Cytoskeletal assembly and vinculin-cytoskeleton interaction in different phases of the activation of bovine platelets.
Vinculin is an Mr 130 kDa protein that has been implicated in membrane-cytoskeleton interaction in various cell types. It has been demonstrated that vinculin is not a cytoskeletal component in resting platelets, but part of it becomes associated with the cytoskeleton during thrombin-induced activation. In this study, using a quantitative immunoblotting technique, the relation of vinculin to the cytoskeleton in different phases of activation of bovine platelets was explored, and the process of incorporation of vinculin into the cytoskeleton was related to that of cytoskeletal assembly. The assembly of cytoskeleton proceeded at a significantly faster rate than the association of vinculin with it, which shows that the latter process is not due to passive trapping of vinculin into the Triton-insoluble residue, but certain biochemical changes had to occur before such an interaction became possible. When the formation of pseudopodia was prevented by cytochalasin B, but neither aggregation nor the release reaction induced by thrombin were inhibited, the recovery of vinculin in the Triton-insoluble residue even increased. In both time- and thrombin-concentration-dependent studies, poor correlation was found between vinculin-cytoskeleton association and the extent of aggregation. Activation with phorbol-myristate-acetate, which is a strong stimulus for aggregation but produces only a slight release in the granular content, resulted in the association of only a negligible amount of vinculin with the cytoskeletal fraction. The incorporation of vinculin into the cytoskeletal fraction of thrombin activated platelets started with the release reaction but still proceeded, and the greatest part of the reaction occurred after secretion had gone to completion.(ABSTRACT TRUNCATED AT 250 WORDS)
Horvath
A R
AR
Department of Clinical Chemistry, University School of Medicine, Debrecen, Hungary.
Asijee
G M
GM
Muszbek
L
L
D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D002417:Cattle; D002465:Cell Movement; D003571:Cytochalasin B / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D015151:Immunoblotting; D015539:Platelet Activation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D011554:Pseudopodia / Q000378:metabolism; D016596:Vinculin / Q000378:metabolism*
D016596:Vinculin; D003571:Cytochalasin B