PROTEIN-KINASE-C MEDIATED ACTIVATION AND PHOSPHORYLATION OF CA-2+ PUMP IN CARDIAC SARCOLEMMA

The effects of purified protein kinase C (PKC) on the Ca2+-pumping ATPase of cardiac sarcolemma were investigated. The addition of PKC to sarcolemmal vesicles resulted in a significant increase in ATP-dependent Ca2+ uptake, by increasing the calcium affinity by 2.8-fold (K(m) 0.14 vs. 0.4 muM for control) and by increasing V(max) from 5 to 6.8 nmol . mg protein-1 . min-1. The addition of PKC also stimulated Ca2+ ATPase activity in sarcolemmal preparations. This activity was increased further upon the addition of calmodulin. These results suggest that PKC stimulates Ca2+ ATPase through a kinase-directed phosphorylation. The addition of PKC to a purified preparation of Ca2+ ATPase in the presence of [gamma-P-32]ATP resulted in a 100% increase in phosphorylation that was dependent on the presence of Ca2+ ,phosphatidylserine, and phorbol 12,13-dibutyrate. These results demonstrate that the Ca2+ ATPase of canine cardiac muscle can be phosphorylated by PKC in vitro, resulting in increased affinity of the Ca2+ ATPase for Ca2+ and increase in the Ca2+ pump pumping rate. The results suggest that the Ca2+-pumping ATPase in heart tissue can be stimulated by PKC, thereby regulating the intracellular Ca2+ levels in whole heart.