Phosphorylation of protein kinase C sites Ser42/44 decreases Ca 2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes

Paul J.M. Wijnker, Vasco Sequeira, E. Rosalie Witjas-Paalberends, D. Brian Foster, Cristobal G. Dos Remedios, Anne M. Murphy, Ger J.M. Stienen, Jolanda Van Der Velden

Research output: Contribution to journalArticlepeer-review

Abstract

Protein kinase C (PKC)-mediated phosphorylation of troponin I (cTnI) at Ser42/44 is increased in heart failure. While studies in rodents demonstrated that PKC-mediated Ser42/44 phosphorylation decreases maximal force and ATPase activity, PKC incubation of human cardiomyocytes did not affect maximal force. We investigated whether Ser42/44 pseudo-phosphorylation affects force development and ATPase activity using troponin exchange in human myocardium. Additionally, we studied if pseudo-phosphorylated Ser42/44 modulates length-dependent activation of force, which is regulated by protein kinase A (PKA)-mediated cTnI-Ser23/24 phosphorylation. Isometric force was measured in membrane-permeabilized cardiomyocytes exchanged with human recombinant wild-type troponin or troponin mutated at Ser42/44 or Ser23/24 into aspartic acid (D) or alanine (A) to mimic phosphorylation and dephosphorylation, respectively. In troponin-exchanged donor cardiomyocytes experiments were repeated after PKA incubation. ATPase activity was measured in troponin-exchanged cardiac muscle strips. Compared to wild-type, 42D/44D decreased Ca2+-sensitivity without affecting maximal force in failing and donor cardiomyocytes. In donor myocardium, 42D/44D did not affect maximal ATPase activity or tension cost. Interestingly, 42D/44D blunted the length-dependent increase in Ca 2+-sensitivity induced upon PKA-mediated phosphorylation. Since the drop in Ca2+-sensitivity at physiological Ca2+- concentrations is relatively large phosphorylation of Ser42/44 may result in a decrease of force and associated ATP utilization in the human heart.

Original languageEnglish (US)
Pages (from-to)11-21
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume554
DOIs
StatePublished - Jul 15 2014

Keywords

  • Cardiomyocyte
  • Myofilament function
  • Protein kinase C
  • Protein phosphorylation
  • Troponin I

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

Fingerprint Dive into the research topics of 'Phosphorylation of protein kinase C sites Ser42/44 decreases Ca <sup>2+</sup>-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes'. Together they form a unique fingerprint.

Cite this