Protein-Tyrosine Phosphorylation and p72 ^syk Activation in Human Platelets Stimulated with Collagen Is Dependent upon Glycoprotein Ia/IIa and Actin Polymerization

Summary

In human platelets treated with acetylsalicylic acid, collagen induced protein-tyrosine-phosphorylation of several proteins. The major 75 kDa band included cortactin and autophosphorylated p72 ^syk . p72 ^syk activity rapidly increased upon collagen stimulation, whereas p^60c-src activation was below detectable levels. A combination of inhibitors to remove the effects of extracellular and intracellular Ca²⁺, released ADP, and fibrinogen binding to GPIIb/IIIa delayed and attenuated the major 75 kDa band. By contrast, p72 ^syk activation was not inhibited by these treatments. Cytochalasin D completely inhibited protein tyrosine phosphorylation and p72 ^syk activation. It also potently inhibited aggregation and [Ca²⁺]i elevation. Anti-GPMIa/IIa MoAb in a concentration-dependent manner partially attenuated protein tyrosine phosphorylation and p72 ^syk activation. Its inhibitory effects on intracellular Ca²⁺ mobilization, release of intracellular granule contents, and aggregation also were partial. No tyrosine kinase activity was coprecipitated with GPIa/IIa. These results suggest that p72 ^syk activation lies upstream of protein tyrosine phosphorylation, Ca²⁺ mobilization, ADP release, thromboxane A₂ production and aggregation. GPIa/IIa plays a key role in p72 ^syk activation induced by collagen, but other collagen receptors may work in synergy to fully activate p72 ^syk . Actin polymerization is a prerequisite for both p72 ^syk activation and other intracellular signal transduction pathways.

• References

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