Regulation of Endogenous Phosphorylation of Specific Proteins in Synaptic Membrane Fractions from Rat Brain by Adenosine 3’: 5’-Monophosphate*

The phosphorylation of specific membrane proteins by endogenous protein kinase in synaptic membrane fractions from rat cerebrum has been studied using the technique of acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The endogenous phosphorylation of two specific membrane proteins was found to be regulated by adenosine 3’ : 5’-monophosphate (cyclic AMP). The minimal molecular weights of these proteins were determined, by gel electrophoresis in the presence of sodium dodecyl sulfate, to be 86,000 (Protein I) and 48,000 (Protein II). The endogenous phosphorylation of Protein I and Protein II was stimulated by cyclic AMP in the presence of magnesium ions. Approximately 80% and 75 % of the azP incorporated into Protein I and Protein II, respectively, was found in phosphoserine. The concentration of cyclic AMP required for maximal stimulation of the phosphorylation of both Protein I and Protein II was approximately 5 x 10m6 rd. Guanosine 3’:5’-monophosphate had no significant effect, at concentrations up to 10m4 M, on the phosphorylation of either of these proteins. The endogenous phosphorylation occurred rapidly, the phosphorylation of both Protein I and Protein II reaching maximal levels within 5 sec. After reaching a maximal level of phosphorylation, Protein II underwent rapid dephosphorylation. The cyclic AMP-stimulated endogenous phosphorylation of a protein corresponding in position on acrylamide gel electrophoresis to Protein I was ako observed in membrane fractions prepared from other neural tissues known to contain synapses, but membrane fractions prepared from several neural and non-neural tissues devoid of synapses, including lingual nerve, lung, liver, spleen, and kidney, all failed to reveal cyclic AMP-dependent phosphorylation of a protein band corresponding in position to Protein I. In contrast, cyclic AMP-dependent phospborylation of a protein band corresponding in position to Protein II was observed in several neural and non-neural tissues. The results are compatible with the hypothesis that the mediation of certain types of synaptic transmission may involve regulation by cyclic AMP of the level of phosphorylation of specific protein constituents of the synaptic membranes.