Calcium-and phorbol ester-dependent calponin phosphorylation in homogenates of swine carotid artery.
Calponin inhibits actin-activated myosin adenosinetriphosphatase (ATPase) activity, and phosphorylation reverses this inhibition. Calponin phosphorylation has been demonstrated in reconstituted contractile protein systems, but studies using intact smooth muscle have produced mixed results. The goal of this study was to determine if vascular smooth muscle contains the necessary biochemical machinery to catalyze calponin phosphorylation. We used swine carotid homogenate, which allows access to the intracellular components and contains all endogenous proteins and enzymes in physiologically relevant concentrations. We demonstrated that calponin is phosphorylated in response to Ca2+ (0.27 +/- 0.04 mol P(i)/mol calponin) and in response to phorbol 12,13-dibutyrate in the presence or absence of Ca2+ (0.48 +/- 0.09 mol P(i)/mol calponin). Calponin phosphorylation was inhibited by the protein kinase C inhibitor staurosporine but not by the Ca(2+)- and calmodulin-dependent protein kinase II inhibitor KN-62. We conclude that Ca(2+)-dependent and -independent isoforms of protein kinase C but not the Ca(2+) -and calmodulin-dependent protein kinase II catalyze calponin phosphorylation in the swine carotid artery.
Rokolya
A
A
Bockus Research Institute, Graduate Hospital, Philadelphia, Pennsylvania 19146, USA.
Walsh
M P
MP
Moreland
R S
RS
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002339:Carotid Arteries / Q000378:metabolism*; D004586:Electrophoresis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008840:Microfilament Proteins; D009124:Muscle Proteins / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019311:Staurosporine / Q000494:pharmacology; D013552:Swine; D000096985:Calponins
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D002118:Calcium; D002135:Calcium-Binding Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D004791:Enzyme Inhibitors; D008840:Microfilament Proteins; D009124:Muscle Proteins; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D019311:Staurosporine; D000096985:Calponins; C063302:KN 62