Protein Kinase C-ζ Phosphorylates Insulin Receptor Substrate-1 and Impairs Its Ability to Activate Phosphatidylinositol 3-Kinase in Response to Insulin

Abstract

Protein kinase C-(PKC-) is a serine/threonine kinase downstream from phosphatidylinositol 3-kinase in insulin signaling pathways. However, specific substrates for PKC-that participate in the biological actions of insulin have not been reported. In the present study, we identified insulin receptor substrate-1 (IRS-1) as a novel substrate for PKC-. Under in vitro conditions, wild-type PKC-(but not kinase-deficient mutant PKC-) significantly phosphorylated IRS-1. This phosphorylation was reversed by treatment with the serine-specific phosphatase, protein phosphatase 2A. In addition, the overexpression of PKC-in NIH-3T3 IR cells caused significant phosphorylation of cotransfected IRS-1 as demonstrated by [ P]orthophosphate labeling experiments. In rat adipose cells, endogenous IRS-1 coimmunoprecipitated with endogenous PKC-, and this association was increased 2-fold upon insulin stimulation. Furthermore, the overexpression of PKC-in NIH-3T3IR cells significantly impaired insulin-stimulated tyrosine phosphorylation of cotransfected IRS-1. Importantly, this was accompanied by impaired IRS-1-associated phosphatidylinositol 3-kinase activity. Taken together, our results raise the possibility that IRS-1 is a novel physiological substrate for PKC-. Because PKC-is located downstream from IRS-1 and phosphatidylinositol 3-kinase in established insulin signaling pathways, PKC-may participate in negative feedback pathways to IRS-1 similar to those described previously for Akt and GSK-3.

Pleiotropic effects of fatty acids on pancreatic β‐cells

“…Expression of PKCz seems to be regulated by LC-CoA in insulin secretion stimulated by palmitate (Yaney et al, 2000). The isoform PKCz is also activated by insulin and mediates the phosphorylation of IRS proteins (Liu et al, 2001;Ravichandran et al, 2001). As a consequence the IR-IRS complexe dissociates, inhibiting the ability of IRS proteins to induce tyrosine phosphorylation (Liu et al, 2001).…”

Identification of an in vitro insulin receptor substrate-1 phosphorylation site by negative-ion μLC/ES-API-CID-MS hybrid scan technique

“…Insulin receptor substrate-1 (IRS-1), one major intracellular substrate of the insulin receptor kinase, interacts with the insulin receptor at sites adjacent to the N-terminus. Recently it was found that during insulin signaling IRS-1 is phosphorylated by protein kinase C (PKC)-, thereby attenuating the insulin signal transduction [2,3]. To disclose these phosphorylation sites, we developed a mass spectrometric strategy combining two independent MS methods.…”

Activation of opioid μ-receptors by loperamide to improve interleukin-6-induced inhibition of insulin signals in myoblast C2C12 cells

“…In fact, atypical PKCs are essential for insulin stimulation of glucose transport in muscle tissue [41]. Although atypical PKCs are located downstream of IRS-1 and PI3-kinase in insulin signalling, IRS-1 is also a novel substrate for atypical PKCs [42]. Moreover, suppressors of cytokine signalling (SOCS) proteins have been implicated in the insulin signalling network [43].…”