Inhibition of Binding of Fibronectin to Matrix Assembly Sites by Anti-Integrin Antibodies

Fibroblasts have cell surface sites that mediate assembly of plasma and cellular fibronectin into the extracellular matrix. Cell adhesion to fibronectin can be mediated by the interaction of an integrin (oL5/3~) with the Arg-Gly-Asp-Ser (RGDS)-containing cell adhesion region of fibronectin. We have attempted to elucidate the role of the otsfll fibronectin receptor in assembly of fibronectin in matrices. Rat monoclonal antibody mAb 13, which recognizes the integrin fll subunit, completely blocked binding and matrix assembly of 125I-fibronectin as well as binding of the 125170-kD amino-terminal fragment of fibronectin (70 kD) to fibroblast cell layers. Fab fragments of the anti-/~ antibody were also inhibitory. Antibody mAb 16, which recognizes the integrin ors subunit, partially blocked binding of 125I-fibronectin and 12q-70-kD. When cell layers were coincubated with fluoresceinated fibronectin and either anti-13~ or anti-us, anti-131 was a more effective inhibitor than anti-o~s of binding of labeled fibronectin to the cell layer. Inhibition of ~2q-fibmnectin binding by anti-/31 IgG occurred within 20 min. Inhibition of 125I-fibronectin binding by anti-/3t Fab fragments or IgG could not be overcome with increasing concentrations of fibronectin, suggesting that anti-131 and exogenous fibronectin may not compete for the same binding site. No/31-containing integrin bound to immobilized 70 kD. These data indicate that the/31 subunit plays an important role in binding and assembly of exogenous fibronectin, perhaps by participation in the organization, regeneration, or cycling of the assembly site rather than by a direct interaction with