Regulation of plasma membrane Ca2+-ATPase in human platelets by calpain.
The plasma membrane Ca(2+)-ATPase (PMCA) plays an essential role in maintaining low cytosolic Ca(2+) in resting human platelets by extruding Ca(2+) from the cytoplasm across the plasma membrane. Since PMCA is the main agent of Ca(2+) efflux in platelets, it is a key point for regulation of platelet Ca(2+) metabolism. PMCA has been shown to be an excellent substrate for the Ca(2+)-activated cysteine protease calpain, a major platelet protein that is turned on during platelet activation. The objectives of the present work were to determine if PMCA is degraded during thrombin- and collagen-mediated platelet activation, and if calpain is responsible. The kinetics of PMCA degradation during platelet activation were analysed using SDS polyacrylamide gel electrophoresis and immunoblotting. The role of calpain was tested using the calpain inhibitors calpeptin and ALLN. Platelet activation mediated by both collagen and thrombin resulted in degradation of 60% of platelet PMCA within 18 minutes. Calpeptin and ALLN significantly inhibited the rate and extent of PMCA degradation. We conclude that calpain-mediated degradation of PMCA during platelet activation likely contributes significantly to Ca(2+) regulation and, therefore, to platelet function.
Brown
Charles S
CS
Department of Biochemistry & Molecular Biology, University of Louisille School of Medicine, Louisville, KY 40292, USA.
Dean
William L
WL
D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology*; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002154:Calpain / Q000037:antagonists & inhibitors / Q000502:physiology*; D002462:Cell Membrane / Q000201:enzymology*; D003094:Collagen / Q000494:pharmacology; D004151:Dipeptides / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects / Q000502:physiology*; D013917:Thrombin / Q000494:pharmacology
D004151:Dipeptides; D007976:Leupeptins; C063261:acetylleucyl-leucyl-norleucinal; C071482:calpeptin; D003094:Collagen; D013917:Thrombin; D002154:Calpain; D000252:Calcium-Transporting ATPases; D002118:Calcium