Differential regulation of eEF2 and p70S6K by AMPKalpha2 in heart

Eukaryotic elongation factor 2 (eEF-2) and mammalian target of rapamycin (mTOR)-p70 ribosomal protein S6 kinase (p70S6K) signaling pathways control protein synthesis and are inhibited during myocardial ischemia, Intracellular acidosis and AMP-activated protein kinase (AMPK) activation, both occurring during ischemia, have been proposed to participate in this inhibition. We evaluated the contribution of AMPKa2, the main cardiac AMPK catalytic subunit isoform, in eEF2 and mTOR-p70S6K regulation using AMPK0t2 KO mice. Hearts were perfused ex vivo with or without insulin, and then submitted or not to ischemia. Insulin pre-incubation was necessary to activate mTOR-p70S6K and evaluate their subsequent inhibition by ischemia. Ischemia decreased insulininduced mTOR-p70S6K phosphorylation in WT and AMPKa2 KO mice to a similar extent. This AMPKa2independent p7056K inhibition correlated well with the inhibition of PKB/Alct, located upstream of mTORp7056K and can be mimicked in cardiomyocytes by decreasing pH. By contrast, ischemia-induced inhibitory phosphorylation of eEF-2 was drastically reduced in AMPKa2 KO mice. Interestingly, AMPKa2 also played a role under normoxia. Its deletion increased the insulin-induced p70S6K stimulation. This p7056K overstimulation was associated with a decrease in inhibitory phosphorylation of Raptor, an mTOR partner identified as an AMPK target. In conclusion, AMPKa2 controls cardiac p70S6K under normoxia and regulates eEF-2 but not the mTOR-p70S6K pathway during ischemia. This challenges the accepted notion that mTOR-p70S6K is inhibited by myocardial ischemia mainly via an AMPK-dependent mechanism. (C) 2013 Elsevier B.V. All rights reserved.