FIGURE 4.

c-Src phosphorylation induced by anti-αIIb TM requires the presence of αIIbβ3. A, WT and GT platelets were incubated for 1 min with 10 μg/ml collagen, 2 μm anti-αIIb TM, or 2 μm anti-αIIb TM plus 10 μm PP2. Platelet lysates were then immunoblotted (IB) for β3, c-Src Tyr(P)⁴¹⁹, and c-Src. MW, molecular weight. B, washed platelets were incubated with buffer, 2 μm anti-αIIb TM, 1 units/ml thrombin, or 2 μm MS1 for 1 min. Platelet lysates were then immunoblotted for c-Src and c-Src Tyr(P)⁴¹⁹. C, washed platelets were incubated with buffer or 2 μm anti-αIIb TM for 1 min in the absence or the presence of 5 mm EDTA, 10 μm eptifibatide, 4 μm R406, 10 μm cytochalasin D (Cyto D), 2 μm latrunculin (Lat A), 10 μm PP2, or 10 μm PP3. c-Src was immunoprecipitated from platelet lysates and immunoblotted for c-Src and c-Src Tyr(P)⁴¹⁹. Top panel, c-Src Tyr(P)⁴¹⁹. Bottom panel, c-Src loading control. The experiments were performed three times. D, the bar graph was generated from densitometry of the immunoblots from the three experiments and represents the mean ± S.E. of c-Src Tyr(P)⁴¹⁹ normalized using the densitometry of the corresponding c-Src band.