Interaction of protein kinase C and cAMP-dependent pathways in the phosphorylation of the Na,K-ATPase

To test the hypothesis that there is cross-talk between the protein kinase C (PKC) and protein kinase A (PKA) pathways in the regulation of the Na,K-A TPase, we measured its phosphorylation in mammalian cell cultures. Phosphor ylation of the PKC site, Ser-18, appeared to be due to the activation of th e a isoform of the kinase. In NRK-52E and L6 cells, this phosphorylation wa s reduced by prior activation of a cAMP-dependent signaling pathway with fo rskolin. In principle this would be consistent with direct interaction betw een the two phosphorylation sites, but further investigation suggested a mo re indirect mechanism. First, phosphorylation of Ser-938, the PKA site, cou ld not be detected despite the presence of active PKA. Second, there was a major reduction in the phosphorylation of unrelated phosphoproteins as a co nsequence of elevation of cAMP, suggesting generalized reduction of kinase activity or activation of phosphatase activity. In NRK-52E and L6, phosphor ylation of the Na,K-ATPase at Ser-18 paralleled this global change. In C6 c ells, in contrast, there was no cAMP effect on Na,K-ATPase phosphorylation at Ser-18 and no global cAMP effect on other phosphoproteins. The cross-tal k is evidently mediated by events occurring at the cellular level.