The properdin-like type I repeats of human thrombospondin contain a cell attachment site.

Abstract

Abstract. Thrombospondin (TS) is a modular adhesive glycoprotein that contains three domains previously implicated in the attachment of cells to "IS. These include the amino-terminal heparin-binding domain, the carboxy terminal cell or platelet-binding domain, and an RGDA sequence of TS. We have characterized a mAb against human TS, designated A4.1, which inhibits the attachment of human melanoma cells (G361) to TS. The epitope for A4.1 lies within the amino terminal half of the central stalklike region of "IS which is distinct from the three known cell attachment sites. This region of TS is recovered in a 50-kD peptide after chymotryptic digestion of "IS in EDTA. It contains the procollagen-like domain of TS as well as three type I repeats of a 60-residue segment homologous to two malarial proteins and the complement proteins properdin, and factors C6 through C9. The purified chymotryptic fragment is an effective attachment factor for G361 cells. A4.1 blocks adhesion to the 50-kD domain, as do some sulfated glycoconjugates. RGD (and RGE) peptides and mAbs against other domains of TS are not inhibitory. Peptides (19 mers) based on the core homology sequence of the three type I repeats of TS are potent attachment factors for these cells, and this adhesion is also inhibited by sulfated glycoconjugates. A polyclonal antibody raised against one of these peptides inhibits adhesion of G361 cells to the peptides, to the 50-kD fragment and to intact "IS. Thus a new cell-adhesion site has been identified in TS whose sequence is very similar to the site identified in region II of the circumsporozoite protein of malaria parasites (Rich, K. A., E W. George IV, J. L. Law, and W. J. Martin. 1990. Science (Wash. DC) 249:1574-1577. Thus there may be a common receptor which binds TS, malarial proteins, and properdin.

Differential expression of thrombospondin 1, 2, and 3 during murine development

“…Further investigation established that TSP was also secreted by a variety of cultured cells that include endothelial cells (McPherson et al, 1981;Mosher et al, 1982;Canfield et al, 1990), smooth muscle cells and fibroblasts (Raugi et al, 1982;Jaffe et al, 1983), keratinocytes (Wikner et al, 1987), macrophages (Jaffe et al, 1985), and glial cells (Asch et al, 1986). These findings, coupled with reports describing several cell-surface receptors for the protein Taraboletti et al, 1987;Lawler et al, 1988;Kaesberg et al, 1989;Frazier, 1991;Prater et al, 1991;Guo et al, 1992;Leung et al, 1992), indicated a broader spectrum of functions than that initially recognized in coagulation.…”

Cooperation between Thrombospondin-1 Type 1 Repeat Peptides and αvβ3 Integrin Ligands to Promote Melanoma Cell Spreading and Focal Adhesion Kinase Phosphorylation

“…Recombinant fragments and GST or T7 fusion proteins expressing fragments of TSP1 were prepared as described (15,19,20). Synthetic peptides containing TSP1 sequences were prepared as described previously (2,14,(21)(22)(23)(24). Bovine type I collagen was obtained from Collaborative Research, and human vitronectin was from Sigma.…”

“…The Mal II peptide promotes adhesion of these cells (23), but it does not activate ␣ v ␤ 3 integrin. Direct adhesion to peptides 246 and 7N3 in the absence of an ␣ v ␤ 3 integrin ligand induces a signal that opposes that induced by the same peptide when added in solution.…”

Stimulation of integrin-mediated adhesion of T lymphocytes and monocytes: two mechanisms with divergent biological consequences.

“…After a 30-rain incubation at 37~ the nonadherent cells were washed offwith 3 rounds of gentle pipetting. The residual adherent cells were checked by visual inspection, and quantified with a colorimetric reaction using endogenous cellular acid phosphatase activity (52). Optical densities were determined using an ELISA plate reader (Molecular Devices Corp., Menlo Park, CA) with a 410-nM filter.…”