Protein kinase C-zeta phosphorylates insulin receptor substrate-1 and impairs its ability to activate phosphatidylinositol 3-kinase in response to insulin.
Protein kinase C-zeta (PKC-zeta) is a serine/threonine kinase downstream from phosphatidylinositol 3-kinase in insulin signaling pathways. However, specific substrates for PKC-zeta 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-zeta. Under in vitro conditions, wild-type PKC-zeta (but not kinase-deficient mutant PKC-zeta) significantly phosphorylated IRS-1. This phosphorylation was reversed by treatment with the serine-specific phosphatase, protein phosphatase 2A. In addition, the overexpression of PKC-zeta in NIH-3T3(IR) cells caused significant phosphorylation of cotransfected IRS-1 as demonstrated by [(32)P]orthophosphate labeling experiments. In rat adipose cells, endogenous IRS-1 coimmunoprecipitated with endogenous PKC-zeta, and this association was increased 2-fold upon insulin stimulation. Furthermore, the overexpression of PKC-zeta in NIH-3T3(IR) 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-zeta. Because PKC-zeta is located downstream from IRS-1 and phosphatidylinositol 3-kinase in established insulin signaling pathways, PKC-zeta may participate in negative feedback pathways to IRS-1 similar to those described previously for Akt and GSK-3.
Ravichandran
L V
LV
Cardiology Branch, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Esposito
D L
DL
Chen
J
J
Quon
M J
MJ
D016475:3T3 Cells; D000818:Animals; D004789:Enzyme Activation; D006801:Humans; D007328:Insulin / Q000502:physiology*; D055504:Insulin Receptor Substrate Proteins; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013379:Substrate Specificity; D014443:Tyrosine / Q000378:metabolism; D000099221:Protein Kinase C zeta
D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D011493:Protein Kinase C; D014443:Tyrosine; D000099221:Protein Kinase C zeta; C526292:IRS1 protein, human; C526295:Irs1 protein, mouse; C526296:Irs1 protein, rat