Coordinated phosphorylation of insulin receptor substrate-1 by glycogen synthase kinase-3 and protein kinase CβII in the diabetic fat tissue

Ziva Liberman, Batya Plotkin, Tamar Tennenbaum, Hagit Eldar-Finkelman

Research output: Contribution to journalArticlepeer-review

Abstract

Serine/threonine phosphorylation of insulin receptor substrate-1 (IRS-1) is an important negative modulator of insulin signaling. Previously, we showed that glycogen synthase kinase-3 (GSK-3) phosphorylates IRS-1 at Ser 332. However, the fact that GSK-3 requires prephosphorylation of its substrates suggested that Ser336 on IRS-1 was the "priming" site phosphorylated by an as yet unknown protein kinase. Here, we sought to identify this "priming kinase" and to examine the phosphorylation of IRS-1 at Ser336 and Ser332 in physiologically relevant animal models. Of several stimulators, only the PKC activator phorbol ester PMA enhanced IRS-1 phosphorylation at Ser336. Treatment with selective PKC inhibitors prevented this PMA effect and suggested that a conventional PKC was the priming kinase. Overexpression of PKCα or PKCβII isoforms in cells enhanced IRS-1 phosphorylation at Ser336 and Ser332, and in vitro kinase assays verified that these two kinases directly phosphorylated IRS-1 at Ser336. The expression level and activation state of PKCβII, but not PKCα, were remarkably elevated in the fat tissues of diabetic ob/ob mice and in high-fat diet-fed mice compared with that from lean animals. Elevated levels of PKCβII were also associated with enhanced phosphorylation of IRS-1 at Ser336/332 and elevated activity of GSK-3β. Finally, adenoviral mediated expression of PKCβII in adipocytes enhancedphosphorylation of IRS-1 at Ser336. Taken together, our results suggest that IRS-1 is sequentially phosphorylated by PKCβII and GSK-3 at Ser336 and Ser332. Furthermore, these data provide evidence for the physiological relevance of these phosphorylation events in the pathogenesis of insulin resistance in fat tissue.

Original languageEnglish
Pages (from-to)E1169-E1177
JournalAmerican Journal of Physiology - Endocrinology and Metabolism
Volume294
Issue number6
DOIs
StatePublished - Jun 2008

Keywords

  • Insulin resistance
  • Insulin signaling
  • Obesity

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