Laminin modulates morphogenic properties of the collagen XVIII endostatin domain.

We have shown previously that the oligomeric endostatin domain of collagen XVIII (NC1) functioned as a motility-inducing factor regulating the extracellular matrix-dependent morphogenesis of endothelial cells. This motogenic activity gave rise to structures resembling filipodia and lamellipodia and was dependent on Rac, Cdc42, and mitogen-activated protein kinase. Here, we demonstrate that these properties of endostatin are primarily mediated by laminin in the basement membrane and heparan sulfates on the cell surface. The sites of interaction between laminin and oligomeric endostain include the N-terminal regions of all three laminin chains (amino acids 204-1243 of the alpha chain, 932-1161 of the beta chain, and 150-965 of the gamma chain). A monoclonal antibody that blocks the interactions between endostatin and laminin was utilized to inhibit the motogenic activity of endostatin. In parallel, we have engineered selective point mutations and produced recombinant forms that lack binding to heparan sulfates on the cell surface. Our data are consistent with a model of endostatin with two binding sites: one mainly to laminin in the basement membrane and the other to heparan sulfates on the cell surface. The two binding domains on endostatin appear to be separate with the possibility of some overlap between the two sites.