Molecular mechanisms of immune cell trafficking

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To elucidate the regulatory mechanisms of integrin adhesion receptors by the small GTPase Rap1 and its effector molecule RAPL, we isolated Mst1, which belongs to the ste20-like kinase family. Mst1 was associated with and activated by RAPL through its coiled-coil region. Mst1 kinase activity was stimulated by activated Rap1 and also by chemokines and TCR ligation. However, in RAPL-deficient lymphocytes, Mst1 was hardly activated by chemokines and TCR ligation, indicating that Mst1 is activated by these stimulation in a RAPL-dependent manner. Overexpression of Mst1 led to development of the leading edge and uropod, and LFA-1 clustering in the leading edge, resulting in increased adhesion by LFA-1, all of which required the Mst1 kinase activity. Conversely, Mst1 knockdown by shRNA impaired cell polarity development and augmentation of adhesion by chemokines and TCR ligation. These results indicate that Mst1 is a critical downstream protein kinase responsible for cell polarity development … More and LFA-1 regulation triggered by chemokines and the TCR. In RAPL-deficient mice, the T and B lymphocyte numbers in peripheral lymph nodes was reduced due to defective lymphocyte homing. In vitro flow adhesion assays with HEV-like endothelial cells and intravital microscopic analysis of lymphocyte trafficking indicate that Rap1 plays a critical role in arrest from rolling, and RAPL is required for firm adhesion by LFA-1 and α4β7. The β2 and αL cytoplasmic domains is involved in arrest and firm adhesion, respectively. Furthermore, intravital two-photon microscopy revealed that RAPL-deficient T and B cells moved at slower velocities with reduced displacement within lymph nodes. Taken together, these studies demonstrate that the Rap1-RAPL signaling regulates lymphocyte cell polarity and LFA-1 activity through Mst1 and control lymphocyte adhesion to endothelial cells and interstitial migration within lymph nodes in vivo. Based on these functions, we are currently examining lymphocyte growth and differentiation in mice genetically engineered on RAPL and Mst1 loci in order to clarify the relationship of lymphocyte adhesion with cell growth/differentiation and immune diseases. Less