TY - JOUR
T1 - Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium
AU - Van Der Velden, Jolanda
AU - Narolska, Nadiya A.
AU - Lamberts, Regis R.
AU - Boontje, Nicky M.
AU - Borbély, Attila
AU - Zaremba, Ruud
AU - Bronzwaer, Jean G.F.
AU - Papp, Zoltan
AU - Jaquet, Kornelia
AU - Paulus, Walter J.
AU - Stienen, Ger J.M.
PY - 2006/3/1
Y1 - 2006/3/1
N2 - Objective: In human heart failure β-adrenergic-mediated protein kinase A (PKA) activity is down-regulated, while protein kinase C (PKC) activity is up-regulated. PKC-mediated myofilament protein phosphorylation might be detrimental for contractile function in cardiomyopathy. This study was designed to reveal the effects of PKC on myofilament function in human myocardium under basal conditions and upon modulation of protein phosphorylation by PKA and phosphatases. Methods: Isometric force was measured at different [Ca 2+] in single permeabilized cardiomyocytes from non-failing and failing human left ventricular tissue. Basal phosphorylation of myofilament proteins and the influence of PKC, PKA, and phosphatase treatments were analyzed by one- and two-dimensional gel electrophoresis, Western immunoblotting, and ELISA. Results: Troponin I (TnI) phosphorylation at the PKA sites was decreased in failing compared to non-failing hearts and correlated well with myofilament Ca2+ sensitivity (pCa50). Incubation with the catalytic domain of PKC slightly decreased maximal force under basal conditions, but not following PKA and phosphatase pretreatments. PKC reduced Ca2+ sensitivity to a larger extent in failing (ΔpCa50 = 0.19 ± 0.03) than in non-failing (ΔpCa50 = 0.08 ± 0.01) cardiomyocytes. This shift was reduced, though still significant, when PKC was preceded by PKA, while PKA following PKC did not further decrease pCa 50. Protein analysis indicated that PKC phosphorylated PKA sites in human TnI and increased phosphorylation of troponin T, while myosin light chain phosphorylation remained unaltered. Conclusion: In human myocardium PKC-mediated myofilament protein phosphorylation only has a minor effect on maximal force development. The PKC-mediated decrease in Ca2+ sensitivity may serve to improve diastolic function in failing human myocardium in which PKA-mediated TnI phosphorylation is decreased.
AB - Objective: In human heart failure β-adrenergic-mediated protein kinase A (PKA) activity is down-regulated, while protein kinase C (PKC) activity is up-regulated. PKC-mediated myofilament protein phosphorylation might be detrimental for contractile function in cardiomyopathy. This study was designed to reveal the effects of PKC on myofilament function in human myocardium under basal conditions and upon modulation of protein phosphorylation by PKA and phosphatases. Methods: Isometric force was measured at different [Ca 2+] in single permeabilized cardiomyocytes from non-failing and failing human left ventricular tissue. Basal phosphorylation of myofilament proteins and the influence of PKC, PKA, and phosphatase treatments were analyzed by one- and two-dimensional gel electrophoresis, Western immunoblotting, and ELISA. Results: Troponin I (TnI) phosphorylation at the PKA sites was decreased in failing compared to non-failing hearts and correlated well with myofilament Ca2+ sensitivity (pCa50). Incubation with the catalytic domain of PKC slightly decreased maximal force under basal conditions, but not following PKA and phosphatase pretreatments. PKC reduced Ca2+ sensitivity to a larger extent in failing (ΔpCa50 = 0.19 ± 0.03) than in non-failing (ΔpCa50 = 0.08 ± 0.01) cardiomyocytes. This shift was reduced, though still significant, when PKC was preceded by PKA, while PKA following PKC did not further decrease pCa 50. Protein analysis indicated that PKC phosphorylated PKA sites in human TnI and increased phosphorylation of troponin T, while myosin light chain phosphorylation remained unaltered. Conclusion: In human myocardium PKC-mediated myofilament protein phosphorylation only has a minor effect on maximal force development. The PKC-mediated decrease in Ca2+ sensitivity may serve to improve diastolic function in failing human myocardium in which PKA-mediated TnI phosphorylation is decreased.
KW - Heart failure
KW - Myofilament function
KW - Protein kinase C
KW - Protein phosphorylation
UR - http://www.scopus.com/inward/record.url?scp=32444442576&partnerID=8YFLogxK
U2 - 10.1016/j.cardiores.2005.11.021
DO - 10.1016/j.cardiores.2005.11.021
M3 - Article
C2 - 16376870
AN - SCOPUS:32444442576
SN - 0008-6363
VL - 69
SP - 876
EP - 887
JO - Cardiovascular Research
JF - Cardiovascular Research
IS - 4
ER -