Identification of a new cell adhesion motif in two homologous peptides from the COOH-terminal cell binding domain of human thrombospondin.

Thrombospondin-1 (TS1) contains at least four domains that support cell attachment. The COOH-terminal cell binding domain (CBD) was first identified with a monoclonal antibody against TS1 that blocked secretion-dependent platelet aggregation. Subsequently, this domain of TS1 has been found to bind a number of normal and transformed cells. We have localized attachment sites for human melanoma cells (G361) within the CBD to two noncontiguous 30-residue peptides designated C4 and C7 (Kosfeld, M. D., and Frazier, W. A. (1991) J. Biol. Chem. 267, 16230-16236). Here we report studies to define the active sequences within C4 and C7. An octapeptide, RFYVVMWK (4N1-1), from C4 and a pentapeptide, IRVVM (7N3-1), from C7 were found to support attachment of G361 melanomas, K562 erythroleukemia cells, HT1080 fibrosarcomas, C32 amelanotic melanomas, and endothelial cells. These peptides also inhibit the adhesion of cells to the recombinant CBD of TS1. The hexapeptide RFYVVM (4N1-2) also inhibits cell attachment. The inhibitory effect of combinations of C4- and C7-derived peptides is synergistic. The sequences 4N1-1 and 7N3-1 of TS1 share homology with two cell adhesive peptides from laminin (LM), LMF9 and LMPA22-2, respectively. These TS1 and LM peptides are interchangeable in inhibiting the adhesion of G361 cells to LM or TS1, suggesting a possible sharing of receptors by LM and TS1. K562 cells, however, bound only to TS1, and this binding was inhibited preferentially by the TS1 CBD peptides, indicating a receptor specific for TS1 which does not recognize LM. The active TS1 peptides are highly conserved among five species and four isoforms of TS1. Homologs of the TS1 peptides are found in tenascin, a matrix protein that shares several properties with TS1 and in factor VIII, alpha 2-macroglobulin, and von Willebrand factor.