Protein kinase D increases maximal Ca²⁺-activated tension of cardiomyocyte contraction by phosphorylation of cMyBP-C-Ser³¹⁵

Abstract

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.

Emergency Spatiotemporal Shift: The Response of Protein Kinase D to Stress Signals in the Cardiovascular System

“…A number of PKD substrates have been identified thus far: the titin-cap protein telethonin (Candasamy et al, 2014), troponin I (TnI; Cuello et al, 2007) and Myosin Binding Protein c (MyBPc; Bardswell et al, 2010; Dirkx et al, 2012a). Upon phosphorylation of TnI at Serine 22 and 23, the sites also targeted by PKA, myofilament Ca 2+ sensitivity is reduced.…”

Phosphorylation of Cardiac Myosin-Binding Protein-C Is a Critical Mediator of Diastolic Function

“…5, 6 cMyBP-C is readily phosphorylated by a host of kinases such as protein kinase A (PKA) 7 , PKC 8 , PKD 9 , calcium calmodulin kinase IIδ (CAMK2δ) 10 , glycogen synthase kinase 3β (GSK3β) 11 and ribosomal S6 kinase (RSK6) 12 . Unphosphorylated cMyBP-C appears to repress both cross-bridge attachment and detachment.…”

Myofilament Calcium Sensitivity: Role in Regulation of In vivo Cardiac Contraction and Relaxation

“…One possibility might be various kinases that increase the calcium sensitivity of myofilaments. For example, protein kinase D (PKD) has been reported to increase calcium sensitivity via phosphorylation of Ser 315 cardiac myosin binding protein C (cMyBP-C) (Dirkx et al, 2012). However, PKD can also reduce calcium sensitivity via phosphorylation of troponin I (TnI) Ser 23/24 (Cuello et al, 2007).…”