Elucidation of mechanism of NK cell activation

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NK cells play important roles in anti-tumor immune response. However, the exact mechanism for their activation still remains unclear. In this study, we have analyzed activation mechanism of NK cells using CD3ζ-deficient mice and a newly developed method for identifying signaling molecules. From the analysis of CD3ζ-deficient mice, we found that CD3ζ negatively regulate the expression and function of FcγRIII on NK cells. Next, we developed a new method using NFAT-GFP transfected reporter cells and unique CD8 chimera cDNA library, in which retrovirally infected reporter cells were crosslinked with anti-CD8 antibody and those become positive for GFP were cloned. cDNAs recovered from these cells included Igα, Igβ, CD3ε, ZAP70, Syk when a library prepared from spleen were used. On the other hand, FcRγ, DAP12, ZAP70 were identified from a library prepared from NK cells. These results show that this method is in fact a powerful method for cloning signaling molecules of lymphocytes. We named this method as NFAT Activating gene Cloning System (NACS). In addition to these already known molecules, we have identified several unknown molecules that activate NFAT-driven transcription upon crosslinking. In future, we will further analyze these newly cloned molecules to exploire the activation mechanism of NK cells.