TNFα activation of PKCδ, mediated by NFκB and ER stress, cross-talks with the insulin signaling cascade

TNF alpha plays key roles in the regulation of inflammation, cell death, and proliferation and its signaling cascade cross-talks with the insulin signaling cascade. PKC delta, a novel PKC isoform, is known to participate in proximal TNF alpha signaling events. However, it has remained unclear whether PKC delta plays a role in distal TNF alpha signaling events. Here we demonstrate that PKCS is activated by TNF alpha in a delayed fashion that is temporally associated with JNK activation. To investigate the signaling pathways activating PKC delta and JNK we used pharmacological and genetic inhibitors of NF kappa B. We found that inhibition of NF kappa B attenuated PKC delta and JNK activations. Further analysis revealed that ER stress contributes to TNF alpha-stimulated PKC delta and JNK activations. To investigate the role of PKC delta in TNF alpha action. we used 29-mer shRNAs to silence PKC delta expression. A reduction of similar to 90% in PKC delta protein levels reduced TNF alpha-stimulated stress kinase activation, including JNK. Further, PKC delta was necessary for thapsigargin-stimulated JNK activation. Because thapsigargin is a potent inducer of ER stress, we determined whether PKC delta was necessary for induction of the UPR. Indeed, a reduction in PKC delta protein levels reduced thapsigargin-stimulated CHOP induction, a hallmark of the UPR. but not BiP/GRP78 induction, suggesting that PKC delta does not globally regulate the UPR. Next, the role of PKC delta in TNF alpha mediated cross-talk with the insulin signaling pathway was investigated in cells expressing human IRS-1 and a 29-mer shRNA to silence PKC delta expression. We found that a reduction in PKC delta protein levels reversed the TNF alpha-mediated reduction in insulin-stimulated IRS-1 Tyr phosphorylation, Akt activation, and glycogen synthesis. In addition. TNF alpha-stimulated IRS protein Ser/Thr phosphorylation and degradation were blocked. Our results indicate that: 1) NF kappa B and ER stress contribute in part to PKC delta activation; 2) PKC6 plays a key role in the propagation of the TNF alpha signal: and 3) PKC delta contributes to TNF alpha-induced inhibition of insulin signaling events. (C) 2009 Elsevier Inc. All rights reserved.