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 language | English (US) |
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Pages (from-to) | 954-963 |
Number of pages | 10 |
Journal | Journal of Molecular and Cellular Cardiology |
Volume | 48 |
Issue number | 5 |
DOIs | |
State | Published - 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