p21-activated kinase increases the calcium sensitivity of rat triton-skinned cardiac muscle fiber bundles via a mechanism potentially involving novel phosphorylation of troponin I.
Phosphorylation of myofilament proteins by kinases such as cAMP-dependent protein kinase and protein kinase C has been shown to lead to altered thin-filament protein-protein interactions and modulation of cardiac function in vitro. In the present study, we report that a small GTPase-dependent kinase, p21-activated kinase (PAK), increases the calcium sensitivity of Triton-skinned cardiac muscle fiber bundles. Constitutively active PAK3 caused an average 1.25-fold (25.0+/-6.0%, n=6) increase in force at pCa 5.75, 1.44-fold (44.0+/-7.78%, n=6) at pCa 6.25, and 2.41-fold (141.2+/-23.7%, n=4) at pCa 6.5, representing a change in pCa50 value of approximately 0.25. Constitutively active PAK3 produced no change in force under conditions of relaxation (pCa 8.0) or maximal contraction (pCa 4.5). Furthermore, an inactive, kinase-dead form of PAK3 failed to produce any change in force development at any pCa value. The myofilament proteins phosphorylated by PAK3, at pCa 6.5, are desmin, troponin T, troponin I, and an unidentified 70-kDa protein. Importantly, cardiac troponin I was found to be phosphorylated at serine 149 of human cardiac troponin I, representing a novel phosphorylation site. These findings suggest a novel mechanism of modulating the calcium sensitivity of cardiac muscle contraction.
Buscemi
Nina
N
Department of Physiology, Queen's University, Kingston, Ontario, Canada.
Foster
D Brian
DB
Neverova
Irina
I
Van Eyk
Jennifer E
JE
D019943:Amino Acid Substitution; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000494:pharmacology*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006352:Heart Ventricles / Q000378:metabolism; D018485:Muscle Fibers, Skeletal / Q000187:drug effects* / Q000378:metabolism; D009154:Mutation; D009206:Myocardium / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011092:Polyethylene Glycols / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013552:Swine; D019210:Troponin I / Q000378:metabolism*; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D019210:Troponin I; D011092:Polyethylene Glycols; D005982:Glutathione Transferase; C515911:PAK3 protein, human; C515913:Pak3 protein, rat; D017346:Protein Serine-Threonine Kinases; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins; D002118:Calcium