TRAF2在活化B淋巴细胞NF-kB信号传导系统中的作用

The role of TRAF2 in activation of NF-kB signaling pathway in hmnan B lymphocytes

目的肿瘤坏死因子受体相关因子2（TNF receptor associated factor2，TRAF2）是介导B淋巴细胞表面CD40信号传导的重要因子。目前对TRAF2如何介导CD40核转录因子（NF-kB）信号系统各环节下传还了解不多。本研究利用人B细胞株模型对这方面进行了初步探讨。方法将人类B淋巴细胞株RAMOS细胞转染融合有黄色荧光素（YFP）的野生型（WT-TRAF2）或功能缺失型（DN-TRAF2）TRAF2质粒，过夜培养后用流式细胞仪分离YFP阳性细胞，然后通过激酶实验、Western blot以及ELISA等方法对NF-kB通路的活化进行包括IkB激酶、IkB磷酸化和降解以及NF-kB的细胞核内转录等多方面的研究。结果过度表达WT-TRAF2可诱导IkB激酶（IKKα和IKXi／g）对IKBα（Ser^32.36）的磷酸化和降解以及对p65／RelA的磷酸化和p65、p60、c-Rel的核内转录。然而，过度表达DN-TRAF2只抑制p65的核内转录，而不抑制p50和c-Rel。TRAF2也可通过诱导RAMOS B细胞分泌IgM 而影响其功能。结论TRAF2可选择性地激活人B淋巴细胞CD40介导的NF-kB信号传导系统中的重要因子，在B细胞活化分化中起重要作用。

Objective TRAF2 （TNF receptor associated factor 2） is an essential adaptor for many members of the TNF receptor family, including CD40. The precise mechanism of how TRAF2 mediates CD40 induced NF-kB signaling pathway in human B cells is unknown. In this study, we investigated the role of TRAF2 in human B cell NF-kB signaling pathways. Methods Human B cell line RAMOS cells were tmnsfected with plasmids expressing YFP fusion TRAF2 （wild type TRAF2, WT-TRAF2 or dominant-negative TRAF2, DN-TRAF2）. After transfection, the cells were cultured overnight, and positive cells were sorted by YFP. Activation of NF-kB path-way, including IkB kinase activity, phosphorylation and degradation of IkB members and nuclear tmnslocation of NF-kB subunits were detected by kinase assay, Western blot and ELISA assay. Results Over-expression of WT-TRAF2 induced activation of IkB-kinases （IKKα, IKKi∈）, phosphorylation of Ika （Ser^32.36 ） as well as phospho-rylation of p65/RelA（Ser^536） and nuclear translocation of p65, p50 and c-Rel. However, over-expression of DN-TRAF2 only inhibited nuclear transloeation of p50 and c-Rel. TRAF2 also affected the function of B cells by inducing excretion of IgM. Conclusion TRAF2 selectively activates some important factors in CD40 mediated NF-kB signaling pathway, and plays an important role in human B lymphocytes functions.