The behavior of calpain-generated N- and C-terminal fragments of talin in integrin-mediated signaling pathways

Our previous results showed that the binding of an adhesive ligand to integ rin alpha IIb beta(3) on the surface of platelets triggers the activation o f calpain and the limited proteolysis of talin by calpain. To explore the p hysiological significance of the calpain-mediated cleavage of talin, we ana lyzed the behavior of the calpain-generated fragments of talin (N-terminal 47 kDa and C-terminal 190 kDa) during platelet activation by biochemical an d immunoelectron microscopic studies. Intact talin and mu-calpain transloca te from the Triton X-100-soluble fraction to the insoluble fraction upon pl atelet stimulation by thrombin, and the limited proteolysis of talin occurs in the Triton X-100-insoluble fraction, the cytoskeletal fraction. The ful ly autolyzed 76-kDa mu-calpain (active form) is found predominantly in the Triton X-100-insoluble fraction in stimulated platelets. While the N-termin al 47-kDa fragment remains in the Triton X-100-insoluble fraction, the C-te rminal 190-kDa fragment is released into the Triton X-100-soluble fraction in a time-dependent manner. Immunoelectron microscopic observations reveale d that the 47-kDa fragment locates on the submembrane zone just beneath the plasma membrane, including the open canalicular systems, while most of the 190-kDa fragment exists diffusely in the cytoplasm in thrombin-stimulated platelets. These findings suggest that calpain may contribute to the reorga nization of the cytoskeleton in an integrin-mediated signaling pathway thro ugh the redistribution of the functional domain of talin. (C) 1999 Academic Press.