Relocation of Syk protein-tyrosine kinase to the actin filament network and subsequent association with Fak.
Previous studies demonstrated that Syk protein-tyrosine kinase (Syk) is activated by thrombin in platelets. To elucidate the function of Syk in platelets, we have biochemically examined the intracellular location of Syk and the molecules associated with Syk, following platelet activation. In human platelets, thrombin induces the relocation of Syk to the cytoskeletal fraction presumably via Syk tyrosine phosphorylation. Relocated Syk is associated with the actin filament network, and the early phase (10-90 s) of this association can be partially inhibited by the pretreatment of platelets with cytochalasin D, an inhibitor of actin polymerization. Upon thrombin stimulation, Syk becomes associated with Fak as demonstrated by co-immunoprecipitation. The association of both kinases can be inhibited by pretreatment of platelets with cytochalasin D. Interestingly, reconstitution experiments, using COS cells transfected with various porcine Syk mutants, revealed that the kinase domain, but not the kinase activity, of Syk is required for the association of Syk with the actin filament network. These findings suggest that thrombin-induced association of Syk with Fak correlates with the state of actin polymerization, and may play an important role in platelet activation.
Sada
K
K
Department of Biochemistry, Kobe University School of Medicine, Japan.
Minami
Y
Y
Yamamura
H
H
D008841:Actin Cytoskeleton / Q000201:enzymology*; D000199:Actins / Q000378:metabolism*; D000818:Animals; D001792:Blood Platelets / Q000201:enzymology*; D019556:COS Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D015539:Platelet Activation / Q000502:physiology; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000072377:Syk Kinase; D013917:Thrombin / Q000494:pharmacology; D014162:Transfection / Q000235:genetics
D000199:Actins; D015815:Cell Adhesion Molecules; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D019000:Phosphotyrosine; D015638:Cytochalasin D; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; C000606896:SYK protein, human; D000072377:Syk Kinase; D013917:Thrombin