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

Abstract

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.

Relationships between superoxide levels and delayed calcium deregulation in cultured cerebellar granule cells exposed continuously to glutamate

“…However, direct evidence for a mitochondrial permeability transition in glutamate-exposed neurons is equivocal. Cyclosporin A is problematic as a permeability transition diagnostic with intact cells due to its ability to inhibit calcineurin and hyperphosphorylate proteins (Hallak et al 2001). The more selective methyl-4-valine cyclosporin did not afford convincing protection in our hands (Castilho et al 1999) although others report partial protection against deregulation (Khaspekov et al 1999;Vergun et al 1999;Alano et al 2002).…”

Short Communication: Angiotensin II Type 1 Receptor–Mediated Upregulation of Calcineurin Activity Underlies Impairment of Cardioprotective Signaling in Diabetic Hearts

“…10 The hypothesis that calcineurin is responsible for lack of EPO-induced phosphorylation of Jak2 in OLETF rats was based on a report of upregulated calcineurin in the diabetic kidney 11 and a reported function of calcineurin as a Tyr protein phosphatase. 12 This hypothesis was supported by the findings that calcineurin activity was elevated by more than 2-fold in the myocardium of OLETF rats and that inhibition of calcineurin restored the response of Jak2 to EPO receptor activation ( Figure 3). Furthermore, blockade of the AT 1 receptor normalized calcineurin activity, indicating that AT 1 receptor mediates upregulation of calcineurin in OLETF rats.…”

Demonstrated brain insulin resistance in Alzheimer’s disease patients is associated with IGF-1 resistance, IRS-1 dysregulation, and cognitive decline

“…No alterations were found in levels of protein phosphatases (i.e., PP2A, PP2B, and PTP1B) known to act on the IR, IRS-1, and Akt (89)(90)(91)(92).…”