Serine phosphorylation sites on IRS2 activated by angiotensin II and protein kinase C to induce selective insulin resistance in endothelial cells

Mol Cell Biol. 2013 Aug;33(16):3227-41. doi: 10.1128/MCB.00506-13. Epub 2013 Jun 17.

Abstract

Protein kinase C (PKC) activation, induced by hyperglycemia and angiotensin II (AngII), inhibited insulin-induced phosphorylation of Akt/endothelial nitric oxide (eNOS) by decreasing tyrosine phosphorylation of IRS2 (p-Tyr-IRS2) in endothelial cells. PKC activation by phorbol ester (phorbol myristate acetate [PMA]) reduced insulin-induced p-Tyr-IRS2 by 46% ± 13% and, similarly, phosphorylation of Akt/eNOS. Site-specific mutational analysis showed that PMA increased serine phosphorylation at three sites on IRS2 (positions 303, 343, and 675), which affected insulin-induced tyrosine phosphorylation of IRS2 at positions 653, 671, and 911 (p-Tyr-IRS2) and p-Akt/eNOS. Specific PKCβ2 activation decreased p-Tyr-IRS2 and increased the phosphorylation of two serines (Ser303 and Ser675) on IRS2 that were confirmed in cells overexpressing single point mutants of IRS2 (S303A or S675A) containing a PKCβ2-dominant negative or selective PKCβ inhibitor. AngII induced phosphorylation only on Ser303 of IRS2 and inhibited insulin-induced p-Tyr911 of IRS2 and p-Akt/eNOS, which were blocked by an antagonist of AngII receptor I, losartan, or overexpression of single mutant S303A of IRS2. Increases in p-Ser303 and p-Ser675 and decreases in p-Tyr911 of IRS2 were observed in vessels of insulin-resistant Zucker fatty rats versus lean rats. Thus, AngII or PKCβ activation can phosphorylate Ser303 and Ser675 in IRS2 to inhibit insulin-induced p-Tyr911 and its anti-atherogenic actions (p-Akt/eNOS) in endothelial cells.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Angiotensin II / metabolism*
  • Animals
  • Cattle
  • Cell Line
  • Endothelial Cells / metabolism*
  • Enzyme Activation
  • Insulin / metabolism
  • Insulin Receptor Substrate Proteins / chemistry
  • Insulin Receptor Substrate Proteins / metabolism*
  • Insulin Resistance*
  • Male
  • Mice
  • Mice, Transgenic
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphorylation
  • Protein Kinase C / genetics
  • Protein Kinase C / metabolism*
  • Protein Kinase C beta
  • Rats
  • Rats, Zucker
  • Serine / chemistry
  • Serine / metabolism
  • Tetradecanoylphorbol Acetate / metabolism
  • Threonine / chemistry
  • Threonine / metabolism
  • Tyrosine / chemistry
  • Tyrosine / metabolism

Substances

  • Insulin
  • Insulin Receptor Substrate Proteins
  • Irs2 protein, mouse
  • Irs2 protein, rat
  • Angiotensin II
  • Threonine
  • Tyrosine
  • Serine
  • Phosphatidylinositol 3-Kinases
  • Protein Kinase C
  • Protein Kinase C beta
  • Tetradecanoylphorbol Acetate