Ca2+/calmodulin-dependent protein kinase kinase-β acts upstream of AMP-activated protein kinase in mammalian cells

AMP-activated protein kinase (AMPK) is the downstream component of a kinase cascade that plays a pivotal role in energy homeostasis. Activation of AMPK requires phosphorylation of threonine 172 (T172) within the T loop region of the catalytic a subunit. Recently, LKB1 was shown to activate AMPK. Here we show that AMPK is also activated by Ca2+/calmodulin-dependent protein kinase kinase (CaMKK). Overexpression of CaMKK beta in mammalian cells increases AMPK activity, whereas pharmacological inhibition of CaMKK, or downregulation of CaMKK beta using RNA interference, almost completely abolishes AMPK activation. CaMKK beta isolated from rat brain or expressed in E. coli phosphorylates and activates AMPK in vitro. In yeast, CaMKK beta expression rescues a mutant strain lacking the three kinases upstream of Snf1, the yeast homolog of AMPK. These results demonstrate that AMPK is regulated by at least two upstream kinases and suggest that AMPK may play a role in Ca2+-mediated signal transduction pathways.