A role for protein phosphatase 2A-like activity, but not atypical protein kinase C zeta, in the inhibition of protein kinase B/Akt and glycogen synthesis by palmitate

We have shown previously that palmitate treatment of C2C12 skeletal muscle myotubes causes inhibition of the protein kinase B (PKB) pathway and hence reduces insulin-stimulated glycogen synthesis through the elevation of intr acellular ceramide levels. Ceramide is known to activate both atypical prot ein kinase C (aPKC) zeta and protein phosphatase (PP) 2A, and each of these effectors has been reported to inhibit PKB. In the present study, palmitat e pretreatment was found to elevate PP2A-like activity in myotubes and to p revent its inhibition by insulin. Incubation with the phosphatase inhibitor okadaic acid before insulin stimulation protected against the effect of th e fatty acid on PKB phosphorylation. Palmitate was unable to inhibit PKB ac tivity and glycogen synthesis in cells overexpressing the activated PKB mut ant (T308D,S473D)-PKB alpha, which is unaffected by phosphatase. In contras t, PKB activity and glycogen synthesis were still inhibited by palmitate in cells overexpressing a membrane-targeted and, hence, activated PKB mutant that retains sensitivity to phosphatase. Although aPKC activity was also in creased in palmitate-treated cells, overexpression of wild-type or kinase-d ead aPKQ did not alter the inhibitory effects of the lipid on either stimul ation of PKB or glycogen synthesis by insulin. We conclude that palmitate d isrupts insulin signaling in C2C12 myotubes by promoting PP2A-like activity and, therefore, the dephosphorylation of PKB, which in turn reduces the st imulation of glycogen synthesis.