Phosphorylation of endogenous proteins by adenosine 3':5'-monophosphate-dependent protein kinase in mouse neuroblastoma cells.

Increased intracellular adenosine 3':5'-monophosphate (cAMP) levels and activation of cAMP-dependent protein kinases (ATP:protein phosphotransferase, EC 2.7.1.37) in vivo were correlated in mouse neuroblastoma cells grown in the presence of 1 mM-6N,O2'-dibutyryl 3':5'-monophosphate (Bt2cAMP). The time course for activation showed that cAMP-dependent protein kinases were activated by 30 min. A heat-stable inhibitor protein inhibited a majority of activated cAMP-dependent protein kinase. Activation of cAMP-dependent protein kinase caused additional phosphorylation of proteins when compared with untreated control cells, as demonstrated by endogenous phosphorylation of proteins in vitro using [gamma-32P]ATP and analysis by two-dimensional polyacrylamide gel electrophoresis. The phosphorylation data show selective phosphorylation of specific proteins by cAMP-independent and cAMP-dependent protein kinase. Among the proteins in the postmitochondrial supernatant fraction phosphorylated by cAMP-dependent protein kinases, two proteins with a molecular weight of 43,000 were heavily phosphorylated. It is suggested that phosphorylation of cellular proteins by cAMP-dependent protein kinases might be involved in the cAMP-modulated biochemical changes in neuroblastoma cells.