Protein kinase D increases maximal Ca 2+ -activated tension of cardiomyocyte contraction by phosphorylation of cMyBP-C-Ser 315

Cardiac myosin-binding protein C (cMyBP-C) is involved in the regulation of cardiac myofilament contraction. Recent evidence showed that protein kinase D (PKD) is one of the kinases that phosphorylate cMyBP-C. However, the mechanism by which PKD-induced cMyBP-C phosphorylation affects cardiac contractile responses is not known. Using immunoprecipitation, we showed that, in contracting cardiomyocytes, PKD binds to cMyBP-C and phosphorylates it at Ser 315 . The effect of PKD-mediated phosphorylation of cMyBP-C on cardiac myofilament function was investigated in permeabilized ventricular myocytes, isolated from wild-type (WT) and from cMyBP-C knockout (KO) mice, incubated in the presence of full-length active PKD. In WT myocytes, PKD increased both myofilament Ca 2+ sensitivity (pCa 50 ) and maximal Ca 2+ -activated tension of contraction (T max ). In cMyBP-C KO skinned myocytes, PKD increased pCa 50 but did not alter T max . This suggests that cMyBP-C is not involved in PKD-mediated sensitization of myofilaments to Ca 2+ but is essential for PKD-induced increase in T max . Furthermore, the phosphorylation of both PKD-Ser 916 and cMyBP-C-Ser 315 was contraction frequency-dependent, suggesting that PKD-mediated cMyBP-C phosphorylation is operational primarily during periods of increased contractile activity. Thus, during high contraction frequency, PKD facilitates contraction of cardiomyocytes by increasing Ca 2+ sensitivity and by an increased T max through phosphorylation of cMyBP-C.