Competition between TRAF2 and TRAF6 regulates NF-kappaB activation in human B lymphocytes

Chin Med Sci J. 2010 Mar;25(1):1-12. doi: 10.1016/s1001-9294(10)60013-2.


Objective: To investigate the role of TNF receptor-associated factor 2 (TRAF-2) and TRAF6 in CD40-induced nuclear factor-kappaB (NF-kappaB) signaling pathway and whether CD40 signaling requires TRAF2.

Methods: Human B cell lines were transfected with plasmids expressing wild type TRAF2 or dominant negative TRAF2, TRAF2-shRNA, or TRAF6-shRNA. The activation of NF-kappaB was detected by Western blot, kinase assay, transfactor enzyme-linked immunosorbent assay (ELISA), and fluorescence resonance energy transfer (FRET). Analysis of the role of TRAF-2 and TRAF-6 in CD40-mediated NF-kappaB activity was examined following stimulation with recombinant CD154.

Results: TRAF2 induced activity of IkappaB-kinases (IKKalpha, IKKi/epsilon), phosphorylation of IkappaBalpha, as well as nuclear translocation and phosphorylation of p65/RelA. In contrast, TRAF6 strongly induced NF-kappaB activation and nuclear translocation of p65 as well as p50 and c-Rel. Engagement of CD154-induced nuclear translocation of p65 was inhibited by a TRAF6-shRNA, but conversely was enhanced by a TRAF2-shRNA. Examination of direct interactions between CD40 and TRAFs by FRET documented that both TRAF2 and TRAF6 directly interacted with CD40. However, the two TRAFs competed for CD40 binding.

Conclusions: These results indicate that TRAF2 can signal in human B cells, but it is not essential for CD40-mediated NF-kappaB activation. Moreover, TRAF2 can compete with TRAF6 for CD40 binding, and thereby limit the capacity of CD40 engagement to induce NF-kappaB activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • B-Lymphocytes / cytology
  • B-Lymphocytes / physiology*
  • CD40 Antigens / metabolism
  • Cell Line
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Fluorescence Resonance Energy Transfer
  • Humans
  • I-kappa B Kinase / metabolism
  • NF-kappa B / genetics
  • NF-kappa B / metabolism*
  • Proto-Oncogene Proteins c-fos / metabolism
  • Signal Transduction / physiology*
  • TNF Receptor-Associated Factor 2 / genetics
  • TNF Receptor-Associated Factor 2 / metabolism*
  • TNF Receptor-Associated Factor 6 / genetics
  • TNF Receptor-Associated Factor 6 / metabolism*
  • Transcription Factor RelA / metabolism
  • p38 Mitogen-Activated Protein Kinases / metabolism


  • CD40 Antigens
  • NF-kappa B
  • Proto-Oncogene Proteins c-fos
  • TNF Receptor-Associated Factor 2
  • TNF Receptor-Associated Factor 6
  • Transcription Factor RelA
  • I-kappa B Kinase
  • Extracellular Signal-Regulated MAP Kinases
  • p38 Mitogen-Activated Protein Kinases