Identification of talin as a major cytoplasmic protein implicated in platelet activation

Abstract

During platelet activation there is a major reorganization in the platelet cytoskeleton that accompanies a rapid change in platelet shape^1,2. Many of the events associated with activation are attributed to a rise in calcium concentration within the platelet cytoplasm^3,4. One direct consequence of the elevated calcium is the activation of a calcium-dependent protease that cleaves a major platelet protein of relative molecular mass (M_r) ∼235,000 (235K) to 200K (refs 5, 6). This protein, P235, has been purified⁶ and reported to interact with actin⁷, but the significance of the proteolytic cleavage is unknown. Talin, a cytoskeletal protein in smooth muscle and fibroblasts^8,9, binds vinculin¹⁰ and, together with vinculin^8,9,11,12, is localized in fibroblasts at sites of actin–membrane attachment. Talin and P235 have similar purification procedures, sedimentation coefficients and Stokes' radii (ref. 6 and Molony et al., unpublished observations). Of particular significance, talin is readily cleaved by proteases from ∼215K to a fragment of ∼190K²⁷. Given these similarities we have investigated the possible relationship between these proteins. Here we demonstrate that platelet P235 is recognized by anti-talin antibody and that it binds vinculin. Both proteins are cleaved in vitro by the calcium-activated protease to yield similar fragments. We conclude that P235 corresponds to the platelet form of talin.