POSTRECEPTOR OCCUPANCY EVENTS IN LEUKOCYTES DURING BETA-1 INTEGRIN-LIGAND INTERACTIONS

Leukocyte adhesion to and subsequent spreading on the endothelium are the initial steps in the migration of these cells to the surrounding t issues. We have investigated the participation of different VLA hetero dimers in cell spreading by using the anti-beta1 TS2/16 monoclonal ant ibody (mAb) which induces a conformational change of different VLA int egrin receptors, enabling a high-affinity interaction with their ligan ds. Both VLA-4 and VLA-5 fibronectin (FN), as well as VLA-2 collagen ( COL) receptors mediated cell spreading and morphological changes. The spreading of U-937 and alpha4-transfected K-562 cells was induced in b oth FN and VCAM-1, suggesting that the morphological changes may be in duced by cell-cell as well as cell-extracellular matrix (ECM) interact ions. Furthermore, the beta1-regulated cell spreading on VCAM-1 and CO L took place independently of the VLA-5 FN receptor function. The enha ncing effect on cell attachment induced by anti-beta1 TS2/16 mAb was o bserved in the presence of different doses of cytochalasin D, whereas cell spreading was abolished. Signal transduction during beta1-stimula ted integrin-ligand interaction was also investigated. We have found t he co-localization of beta1 integrins and tyrosine-phosphorylated prot eins during the spreading of U-937 and alpha2- and alpha4-transfected K-562 cells on both ECM (FN and COL) and cellular (VCAM-1) ligands. Ki netic studies showed that tyrosine phosphorylation was almost coincide nt with cellular spreading. The tyrosine phosphorylation of polypeptid es of 130 kDa and 77 kDa was triggered in U-937 cells by the interacti on of FN with the VLA-5 receptor in a high-affinity conformation. Howe ver, no signaling was observed by inducing the high-affinity state of receptor in the absence of appropriate ligand. These data suggest that tyrosine kinase activation is a post-receptor occupancy event that mi ght be critical in regulating the adhesive properties of integrins.