Early Lethality, Functional NF-κB Activation, and Increased Sensitivity to TNF-Induced Cell Death in TRAF2-Deficient Mice

Amgen Institute, Department of Medical Biophysics, University of Toronto, Ontario, Canada.
Immunity (Impact Factor: 21.56). 12/1997; 7(5):715-25. DOI: 10.1016/S1074-7613(00)80391-X
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TRAF2 is an intracellular signal-transducing protein recruited to the TNFR1 and TNFR2 receptors following TNF stimulation. To investigate the physiological role of TRAF2, we generated TRAF2-deficient mice. traf2-/- mice appeared normal at birth but became progressively runted and died prematurely. Atrophy of the thymus and spleen and depletion of B cell precursors also were observed. Thymocytes and other hematopoietic progenitors were highly sensitive to TNF-induced cell death and serum TNF levels were elevated in these TRAF2-deficient animals. Examination of traf2-/- cells revealed a severe reduction in TNF-mediated JNK/SAPK activation but a mild effect on NF-kappaB activation. These results suggest that TRAF2-independent pathways of NF-kappaB activation exist and that TRAF2 is required for an NF-kappaB-independent signal that protects against TNF-induced apoptosis.

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    • "Cells (African green monkey kidney epithelioid [BS-C-1] cells, human cervix carcinoma epithelioid [HeLa] cells, and MEFs) were grown in Dulbecco's modified Eagle's medium (DMEM; Life Technologies) containing 50 IU/ml penicillin, 50 μg/ml streptomycin (Sigma), and 10% fetal bovine serum (FBS; Life Technologies). TRAF2+/+ and TRAF2−/− MEFs (42) were sent by Søren R. Paludan (University of Aarhus, Aarhus, Denmark) with the permission of Tak W. Mak (Campbell Family Institute for Breast Cancer Research, Canada). Cells were incubated at 37°C in a 5% CO2 incubator. "
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    ABSTRACT: Tumor necrosis factor receptor (TNFR)-associated factor 2 (TRAF2) is a pivotal intracellular mediator of signaling pathways downstream of TNFR1 and -2 with known pro- and antiviral effects. We investigated its role in the replication of the prototype poxvirus vaccinia virus (VACV). Loss of TRAF2 expression, either through small interfering RNA treatment of HeLa cells or through genetic knockout in murine embryonic fibroblasts (MEFs), led to significant reductions in VACV growth following low-multiplicity infection. In single-cycle infections, there was delayed production of both early and late VACV proteins as well as accelerated virus-induced alterations to cell morphology, indicating that TRAF2 influences early stages of virus replication. Consistent with an early role, uncoating assays showed normal virus attachment but delayed virus entry in the absence of TRAF2. Although alterations to c-Jun N-terminal kinase (JNK) signaling were apparent in VACV-infected TRAF2−/− MEFs, treatment of wild-type cells with a JNK inhibitor did not affect virus entry. Instead, treatment with an inhibitor of endosomal acidification greatly reduced virus entry into TRAF2−/− MEFs, suggesting that VACV is reliant on the endosomal route of entry in the absence of TRAF2. Thus, TRAF2 is a proviral factor for VACV that plays a role in promoting efficient viral entry, most likely via the plasma membrane. IMPORTANCE Tumor necrosis factor receptor-associated factors (TRAFs) are key facilitators of intracellular signaling with roles in innate and adaptive immunity and stress responses. We have discovered that TRAF2 is a proviral factor in vaccinia virus replication in both HeLa cells and mouse embryonic fibroblasts and that its influence is exercised through promotion of efficient virus entry.
    Journal of Virology 01/2014; 88(7). DOI:10.1128/JVI.03013-13 · 4.44 Impact Factor
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    • "TRAF2 and TRAF3 play co-operative, but distinct roles in facilitating both the constitutive suppression of NF-κB2 signaling and the BAFFR-mediated activation of this pathway, the molecular details of which are described above. Mice completely lacking either TRAF are perinatally lethal (99, 100). Thus the use of a B cell specific deletion system to investigate their role in B cells has been important in furthering our understanding of these molecules. "
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    ABSTRACT: It has been more than a decade since it was recognized that the nuclear factor of kappa light polypeptide gene enhancer in B cells (NF-κB) transcription factor family was activated by two distinct pathways: the canonical pathway involving NF-κB1 and the non-canonical pathway involving NF-κB2. During this time a great deal of evidence has been amassed on the ligands and receptors that activate these pathways, the cytoplasmic adapter molecules involved in transducing the signals from receptors to nucleus, and the resulting physiological outcomes within body tissues. In contrast to NF-κB1 signaling, which can be activated by a wide variety of receptors, the NF-κB2 pathway is typically only activated by a subset of receptor and ligand pairs belonging to the tumor necrosis factor (TNF) family. Amongst these is B cell activating factor of the TNF family (BAFF) and its receptor BAFFR. Whilst BAFF is produced by many cell types throughout the body, BAFFR expression appears to be restricted to the hematopoietic lineage and B cells in particular. For this reason, the main physiological outcomes of BAFF mediated NF-κB2 activation are confined to B cells. Indeed BAFF mediated NF-κB2 signaling contributes to peripheral B cell survival and maturation as well as playing a role in antibody responses and long term maintenance plasma cells. Thus the importance BAFF and NF-κB2 permeates the entire B cell lifespan and impacts on this important component of the immune system in a variety of ways.
    Frontiers in Immunology 01/2014; 4:509. DOI:10.3389/fimmu.2013.00509
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    • "Previous studies have shown that NF-κB is a powerful transcription factor that blocks apoptosis [15]. TNF-α can induce NF-κB transactivation via IκB kinase (IKK) complex phosp orylation, which lead to degradation of IκBs and the consequent translocation of NF-κB to nucleus [9,10,16,17]. We examined TNF-α mediated NF-κB transactivation in Hep3B and SMMC-7721 cells. "
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    ABSTRACT: Tumor microenviroment is characteristic of inflammation, ischemia and starvation of nutrient. TNF-alpha, which is an extraordinarily pleiotropic cytokine, could be an endogenous tumor promoter in some tumor types. The basic objective of this study was to investigate the effects of TNF-alpha on the cell viability and apoptosis of hepatocellular carcinoma cells under serum starvation, and to identify the molecular mechanisms involved. For this purpose, five different concentrations of TNF-alpha and two different serum settings (serum-cultured and serum-deprived) were used to investigate the effects of TNF-alpha on the cell viability and apoptosis of Hep3B and SMMC-7721 cells. TNF-alpha (10 ng/ml) attenuated serum starvation-induced apoptosis of hepatocellular carcinoma cells, and autophagy conferred this process. BAY11-7082, a specific inhibitor of NF-kappaB, reversed the suppression of serum starvation-induced apoptosis by TNF-alpha. Moreover, TNF-alpha-induced NF-kappaB transactivation was suppressed by autophagy inhibitor 3-MA. In addition, TNF-alpha up-regulated Ferritin heavy chain (FHC) transiently by NF-kappaB activation and FHC levels were correlated with the TNF-alpha-induced protection against serum starvation-mediated apoptosis of hepatocellular carcinoma cells. Furthermore, FHC-mediated inhibition of apoptosis depended on suppressing ROS accumulation. Our findings suggested that autophagy conferred the TNF-alpha protection against serum starvation-mediated apoptosis of hepatocellular carcinoma cells, the mechanism involved with the activation of the TNF-alpha/ NF-kappaB /FHC signaling pathway.
    BMC Cancer 09/2013; 13(1):438. DOI:10.1186/1471-2407-13-438 · 3.36 Impact Factor
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