Greater insulin-mediated Akt phosphorylation concomitant with heterogeneous effects on phosphorylation of Akt substrates in soleus of calorie-restricted rats

Akt is a serine/threonine kinase that plays a key role in numerous cellular functions including metabolism, growth, protein synthesis, apoptosis, and cell proliferation. The most consistent and robust effect of moderate calorie restriction (CR; ~60% of ad libitum, AL, food consumption) on insulin signaling in rodent muscle has been enhanced insulin-induced phosphorylation of Akt (pAkt). However, there is limited knowledge regarding the mechanism for this enhancement and its consequences in predominantly slow-twitch muscle. Accordingly, in soleus muscle of 9-mo-old rats, we analyzed the effect of CR and insulin on important signaling events that are proximal to Akt activation including: pIR^Tyr1162/1163, pIRS1^Tyr, pIRS1^Ser312, IRS1-associated phosphatidylinositol 3-kinase activity, or pPTEN^Ser380. In addition, we analyzed the effect of CR and insulin on Akt substrates that have established or putative roles in glucose metabolism, cellular growth, maintenance of muscle structure, or protein synthesis including pGSK3α^Ser21, pGSK3β^Ser9, pTSC2^Ser939, pP70S6K^Thr412, pAS160^Thr642, and pFLNc^Ser2213. The current study demonstrated that the CR-induced increase in pAkt in isolated soleus muscles from 9-mo-old rats can occur without concomitant enhancement of several important insulin signaling events that are proximal to Akt activation. These results suggest that the greater pAkt in the soleus muscles from CR rats was attributable to an alternative mechanism. We also observed that the effects of CR were not uniform for phosphorylation of six insulin-regulated Akt substrates in the soleus. The differential response in phosphorylation by Akt substrates likely has important implications for explaining the complex effect of CR diverse cellular functions.