Thrombospondin (TSP) is a member of the platelet adhesive glycoprotein family. This classification is indicated by the lectin- like activity of TSP, as well as the ability of several alpha-TSP antibodies to inhibit platelet aggregation. Despite apparent functional similarities, TSP is a unique platelet adhesive glycoprotein as it associates with the cell surface by two classes of receptors, one of which is divalent ion independent and exists on resting platelets. Moreover, TSP associates with the cell surface of thrombasthenic platelets which lack GPIIb-IIIa. The other platelet adhesive proteins, fibrinogen, fibronectin, and von Willebrand factors, interest only with stimulated platelets in the presence of divalent ions and do not bind to thrombasthenic cells. Moreover, the popular hypothesis that the monoclonal antibody, OKM5, identifies the platelet TSP receptor has been rigorously tested and proven inaccurate. Thus, the first aim of the proposed studies is to identify and TSP receptors on the surface of platelets. An initial approach will be affinity chromatography. Either TSP, an isolated domain of TSP, or TSP peptide retaining platelet binding capacity will be coupled to Sepharose. EDTA and TSP will be used to elute the bound receptor from the column. A second approach will involve chemical cross-linking of TSP, a fragment of TSP, or TSP peptide to platelets followed by immunoprecipitation of the resulting complex with an anti-TSP antibody. The components of the complex will be identified, isolated, and characterized. The second objective of this proposal is to probe the functional significance of TSP-platelet membrane interactions. These studies will require the use of purified TSP, previously characterized anti-TSP antibodies or the generation and use of anti-TSP receptor polyclonal and monoclonal antibodies which will be screened for perturbation of platelet function.
The final aim of this proposal is to determine if the identified receptors are present on the other cell types reported to bind TSP and elucidate the physiological role of these TSP receptors. It is believed that these studies will establish TSP as a cellular adhesive protein with functional significance for the platelet as well as several other cell types.
| Aiken, M L; Ginsberg, M H; Byers-Ward, V et al. (1990) Effects of OKM5, a monoclonal antibody to glycoprotein IV, on platelet aggregation and thrombospondin surface expression. Blood 76:2501-9 |
