Conformational regulation of the fibronectin binding and alpha(3)beta(1) integrin-mediated adhesive activities of thrombospondin-1

The recognition of extracellular matrix components can be regulated by conf ormational changes that alter the activity of cell surface integrins. We no w demonstrate that conformational regulation of the matrix glycoprotein thr ombospondin-1 (TSP1) can also modulate its binding to an integrin receptor. F18 1G8 is a conformation-sensitive TSP1 antibody that binds weakly to sol uble TSP1 in the presence of divalent cations. However, binding of the anti body to melanoma cells was strongly stimulated by adding exogenous TSP1 in the presence of calcium, suggesting that TSP1 undergoes a conformational ch ange following its binding to the cell surface. This conformation was not i nduced by known cell surface TSP1 receptors, whereas binding of F18 was sti mulated when TSP1 bound to fibronectin but not to heparin or fibrinogen. Co nversely, binding of F18 to TSP1 enhanced TSP1 binding to fibronectin. Exog enous fibronectin also stimulated TSP1-dependent binding of F18 to melanoma cells. Binding of the fibronectin-TSP1 complex to melanoma cells was media ted by alpha (4)beta (1) and alpha (5)beta (1) integrins. Furthermore, bind ing to F18 or fibronectin strongly enhanced the adhesive activity of immobi lized TSP1 for some cell types. This enhancement of adhesion was mediated b y alpha (3)beta (1) integrin and required that the alpha (3)beta (1) integr in be in an active state. Fibronectin also enhanced TSP1 binding to purifie d alpha (3)beta (1) integrin. Therefore, both fibronectin and the F18 antib ody induce conformational changes in TSP1 that enhance the ability of TSP1 to be recognized by alpha (3)beta (1) integrin. The conformational and func tional regulation of TSP1 activity by fibronectin represents a novel mechan ism for extracellular signal transduction.