Expression of a structural domain of the beta(2) subunit essential for alpha(M)beta(2) ligand recognition

The beta(2) leukocyte integrins comprise a group of closely related adhesio n receptors that mediate critical events during normal and inflammatory imm une responses. Central to the understanding of beta(2) integrin function is the basis of Ligand recognition, Results from our laboratory and others in dicate the presence of multiple ligand contact points in both the alpha and beta subunit, As an approach to identify and characterize regulatory domai ns of the beta(2) subunit, we have generated two different subdomains of th e beta(2) subunit for expression on the surface of mammalian cells through a phosphatidylinositol glycan anchor. The first subdomain contains the puta tive beta(2) MIDAS motif implicated in ligand binding [beta(2)(LB)], wherea s the second beta(2) subdomain contains the cysteine-rich region [beta(2)(C R)]. Cells expressing alpha(M) and beta(2) constructs singly or cotransfect ed transiently in COS-7 cells were tested for the ability to bind to immobi lized iC3b. Cells bearing the recombinant alpha(M)beta(2)(LB) were capable of adhering to iC3b in a manner similar to that observed with the complete alpha(M)beta(2) heterodimer, in contrast, cells expressing alpha(M)beta(2)( CR) failed to adhere to immobilized iC3b. Moreover, cells bearing singly tr ansfected alpha or beta chains alone failed to adhere to immobilized iC3b. These results indicate that along with alpha(M), the beta(2)(LB) subdomain contains the sufficient components within the beta(2) subunit essential for ligand recognition. These findings support the hypothesis that the beta(2) submit cooperates with site(s) within the alpha(M) subunit in a receptor/c ation/ligand complex resulting in high-affinity Ligand interaction.