Cardiac troponin I phosphorylation increases the rate of cardiac muscle relaxation.
Cardiac troponin (Tn) I (CTnI), compared with skeletal TnI, contains extra amino acids (32 to 33) at its amino terminus, including two adjacent serine residues. These two serine residues are believed to be phosphorylated by protein kinase A (PKA) upon stimulation of the heart by beta-agonists. In this study, we found that phosphorylation of a cardiac skinned muscle preparation by PKA, mainly at CTnI, results in a decrease in the Ca2+ sensitivity of muscle contraction. The pCa50 decreased by approximately 0.27 +/- 0.06 pCa units upon phosphorylation. To study cardiac muscle relaxation, we used diazo-2, a photolabile Ca2+ chelator with a low Ca2+ affinity in its intact form that is converted to a high-affinity form after photolysis. We found that the rate of cardiac muscle relaxation increased from a time of half-relaxation (t1/2) = 110 +/- 10 milliseconds to t1/2 = 70 +/- 8 milliseconds after CTnI phosphorylation. This result demonstrates that CTnI phosphorylation can be linked with the increased rate of muscle relaxation in a relatively intact muscle preparation. Since CTnI phosphorylation has been shown previously to affect the Ca2+ affinity and Ca2+ off-rate of CTnC in vitro, it is likely that the faster relaxation seen here reflects faster dissociation of Ca2+ from cardiac TnC (CTnC). Model calculations show that increased dissociation of Ca2+ from CTnC, coupled with the faster uptake of Ca2+ by the sarcoplasmic reticulum stimulated by PKA phosphorylation of phospholamban, can account for the faster relaxation seen in the inotropic response of the heart to catecholamines.
Zhang
R
R
Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, FL 33101, USA.
Zhao
J
J
Mandveno
A
A
Potter
J D
JD
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D003198:Computer Simulation; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003979:Diazonium Compounds; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000201:enzymology / Q000378:metabolism*; D010642:Phenoxyacetates; D010766:Phosphorylation; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D013552:Swine; D013997:Time Factors; D014336:Troponin / Q000378:metabolism*; D019210:Troponin I; D000099211:Phospholamban
D000251:Adenosine Triphosphatases; D002118:Calcium; D002135:Calcium-Binding Proteins; D002614:Chelating Agents; D017868:Cyclic AMP-Dependent Protein Kinases; D003979:Diazonium Compounds; D010642:Phenoxyacetates; D014336:Troponin; D019210:Troponin I; D000099211:Phospholamban; C063685:diazo-2