-1 from the rapid action of protein-tyrosine phosphatases and enables it to maintain its Tyr-phosphorylated active conformation. These findings implicate PKB as a positive regulator of IRS-1 functions.
authors.affiliation: Department of Molecular Cell Biology, the Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: K
authors.initials: K
authors.lastname: Paz
authors.forename: Y F
authors.initials: YF
authors.lastname: Liu
authors.forename: H
authors.initials: H
authors.lastname: Shorer
authors.forename: R
authors.initials: R
authors.lastname: Hemi
authors.forename: D
authors.initials: D
authors.lastname: LeRoith
authors.forename: M
authors.initials: M
authors.lastname: Quan
authors.forename: H
authors.initials: H
authors.lastname: Kanety
authors.forename: R
authors.initials: R
authors.lastname: Seger
authors.forename: Y
authors.initials: Y
authors.lastname: Zick
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D018076:DNA, Complementary; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D014638:Vanadates / Q000494:pharmacology; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; C526292:IRS1 protein, human; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D014638:Vanadates; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D000077191:Wortmannin