EZH2 Regulates The Development And Functionof NK Cells Via Repressing NKG2D

Objiective: Explore the mechanisms of histone methyltransferases EZH2 regulating the development and function of NK cell.Methods: Part I: HSCs were isolated from C57BL6 mice, cultured with cytokines and EZH2 inhibitors, western blotting analysis to detect the level of H3K27me3 following the treatment of EZH2 inhibitors. At indicated time points, cells were harvested and analyzed for NK development with surface antigen staining and flow cytometry. h UCB CD34+ HSCs were enriched, cultured in the existence of cytokines and EZH2 inhibitors, western blotting analysis to detect the level of H3K27me3 following the treatment with EZH2 inhibitors. At indicated time points, cells were harvested and analyzed for NK development with surface antigen staining and flow cytometry.Part II: The expression level of NKG2 D in NK cells cultured in vitro was detected by flow cytometry at indicated time points. Ch IP-q PCR analysis of in vitro cultured h UCB CD34+ HSC in the presence of cytokines. The cytotoxic ability of NK cells against target cells was analyzed, to analysis the effect of EZH2 in NK function.Results: Part I: When culture in vitro differentiation, NK cells population was specifically increased after cells were treated by EZH2 inhibitors.Part II: The Expression level of NKG2 D from in vitro derived mouse NK cells and human NK cells was significantly increased after EZH2 inhibitors. Ch IP-q PCR analysis showed that the expression of NKG2 D was regulated by H3K27me3. The killing assay results show that the killing ability of NK cells was significantly increased after cells were treated by EZH2 inhibitors, and it closely related with the expression of NKG2 D.Conclusion: These results imply a novel mechanism that EZH2 could regulate the development and function of NK cells via repressing NKG2 D. EZH2 inhibitors treatment enhances both mouse and human NK cells development in vitro. EZH2 inhibitors treatments both enhance NKG2 D expression on NK cell. NKG2 D expression could be regulated by EZH2 mediated H3K27me3 modification on its gene loci and is essential for EZH2 inhibition improved NK cells development. in vitro treatments by EZH2 inhibitors enhance the NKG2D-dependent killing ability of NK cells.