The vitronectin receptor αvβ3 binds fibronectin and acts in concert with α5β1 in promoting cellular attachment and spreading on fibronectin

Abstract

The vitronectin receptor (αvβ3) is a member of the integrin superfamily of adhesive protein receptors that mediate a wide spectrum of adhesive cellular interactions, including attachment to vitronectin, von Willebrand factor, fibrinogen, and thrombospondin. We have studied the binding of fibronectin to the purified vitronectin receptor, and the role of this receptor in the attachment of cells to fibronectin. A solid-phase microtiter assay was developed to investigate the binding properties of the vitronectin receptor. Purified αvβ3 bound fibronectin with high affinity in a saturable, divalent cation-dependent manner. Binding was inhibited by soluble vitronectin, by RGD-containing peptides, and by LM609, a monoclonal antibody against the vitronectin receptor known to inhibit the binding of adhesive proteins to αvβ3. Immunoinhibition experiments showed that M21 human melanoma cells, which express the fibronectin receptor, α5β1, as well as αvβ3, used both of these integrins to attach and spread on fibronectin. In support of this finding, M21-L cells, a variant cell line that specifically lacks αvβ3 but expresses αvβ1, attached and spread poorly on fibronectin. In addition, αvβ3 from surface-labeled M21 cells was retained, and selectively eluted by RGDS from a fibronectin affinity column. These results indicate that αvβ3 acts in concert with α5β1 in promoting fibronectin recognition by these cells. We conclude that fibronectin binds to the αvβ3 vitronectin receptor specifically and with high affinity, and that this interaction is biologically relevant in supporting cell adhesion to matrix proteins.