Tumor necrosis factor receptor-associated factor (TRAF) 1 regulates CD40-induced TRAF2-mediated NF-kappaB activation.
To investigate CD40 signaling complex formation in living cells, we used green fluorescent protein (GFP)-tagged CD40 signaling intermediates and confocal life imaging. The majority of cytoplasmic TRAF2-GFP and, to a lesser extent, TRAF3-GFP, but not TRAF1-GFP or TRAF4-GFP, translocated into CD40 signaling complexes within a few minutes after CD40 triggering with the CD40 ligand. The inhibitor of apoptosis proteins cIAP1 and cIAP2 were also recruited by TRAF2 to sites of CD40 signaling. An excess of TRAF2 allowed recruitment of TRAF1-GFP to sites of CD40 signaling, whereas an excess of TRAF1 abrogated the interaction of TRAF2 and CD40. Overexpression of TRAF1, however, had no effect on the interaction of TRADD and TRAF2, known to be important for tumor necrosis factor receptor 1 (TNF-R1)-mediated NF-kappaB activation. Accordingly, TRAF1 inhibited CD40-dependent but not TNF-R1-dependent NF-kappaB activation. Moreover, down-regulation of TRAF1 with small interfering RNAs enhanced CD40/CD40 ligand-induced NF-kappaB activation but showed no effect on TNF signaling. Because of the trimeric organization of TRAF proteins, we propose that the stoichiometry of TRAF1-TRAF2 heteromeric complexes ((TRAF2)2-TRAF1 versus TRAF2-(TRAF1)2) determines their capability to mediate CD40 signaling but has no major effect on TNF signaling.
Fotin-Mleczek
Mariola
M
Institute of Cell Biology and Immunology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
Henkler
Frank
F
Hausser
Angelika
A
Glauner
Heike
H
Samel
Dierk
D
Graness
Angela
A
Scheurich
Peter
P
Mauri
Davide
D
Wajant
Harald
H
D019013:CD40 Antigens / Q000502:physiology*; D023201:CD40 Ligand / Q000502:physiology; D002460:Cell Line; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D018124:Receptors, Tumor Necrosis Factor / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D047990:TNF Receptor-Associated Factor 1; D047992:TNF Receptor-Associated Factor 2; D048008:TNF Receptor-Associated Factor 3; D014162:Transfection
D019013:CD40 Antigens; D008164:Luminescent Proteins; D016328:NF-kappa B; D011506:Proteins; D034741:RNA, Small Interfering; D018124:Receptors, Tumor Necrosis Factor; D011993:Recombinant Fusion Proteins; D047990:TNF Receptor-Associated Factor 1; D047992:TNF Receptor-Associated Factor 2; D048008:TNF Receptor-Associated Factor 3; D023201:CD40 Ligand; D049452:Green Fluorescent Proteins