CaMKKα activates AMPK signaling in mouse skeletal muscle in vivo

Abstract

AMP-activated protein kinase (AMPK) is activated by exercise/skeletal muscle contraction, and is a regulator of numerous cellular processes. AMPK is regulated by changes in cellular energy status, however recent studies in cultured cells have suggested that AMPK may also be regulated by Ca2+/calmodulin-dependent protein kinase kinases (CaMKKs), primarily CaMKKβ. We determined whether CaMKKα, the only CaMKK isoform known to be expressed in muscle, activates AMPK signaling in skeletal muscle in vivo. Plasmids containing constitutively active CaMKKα (caCaMKKα) or empty vector were transfected into mouse tibialis anterior muscles by in vivo electroporation. After 2 wks, caCaMKKα protein levels were increased 10-fold over endogenous CaMKKα. caCaMKKα expression resulted in ~2-fold increases in the phosphorylation of CaMKI (Thr177), a CaMKK substrate. To determine whether CaMKKα activates AMPK, we assessed AMPK phosphorylation and activity. caCaMKKα increased AMPK (Thr172) phosphorylation and AMPK α1 and α2 activities by ~2-fold, with no change in the protein levels of AMPK or the AMPK kinase, LKB1. Phosphorylation of the AMPK substrate, acetyl-CoA-carboxylase (ACC, Ser79), was also increased ~2-fold, with no change in ACC protein levels. Neither Akt nor ERK phosphorylation was changed, suggesting that the effects of caCaMKKα expression were specific to AMPK signaling. In conclusion, we demonstrated that the predominant CaMKK isoform in muscle, CaMKKα, can activate AMPK signaling in vivo. These results suggest a novel Ca2+-dependent mechanism for the in vivo regulation of AMPK in skeletal muscle. SUPPORT: R01 AR42238, F32 AR051663.