Role of the atypical protein kinase Cζ in regulation of 5′-AMP-activated protein kinase in cardiac and skeletal muscle

Ussher JR, Jaswal JS, Wagg CS, Armstrong HE, Lopaschuk DG, Keung W, Lopaschuk GD. Role of the atypical protein kinase C zeta in regulation of 5'-AMP-activated protein kinase in cardiac and skeletal muscle. Am J Physiol Endocrinol Metab 297: E349-E357, 2009; doi: 10.1152/ajpendo.00009.2009.-During metabolic stress, phosphorylation and activation of 5'-AMP-activated protein kinase (AMPK) becomes a major regulator of cellular energy metabolism in heart and skeletal muscle. Despite this, the upstream regulation of AMPK in both heart and muscle is poorly understood. Recent work has implicated the atypical protein kinase C zeta (PKC zeta) as a regulator of AMPK in endothelial cells via phosphorylation of LKB1, an upstream AMPK kinase (AMPKK). Our goal was to determine the potential role PKC zeta plays in regulating AMPK in cardiac and skeletal muscle. Cultures of H9c2 myocytes (cardiac) and C2C12 myotubes (skeletal muscle) were pretreated with a selective PKC zeta pseudosubstrate peptide inhibitor and treated with various AMPK activating agents to determine whether PKC zeta regulates AMPK. PKC zeta activity was also examined in isolated working rat hearts subjected to ischemia. We show that PKC zeta is not involved in regulating threonine 172 AMPK phosphorylation induced by metformin or phenformin in either cardiac or skeletal muscle cells but is involved in 5-aminoimidazole-4-carboxamine-1-beta-D-ribofuranoside (AICAR)-induced AMPK phosphorylation in cardiac muscle cells. Activation of PKC zeta with high palmitate concentrations is also insufficient to increase AMPK phosphorylation. Furthermore, we show that the ischemia-induced activation of AMPK is not accompanied by increased PKC zeta activity. Finally, we show that PKC zeta may actually be a downstream target of AMPK in skeletal muscle, since adenoviral expression of a dominant-negative mutant of AMPK prevented metformin-and AICAR-induced phosphorylation of PKC zeta. We conclude that PKC zeta plays a very minor role in the regulation of AMPK in cardiac and skeletal muscle and may actually be a downstream target pre-treatedof AMPK in skeletal muscle.