Tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) by oxidant stress in cerebellar granule neurons: modulation by N-methyl-D-aspartate through calcineurin activity.
Insulin receptor-substrate-1 (IRS-1) is a docking protein for several tyrosine kinase receptors. Upon tyrosine phosphorylation, IRS-1 binds to signaling molecules that express Src homology 2 (SH-2) binding domains, including phosphatidylinositol 3-kinase (PI 3-kinase), phosphotyrosine phosphatase SHP-2 (Syp), Nck, Crk and Grb-2. Hydrogen peroxide (H(2)O(2)) induces tyrosine phosphorylation of key signaling mediators presumably by inhibition of tyrosine phosphatases. In many cell types, the activation of extracellular signal-related kinases (e.g. MAPK) and other protein kinases by H(2)O(2) leads to transcriptional activation. In the current study, we examined the effect of H(2)O(2) on IRS-1 tyrosine phosphorylation in primary cultured rat cerebellar granule neurons. H(2)O(2) stimulated the rapid tyrosine phosphorylation of IRS-1 and p42/p44 MAP kinase, and induced its association with PI 3-kinase. H(2)O(2)-induced IRS-1 phosphorylation was rapidly reversible (5 min) whereas MAPK phosphorylation persisted for up to 1 h. NMDA reversed H(2)O(2)-mediated tyrosine phosphorylation of IRS-1 and its association with PI 3-kinase. The dephosphorylation of IRS-1 by NMDA was calcium-dependent and was inhibited by the calcineurin inhibitor cyclosporine. Calmodulin-dependent tyrosine phosphatase activity of calcineurin was observed in vitro using both immunoprecipitated and recombinant tyrosine-phosphorylated IRS-1 as substrates. These data highlight the role of multiple phosphatases in the regulation of IRS-1 tyrosine phosphorylation and identify a novel functional property of calcineurin.
Department of Pathology, Anatomy and Cell Biology, Jefferson Medical College, Philadelphia 19107, USA.
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Hallak
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Ramadan
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Rubin
D000818:Animals; D019703:Calcineurin / Q000378:metabolism*; D065095:Calcineurin Inhibitors; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016202:N-Methylaspartate / Q000378:metabolism* / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects; D014443:Tyrosine / Q000378:metabolism
D065095:Calcineurin Inhibitors; D004791:Enzyme Inhibitors; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D016877:Oxidants; D010750:Phosphoproteins; D014443:Tyrosine; D016202:N-Methylaspartate; D006861:Hydrogen Peroxide; D019869:Phosphatidylinositol 3-Kinases; D020928:Mitogen-Activated Protein Kinases; C120817:calcineurin phosphatase; D019703:Calcineurin; D010744:Phosphoric Monoester Hydrolases; D002118:Calcium