DIFFERENTIAL CATION REGULATION OF THE ALPHA(5)BETA(1) INTEGRIN-MEDIATED ADHESION OF LEUKEMIC-CELLS TO THE CENTRAL CELL-BINDING DOMAIN OF FIBRONECTIN

Normal and neoplastic leukocytes interact with the central cell-bindin g and carboxyl-terminal regions of fibronectin (FN) primarily via the alpha(4) beta(1) and alpha(5) beta(1) integrins. By using a unique cen trifugation-based cell adhesion assay and affinity chromatography of t he cell surface-labeled integrins, we show in this study that the cons titutive alpha(5) beta(1)-dependent attachment of three leukemic cell lines, BV-173, K562, and Nalm-6, to FN or to a 110-kDa central cell-bi nding fragment of FN was totally inhibited at 37 degrees C by preincub ation of the substrate with either antibodies to the arginine-glycine- aspartic acid-containing region (3Fn-10 module) or to the synergistic region (3Fn-9 module) of FN. Similar results were obtained when assays were carried out at 4 degrees C, suggesting that energy-dependent eve nts were not involved. On the other hand, only the antibody against th e 3Fn-10 module was able to detach most firmly adherent cells. Constit utive cell attachment to the 110-kDa fragment was cation dependent, wi th the order of efficacy of the cation being Mn2+ > Mg2+ > Ca2+, and C a2+ was only effective for BV-173 cells. Antibodies against the alpha( 5) beta(1) integrin or the alpha(5) subunit completely impaired cell a ttachment of all cell lines, whereas several blocking anti-beta 1 subu nit antibodies, including 4B4, P4C10, and AIIB2, differentially pertur bed cell adhesion of BV-173, depending on which cation was present. Th ese anti-beta 1 blocking antibodies, whose epitopes map to a region di stant from the one expected to contain the beta 1 putative cation bind ing sites, seemed to lock the higher activation state of this integrin attained in the presence of Mg2+ and to preserve it during the subseq uent adhesion events irrespectively of the presence of the low avidity state-inducing Ca2+ ion. Because the higher binding avidity displayed by BV-173 could not be explained by a higher degree of preclustering of alpha(5) beta(1) integrin in this cell line compared to K562, these findings suggest that cations might act as allosteric activators of i ntegrin function. Whether this novel cation-dependent parameter of alp ha(5) beta(1) integrin-FN interaction might contribute to the growth c ontrol and/or tissue dissemination of leukemic cells remains to be det ermined.