Study on molecular mechanisms of integrin-regulated adhesion in immune responses

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We have reported that the small GTPase Rap1 is a intracellular signaling molecule that increase integrin adhesiveness that is triggered by stimulation with chemokines or antigens. Rap1 activation by chemokines robustly stimulates lymphocyte motility. We have also demonstrated that Rap1 activation by chemokines is required in lymphocyte transmigration through endothelial monolayers under physiological shear flow. Furthermore, Rap1 activation induces cell polarization, which is accompanied with development of the leading edge and uropod and relocalization of chemokine receptor CXCR4 and CD44 to the leading edge and uropod, respectively. In order to clarigy the molecular mechanism in these processes, we have identified and characterized a novel Rap1 effector RAPL. RAPL is a Rap1-GTP binding 30 kD protein isolated by yeast two-hybrid screening with an active Rap1 mutant. RAPL is preferentially expressed in spleen, lymph nodes, and thymus, in particular T and B cells. RAPL overexpression stimulated LFA-1-mediated adhesion to ICAM-1, whereas a N-terminal deletion mutant acted as a dominant negative mutant and inhibited adhesion to ICAM-1 triggered by TCR and chemokines. RAPL augment ligand-binding affinity of LFA-1 and lymphocyte polarization. Upon stimulation with chemokines, RAPL forms a complex with LFA-1 and translocates to the leading edge, depending on Rap1 activation. Taken together, these results indicate that when RAPL binds to Rap1-GTP, it induce cell polarization and translocates LFA-1, resulting in an increase of adhesiveness through modulation of affinity and clustering of LFA-1.