Abstract: Purpose To investigate the role of TRAF6 in NF- B and AP-1 signaling pathway in human B cell line. Methods Human Ramos B cells were transfected with plasmids expressing YFP fusion dominant-negative TRAF6 (DN-TRAF6), or transfected with shRNA-TRAF6 plasmid. After cultured overnight, cells were either sorted with flowcytometry or screened by G418. Activation of NF- B and AP-1 pathway, including phosphorylation of I B , ERK, JNK and P38, as well as nuclear translocation of NF- B subunits (P65, P50 and c-Rel)and AP-1 subunits(c-Fos, c-Jun, CREB and ATF) were detected by Western blot and ELISA. Results In cells which overexpress DN-TRAF6 or endogenous TRAF6 expression were knocked-down by shRNA, the phosphorylation of I B , as well as phosphorylation of JNK were inhibited. Furthermore, nuclear translocation of NF- B subunits P65, P50, c-Rel,and AP-1 subunits c-fos and c-jun were also inhibited in B cells overexpression of DN-TRAF6. Conclusion TRAF6 selectively activates some kinases in CD40 mediated NF- B and AP-1 signaling pathway, and plays an important role in their activation.