Inhibition of the Ca2+/calmodulin-dependent protein kinase I cascade by cAMP-dependent protein kinase

Several recent studies have shown that Ca2+/calmodulin-dependent protein ki nase I (CaMKI) is phosphorylated and activated by a protein kinase (CaMKK) that is itself subject to regulation by Ca2+/calmodulin. In the present stu dy, we demonstrate that this enzyme cascade is regulated by cAMP-mediated a ctivation of cAMP-dependent protein kinase (PKA), In vitro, CaMKK is phosph orylated by PKA and this is associated with inhibition of enzyme activity. The major site of phosphorylation is threonine 108, although additional sit es are phosphorylated with lower efficiency. In vitro, CaMKK is also phosph orylated by CaMKI at the same sites as PKA, suggesting that this regulatory phosphorylation might play a role as a negative-feedback mechanism, In int act PC12 cells, activation of PKA with forskolin resulted in a rapid inhibi tion of both CaMKK and CaMKI activity. In hippocampal slices CaMKK was phos phorylated under basal conditions, and activation of PKA led to an increase in phosphorylation. Two-dimensional phosphopeptide mapping indicated that activation of PKA led to increased phosphorylation of multiple sites includ ing threonine 108. These results indicate that in vitro and in intact cells the CaMKK/CaMKI cascade is subject to inhibition by PKA-mediated phosphory lation of CaMKK. The phosphorylation and inhibition of CaMKK by PKA is like ly to be involved in modulating the balance between cAMP- and Ca2+-dependen t signal transduction pathways.