Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium

Jolanda Van Der Velden, Nadiya A. Narolska, Regis R. Lamberts, Nicky M. Boontje, A. Borbély, Ruud Zaremba, Jean G F Bronzwaer, Z. Papp, Kornelia Jaquet, Walter J. Paulus, Ger J M Stienen

Research output: Contribution to journalArticle

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Abstract

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.

Original languageEnglish
Pages (from-to)876-887
Number of pages12
JournalCardiovascular Research
Volume69
Issue number4
DOIs
Publication statusPublished - Mar 1 2006

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Myofibrils
Protein Kinase C
Cyclic AMP-Dependent Protein Kinases
Myocardium
Phosphorylation
Troponin I
Phosphoprotein Phosphatases
Proteins
Cardiac Myocytes
Myosin Light Chains
Troponin T
Electrophoresis, Gel, Two-Dimensional
Cardiomyopathies
Adrenergic Agents
Catalytic Domain
Heart Failure
Western Blotting
Enzyme-Linked Immunosorbent Assay

Keywords

  • Heart failure
  • Myofilament function
  • Protein kinase C
  • Protein phosphorylation

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

Cite this

Van Der Velden, J., Narolska, N. A., Lamberts, R. R., Boontje, N. M., Borbély, A., Zaremba, R., ... Stienen, G. J. M. (2006). Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium. Cardiovascular Research, 69(4), 876-887. https://doi.org/10.1016/j.cardiores.2005.11.021

Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium. / Van Der Velden, Jolanda; Narolska, Nadiya A.; Lamberts, Regis R.; Boontje, Nicky M.; Borbély, A.; Zaremba, Ruud; Bronzwaer, Jean G F; Papp, Z.; Jaquet, Kornelia; Paulus, Walter J.; Stienen, Ger J M.

In: Cardiovascular Research, Vol. 69, No. 4, 01.03.2006, p. 876-887.

Research output: Contribution to journalArticle

Van Der Velden, J, Narolska, NA, Lamberts, RR, Boontje, NM, Borbély, A, Zaremba, R, Bronzwaer, JGF, Papp, Z, Jaquet, K, Paulus, WJ & Stienen, GJM 2006, 'Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium', Cardiovascular Research, vol. 69, no. 4, pp. 876-887. https://doi.org/10.1016/j.cardiores.2005.11.021
Van Der Velden, Jolanda ; Narolska, Nadiya A. ; Lamberts, Regis R. ; Boontje, Nicky M. ; Borbély, A. ; Zaremba, Ruud ; Bronzwaer, Jean G F ; Papp, Z. ; Jaquet, Kornelia ; Paulus, Walter J. ; Stienen, Ger J M. / Functional effects of protein kinase C-mediated myofilament phosphorylation in human myocardium. In: Cardiovascular Research. 2006 ; Vol. 69, No. 4. pp. 876-887.
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abstract = "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.",
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AU - Narolska, Nadiya A.

AU - Lamberts, Regis R.

AU - Boontje, Nicky M.

AU - Borbély, A.

AU - Zaremba, Ruud

AU - Bronzwaer, Jean G F

AU - Papp, Z.

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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.

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