Tyrosine Kinases Regulate the Cytoskeletal Attachment of Integrin a , , & ( Platelet Glycoprotein IIb / IIIa ) and the Cellular Retraction of Fibrin Polymers

Integrins promote cell-substratum and cell-cell adhesion by acting as transmembrane linker molecules between extracellular adhesion proteins and the actin-rich cytoskeleton. The integrin a,,& (platelet glycoprotein IIb/IIIa) is essential for platelet spreading, aggregation, fibrin clot retraction, and for the transduction of extracellular signals. We examined the effect of the specific tyrosine kinase inhibitor herbimycin A on integrin and cytoskeletal-mediated events in thrombin-stimulated platelets. Incubation of washed platelets for 24 h with herbimycin A (5 w) abolished the thrombin-stimulated cytoskeletal enzyme activity of pp60'"" in parallel with a reduction in the tyrosine phosphorylation of multiple platelet proteins, as assessed with anti-phosphotyrosine immunoblots. However, thrombin-induced activation of protein kinase C and the production of thromboxane A, were not altered by herbimycin A. Despite the absence of cytoskeletal pp60c"m enzyme activity, platelet shape change, aggregation, and serotonin release were unaltered following platelet stimulation with thrombin (0.051.0 unit/ml). Herbimycin A-treated platelets also demonstrated normal platelet aggregation in response to collagen (5 pg/ml), ionophore A23187 (2 p ~ ) , and ADP/ adrenaline (10 p~ each). However, the ability of herbimycin A-treated platelets to retract fibrin gels was significantly reduced. This defect in clot retraction was associated with reduced incorporation of integrin armbps into the cytoskeletal fraction of thrombin-aggregated platelets. Our studies suggest that tyrosine kinases in platelets regulate the cytoskeletal attachment of arm&, as an essential process for the transmission of cellular contractile forces to fibrin polymers.