The role of TRAF2 in activation of NF-κB signaling pathway in human B lymphocytes

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-κB signaling pathway in human B cells is unknown. In this study, we investigated the role of TRAF2 in human B cell NF-κB signaling pathways. Methods: Human B cell line RAMOS cells were transfected 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-κB pathway, including IκB kinase activity, phosphorylation and degradation of IκB members and nuclear translocation of NF-κB subunits were detected by kinase assay, Western blot and ELISA assay. Results: Over-expression of WT-TRAF2 induced activation of IκB-kinases (IKKα, IKKi/ε), phosphorylation of IκBα (Ser^{32, 36}) as well as phosphorylation of p65/RelA(Ser⁵³⁶) and nuclear translocation of p65, p50 and c-Rel. However, over-expression of DN-TRAF2 only inhibited nuclear translocation 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-κB signaling pathway, and plays an important role in human B lymphocytes functions.