Effect of troponin I Ser23/24 phosphorylation on Ca2+-sensitivity in human myocardium depends on the phosphorylation background

Viola Kooij, Martina Saes, Kornelia Jaquet, Ruud Zaremba, D. Brian Foster, Anne M. Murphy, Cris dos Remedios, Jolanda van der Velden, Ger J.M. Stienen

Research output: Contribution to journalArticlepeer-review

Abstract

Protein kinase A (PKA)-mediated phosphorylation of Ser23/24 of cardiac troponin I (cTnI) causes a reduction in Ca2+-sensitivity of force development. This study aimed to determine whether the PKA-induced modulation of the Ca2+-sensitivity is solely due to cTnI phosphorylation or depends on the phosphorylation status of other sarcomeric proteins. Endogenous troponin (cTn) complex in donor cardiomyocytes was partially exchanged (up to 66±1%) with recombinant unphosphorylated human cTn and in failing cells similar exchange was achieved using PKA-(bis)phosphorylated cTn complex. Cardiomyocytes immersed in exchange solution without complex added served as controls. Partial exchange of unphosphorylated cTn complex in donor tissue significantly increased Ca2+-sensitivity (pCa50) to 5.50±0.02 relative to the donor control value (pCa50=5.43±0.04). Exchange in failing tissue with PKA-phosphorylated cTn complex did not change Ca2+-sensitivity relative to the failing control (pCa50=5.60±0.02). Subsequent treatment of the cardiomyocytes with the catalytic subunit of PKA significantly decreased Ca2+-sensitivity in donor and failing tissue. Analysis of phosphorylated cTnI species revealed the same distribution of un-, mono- and bis-phosphorylated cTnI in donor control and in failing tissue exchanged with PKA-phosphorylated cTn complex. Phosphorylation of myosin-binding protein-C in failing tissue was significantly lower compared to donor tissue. These differences in Ca2+-sensitivity in donor and failing cells, despite similar distribution of cTnI species, could be abolished by subsequent PKA-treatment and indicate that other targets of PKA are involved the reduction of Ca2+-sensitivity. Our findings suggest that the sarcomeric phosphorylation background, which is altered in cardiac disease, influences the impact of cTnI Ser23/24 phosphorylation by PKA on Ca2+-sensitivity.

Original languageEnglish (US)
Pages (from-to)954-963
Number of pages10
JournalJournal of Molecular and Cellular Cardiology
Volume48
Issue number5
DOIs
StatePublished - May 2010

Keywords

  • Ca-sensitivity
  • Myocardium
  • Phosphorylation
  • Protein kinase A
  • Sarcomere
  • Single cardiomyocyte
  • Troponin I

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

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