Phosphorylation of insulin receptor substrate-1 on multiple serine residues, 612, 632, 662, and 731, modulates insulin action

J Biol Chem. 1996 May 10;271(19):11222-7. doi: 10.1074/jbc.271.19.11222.


Okadaic acid has been described previously as being a negative regulator of insulin signaling, as it inhibits insulin stimulation of glucose transport. In addition, this drug induces on insulin receptor substrate-1 (IRS-1) a decrease in tyrosine phosphorylation, concomitantly with an increase in serine/threonine phosphorylation. The present work was aimed at the identification of the serine/threonine residues that, upon phosphorylation, might be involved in modulating insulin signaling. To this end, we studied double-point mutants of IRS-1, in which serines 612/632 and 662/731 were replaced with alanine. These are four plausible sites of phosphorylation by mitogen-activated protein kinases and are in the immediate proximity of tyrosine residues, which are potential sites of interaction with phosphatidylinositol 3-kinase Src homology 2 domains. Using transient expression in 293 EBNA cells, we demonstrate that serines 612, 632, 662, and 731 and mitogen-activated protein kinases are not involved in the okadaic acid effect on IRS-1. Rather, these serines appear to play a role in modulating basal and insulin-stimulated IRS-1 tyrosine phosphorylation, association of IRS-1, with p85, and phosphatidylinositol 3-kinase activity in the IRS-1.p85 immune complex, since mutation of these sites enhances these events. Our findings suggest the existence of an IRS-1 desensitization mechanism resulting from serine/threonine phosphorylation, occurring at least on serines 612, 632, 662, and 731.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Calcium-Calmodulin-Dependent Protein Kinases / antagonists & inhibitors
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • Cell Line
  • Cloning, Molecular
  • Enzyme Inhibitors / pharmacology
  • Ethers, Cyclic / pharmacology
  • Flavonoids / pharmacology
  • Humans
  • Insulin / pharmacology*
  • Insulin Receptor Substrate Proteins
  • Molecular Sequence Data
  • Okadaic Acid
  • Phosphatidylinositol 3-Kinases
  • Phosphoproteins / chemistry
  • Phosphoproteins / isolation & purification
  • Phosphoproteins / metabolism*
  • Phosphorylation
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Point Mutation
  • Rats
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Serine*
  • Threonine
  • Transfection


  • Enzyme Inhibitors
  • Ethers, Cyclic
  • Flavonoids
  • IRS1 protein, human
  • Insulin
  • Insulin Receptor Substrate Proteins
  • Irs1 protein, rat
  • Phosphoproteins
  • Recombinant Proteins
  • Okadaic Acid
  • Threonine
  • Serine
  • Phosphatidylinositol 3-Kinases
  • Phosphotransferases (Alcohol Group Acceptor)
  • Calcium-Calmodulin-Dependent Protein Kinases
  • 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one