The cell substrate attachment (CSAT) antigen has properties of a receptor for laminin and fibronectin.

Abstract

The cell substrate attachment (CSAT) antigen is an integral membrane glycoprotein complex that participates in the adhesion of cells to extracellular molecules. The CSAT monoclonal antibody, directed against this complex, inhibited adhesion of cardiac and tendon fibroblasts and ske[etat myoblasts to both laminin and fibronectin, thus implicating the CSAT antigen in adhesion to these extracellular molecules.Equilibrium gel filtration was used to explore the hypothesis that the CSAT antigen functions as a cell surface receptor for both laminin and fibronectin. In this technique, designed for rapidly exchanging equilibria, the gel filtration column is pre-equilibrated with extracellular ligand to ensure receptor occupancy during its journey through the column. Both laminin and fibronectin formed complexes with the CSAT antigen. The association with laminin was inhibited by the CSAT monoclonal antibody; the associations with both fibronectin and laminin were inhibited by synthetic peptides containing the fibronectin cell-binding sequence. Estimates of the dissociation constants by equilibrium gel filtration agree well with those available from other measurements. This suggests that these associations are biologically significant. SDS PAGE showed that all three glycoproteins comprising the CSAT antigen were present in the antigen-ligand complexes. Gel filtration and velocity sedimentation were used to show that the three bands comprise an oligomeric complex, which provides an explanation for their functional association. The inhibition of adhesion by the CSAT monoclonal antibody and the association of the purified antigen with extracellular ligands are interpreted as strongly implicating the CSAT antigen as a receptor for both fibronectin and laminin and perhaps for other extracellular molecules as well.The interaction of cells with components of the extracellular matrix plays a major role in determining cell morphology, cell migration, and tissue maintenance (1). Putative sites of cell-matrix contact have been identified and studied using various microscopic techniques. In fibroblasts they occur in regions where the cellular cytoskeleton and associated components interact with the cell surface (2-4). Integral membrane proteins are hypothesized to be present in these sites and to serve as transmembrane links connecting cytoskeletal and the extracellular matrix components. These transmembrane proteins would then serve as dual receptors for cytoskeletal and extracellular matrix components.We have identified a candidate for such a cell surface molecule using an adhesion and morphology perturbing cell 2134 substrate attachment (CSAT) ~ monoclonal antibody (5). This monoclonal antibody alters the adhesion of several different cell types in culture. The nature and degree of perturbation by the CSAT monoclonal antibody is characteristic for different cell types (6). The antigen to which the antibody is directed has been purified by immunoaffinity chromatography and partially characterized (7). It is a complex of th...

Frog egg extracts can do a cell's work

“…He switched to a gel filtration strategy using buffer equilibrated with fibronectin so that the CSAT antigen always saw its ligand. Under these saturating conditions, the two molecules ran off the column bound together as a complex (Horwitz et al, 1985), a result confirmed independently by the Yamada lab (Akiyama et al, 1986). He and Buck used the same method to show that the antigen acted as a laminin receptor, and delivered a "home-run" by establishing the transmembrane link in vitro, with integrin binding both to the ECM fibronectin and to cytoplasmic talin (Horwitz et al, 1986).…”

Alpha 1 beta 1 integrin on neural crest cells recognizes some laminin substrata in a Ca(2+)-independent manner.

“…Consistent with antibody-blocking experiments, the HNK-1 antibody recognizes 165 x 103 and 120 x 103 Mr glycoproteins on neural crest ceils isolated using either /3~ (Lallier and Bronner-Fraser, 1991) or tx~ integrin antibodies. The avian ch integrin subunit isolated from gizzard has been shown previously to have a molecular mass of 165 x 103 (Syfrig et al, 1991) and the ~ subunit has an Mr of 120 x 103 (Horwitz et al, 1985). Although this is the first reported example of an integrin that functions in the absence of soluble divalent cations, our preliminary results suggest that a number of tissues possess divalent cation-independent integrins which mediate attachment to laminin, fibronectin, and collagens (our unpublished observations); this cation-independent attachment is lost as a function of developmental time.…”

“…Integrins are transmembrane, heterodimeric receptors that mediate attachment of cells to a variety of ECM molecules including fibronectin, laminin, vitronectin, various collagens, and tenascin (Horwitz et al, 1985;Buck r al., 1986;Hynes, 1987;Tomaselli et al, 1988;Bourdon and Ruoslahti, 1989). Recendy, a number of laminin-binding integrin heterodimers have been identified and isolated in mammalian ceils, including txz/3~, ot2~, ot3B~, otd3t, and txdL (Forsberg et al, 1990;Hall et al, 1990;Kramer et al, 1990;Toyota et al, 1990;Languino et al, 1989;Elices and Hemler, 1989;Carter et al, 1990;Lotz et al, 1990;Kirchofer et al, 1990;Sonnenberg et al, 1988Sonnenberg et al, , 1990Shaw et al, 1990;Dedhar and Sanlimien, 1990;Shimizu et al, 1990).…”

Expression of normal and mutant avian integrin subunits in rodent cells [published erratum appears in J Cell Biol 1989 Oct;109(4 Pt 1):1187]

“…Those integrins involved in cell-substratum adhesion are found concentrated in or around focal contacts on the ventral cell surface, colocalizing with extracellular matrix (ECM)' molecules and cytoskeleton-associated (CSK) molecules (Chen et al, 1985;Damsky et al, 1985;Singer et al, 1988;Dejanna et al, 1988). Integrins are capable of binding directly to ECM molecules, including fibronectin, vitronectin, or laminin (Pytela et al, 1985a,b;Horwitz et al, 1985;Akiyama et al, 1985;Gardner and Hynes, 1985;Johansson et al, 1987a,b;Wayner and Carter, 1987;Wayner et al, 1988;Gehlsen et al, 1988;Ignatius and Reichardt, 1988;Sonnenberg et al, 1988), and to CSK molecules such as talin . The integrity of the aft complex is required for binding to both ECM and CSK molecules .…”