期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 289, 期 10, 页码 6969-6977出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M113.498253
关键词
Cell Biology; Cell Junctions; ERK; Natural Killer (NK) Cell; Phosphorylation
资金
- Ministry of Science and Technology of China (973 Basic Science Project) [2010CB911901, 2013CB530506]
- Natural Science Foundation of China [91029303, 31021061]
Background: CD226 is an activating receptor on NK cells that mediates NK cell cytotoxicity. Results: The first extracellular domain of CD226 (CD226-ECD1) mediates NK cell recognition, adhesion, immune synapse formation, and cytotoxicity against target cells. Conclusion: CD226-ECD1 retains almost all functions of the full-length CD226 protein. Significance: The conclusion is helpful to understand the mechanism by which CD226 recognizes its ligands. CD226, an activating receptor that interacts with the ligands CD155 and CD112, activates natural killer (NK) cells via its immunoreceptor tyrosine-based activatory motif (ITAM). There are two extracellular domains of CD226; however, the comparative functional relevance of these domains remains unknown. In this study, two different deletion mutants, rCD226-ECD1 (the first extracellular domain) and rCD226-ECD (full extracellular domains), were recombinantly expressed. We observed that rCD226-ECD1, similar to rCD226-ECD, specifically bound to ligand-positive cell lines and that this interaction could be competitively blocked by an anti-CD226 mAb. In addition, rCD226-ECD1 was able to block the binding of CD112 mAb to tumor cells in a competitive binding assay. Importantly, based on surface plasmon resonance (SPR), we determined that rCD226-ECD1, similar to rCD226-ECD, directly bound to its ligand CD155 on a protein chip. Functionally, NK cell cytotoxicity against K562 or HeLa cells was blocked by rCD226-ECD1 by reducing the expression of CD69 and granzyme B, indicating the critical role of ECD1 in NK cell activation. We also examined the role of rCD226-ECD1 in effector/target interactions by using rCD226-ECD to block these interactions. Using flow cytometry, we found that the number of conjugates between IL-2-dependent NKL cells and HeLa cells was reduced and observed that the formation of immune synapses was also decreased under confocal microscopy. In addition, we prepared two anti-rCD226-ECD1 agonistic antibodies, 2E6 and 3B9. Both 2E6 and 3B9 antibodies could induce the phosphorylation of ERK in NK-92 cells. Taken together, our results show that CD226 functions via its first extracellular domain.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据