Protein kinase C contributes to the maintenance of contractile force in human ventricular cardiomyocytes.
Prolonged Ca(2+) stimulations often result in a decrease in contractile force of isolated, demembranated human ventricular cardiomyocytes, whereas intact cells are likely to be protected from this deterioration. We hypothesized that cytosolic protein kinase C (PKC) contributes to this protection. Prolonged contracture (10 min) of demembranated human cardiomyocytes at half-maximal Ca(2+) resulted in a 37 +/- 5% reduction of active force (p < 0.01), whereas no decrease (2 +/- 3% increase) was observed in the presence of the cytosol (reconstituted myocytes). The PKC inhibitors GF 109203X and Gö 6976 (10 micromol/liter) partially antagonized the cytosol-mediated protection (15 +/- 5 and 9 +/- 2% decrease in active force, p < 0.05). Quantitation of PKC isoform expression revealed the dominance of the Ca(2+)-dependent PKCalpha over PKCdelta and PKCepsilon (189 +/- 31, 7 +/- 3, and 7 +/- 2 ng/mg protein, respectively). Ca(2+) stimulations of reconstituted human cardiomyocytes resulted in the translocation of endogenous PKCalpha, but not PKCbeta1, delta, and epsilon from the cytosol to the contractile system (PKCalpha association: control, 5 +/- 3 arbitrary units; +Ca(2+), 39 +/- 8 arbitrary units; p < 0.01, EC(50,Ca) = 645 nmol/liter). One of the PKCalpha-binding proteins were identified as the thin filament regulatory protein cardiac troponin I (TnI). Finally, the Ca(2+)-dependent interaction between PKCalpha and TnI was confirmed using purified recombinant proteins (binding without Ca(2+) was only 28 +/- 18% of that with Ca(2+)). Our data suggest that PKCalpha translocates to the contractile system and anchors to TnI in a Ca(2+)-dependent manner in the human heart, contributing to the maintenance of contractile force.
Division of Clinical Physiology, Institute of Cardiology, Faculty of Medicine, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary.
Andrea
A
Molnár
Attila
A
Borbély
Dániel
D
Czuriga
Siket M
SM
Ivetta
Szabolcs
S
Szilágyi
Zita
Z
Hertelendi
Eniko T
ET
Pásztor
Agnes
A
Balogh
Zoltán
Z
Galajda
Tamás
T
Szerafin
Kornelia
K
Jaquet
Zoltán
Z
Papp
István
I
Edes
Attila
A
Tóth
D000328:Adult; D002118:Calcium / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D006352:Heart Ventricles / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D009119:Muscle Contraction; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000201:enzymology*; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D013379:Substrate Specificity; D019210:Troponin I / Q000378:metabolism
D007527:Isoenzymes; D019210:Troponin I; D011493:Protein Kinase C; D002118:Calcium