Relaxin alters cardiac myofilament function through a PKC-dependent pathway.
The pregnancy hormone relaxin (RLX) is a powerful cardiostimulatory peptide. Despite its well-characterized effects on the heart, the intracellular mechanisms responsible for RLX's positive inotropic effects are unknown. Cardiac myofilaments are the central contractile elements of the heart, and changes in the phosphorylation status of myofilament proteins are known to mediate changes in function. The first objective of this study was to determine whether RLX stimulates myofilament activation and alters the phosphorylation of one or more myofilament proteins. RLX works through a variety of intracellular signaling cascades in different tissue types. Protein kinases A (PKA) and C (PKC) are two common molecules implicated in RLX signaling and are known to affect myofilament function. Thus the second objective of this study was to determine whether RLX mediates its myocardial effects through PKA or PKC activation. Murine myocardium was treated with recombinant H2-RLX, and cardiac myofilaments were isolated. RLX increased cardiac myofilament force development at physiological levels of intracellular Ca(2+) without altering myofilament ATP consumption. Myosin binding protein C, troponin T, and troponin I phosphorylation levels were increased with RLX treatment. Immunoblot analysis revealed an increase in myofilament-associated PKC-delta, decreases in PKC-alpha and -beta(II), but no effect on PKC-epsilon. Inhibition of PKC with chelerythrine chloride or PKC-delta with rottlerin prevented the RLX-dependent changes in myofilament function and protein phosphorylation. PKA antagonism with H-89 had no effect on the myofilament effects of RLX. This study is the first to show that RLX-dependent changes in myofilament-associated PKC alters myofilament activation in a manner consistent with its cardiostimulatory effects.
Department of Biomedical Sciences, Ontario Veterinary College, University of Guelph, Guelph, Ontario, Canada.
Erynn E
EE
Shaw
Philip
P
Wood
Justyna
J
Kulpa
Feng Hua
FH
Yang
Alastair J
AJ
Summerlee
W Glen
WG
Pyle
D008841:Actin Cytoskeleton / Q000187:drug effects*; D000818:Animals; D053119:Benzophenanthridines / Q000494:pharmacology; D015153:Blotting, Western; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D002316:Cardiotonic Agents; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006321:Heart / Q000187:drug effects*; D066298:In Vitro Techniques; D007537:Isometric Contraction / Q000187:drug effects; D007546:Isoquinolines / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D009206:Myocardium / Q000166:cytology / Q000201:enzymology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D051745:Protein Kinase C-delta / Q000502:physiology; D012065:Relaxin / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects*; D013449:Sulfonamides / Q000494:pharmacology
D053119:Benzophenanthridines; D002316:Cardiotonic Agents; D004791:Enzyme Inhibitors; D007546:Isoquinolines; D013449:Sulfonamides; D012065:Relaxin; C016299:chelerythrine; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D017301:Ca(2+) Mg(2+)-ATPase; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide