TNF receptor-associated factor 3 restrains B-cell receptor signaling in normal and malignant B cells.

TRAF3 has diverse signaling functions, which vary by cell type. Uniquely in B lymphocytes, TRAF3 inhibits homeostatic survival. Highlighting the role of TRAF3 as a tumor suppressor, loss-of-function TRAF3 mutations are associated with human B-cell malignancies, while B-cell-specific deletion of TRAF3 in mice leads to autoimmunity and lymphoma development. The role of TRAF3 in inhibiting noncanonical NF-κB activation, CD40 and BAFF-R signaling to B cells is well documented. In contrast, TRAF3 enhances many T-cell effector functions, through associating with and enhancing signaling by the T-cell receptor (TCR)-CD28 complex. The present study was designed to determine the role of TRAF3 in signaling via the B-cell antigen receptor (BCR). The BCR is crucial for antigen recognition, survival, proliferation, and antibody production, and defects in BCR signaling can promote abnormal survival of malignant B cells. Here, we show that TRAF3 is associated with both CD79B and the BCR-activated kinases Syk and Btk following BCR stimulation. BCR-induced phosphorylation of Syk and additional downstream kinases was increased in TRAF3-/- B cells, with regulation observed in both follicular and marginal zone B-cell subsets. BCR stimulation of TRAF3-/- B cells resulted in increased surface expression of MHC-II, CD80, and CD86 molecules. Interestingly, increased survival of TRAF3-/- primary B cells was resistant to inhibition of Btk, while TRAF3-deficient malignant B-cell lines showed enhanced sensitivity. TRAF3 serves to restrain normal and malignant BCR signaling, with important implications for its role in normal B-cell biology and abnormal survival of malignant B cells.

Tcf19 is a novel islet factor necessary for proliferation and survival in the INS-1 ß-cell line.

Recently, a novel type 1 diabetes association locus was identified at human chromosome 6p31.3, and transcription factor 19 (TCF19) is a likely causal gene. Little is known about Tcf19, and we now show that it plays a role in both proliferation and apoptosis in insulinoma cells. Tcf19 is expressed in mouse and human islets, with increasing mRNA expression in nondiabetic obesity. The expression of Tcf19 is correlated with ß-cell mass expansion, suggesting that it may be a transcriptional regulator of ß-cell mass. Increasing proliferation and decreasing apoptotic cell death are two strategies to increase pancreatic ß-cell mass and prevent or delay diabetes. siRNA-mediated knockdown of Tcf19 in the INS-1 insulinoma cell line, a ß-cell model, results in a decrease in proliferation and an increase in apoptosis. There was a significant reduction in the expression of numerous cell cycle genes from the late G1 phase through the M phase, and cells were arrested at the G1/S checkpoint. We also observed increased apoptosis and susceptibility to endoplasmic reticulum (ER) stress after Tcf19 knockdown. There was a reduction in expression of genes important for the maintenance of ER homeostasis (Bip, p58(IPK), Edem1, and calreticulin) and an increase in proapoptotic genes (Bim, Bid, Nix, Gadd34, and Pdia2). Therefore, Tcf19 is necessary for both proliferation and survival and is a novel regulator of these pathways.

Author Correction: TRAF3 regulates the oncogenic proteins Pim2 and c-Myc to restrain survival in normal and malignant B cells.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

TRAF3 regulates the oncogenic proteins Pim2 and c-Myc to restrain survival in normal and malignant B cells.

TRAF3 is a versatile intracellular adapter protein with multiple context-specific roles. Uniquely in B cells, TRAF3 deficiency enhances survival and increases the risk of transformation, as loss of TRAF3 is observed in several types of B cell cancers. Here, we report a new mechanism for TRAF3 in the restraint of B cell survival. We found that TRAF3 deficiency was associated with induction of the pro-survival kinase Pim2 in mouse primary B cells and human malignant B cell lines. The increase in Pim2 was independent of NF-κB2 activation but was ameliorated with inhibition of STAT3 expression or function. TRAF3 deficiency also led to a Pim2-dependent increase in c-Myc protein levels and was associated with reduced c-Myc ubiquitination. TRAF3-deficient primary B cells were less sensitive to cell death induced by the Pim inhibitors SGI-1776 and TP-3654. Interestingly, human malignant B cell lines with low expression of TRAF3 were more sensitive to Pim inhibition-induced cell death. Combination treatment of TRAF3-deficient B cells and B cell tumor lines with c-Myc inhibitors enhanced their sensitivity to Pim inhibition, suggesting a possible therapeutic strategy. TRAF3 thus suppresses a Pim2-mediated B cell survival axis, which can be a potential target for treatment of B cell malignancies.

The oncogenic membrane protein LMP1 sequesters TRAF3 in B-cell lymphoma cells to produce functional TRAF3 deficiency.

Loss-of-function mutations in genes encoding the signaling protein tumor necrosis factor receptor-associated factor 3 (TRAF3) are commonly found in human B-cell malignancies, especially multiple myeloma and B-cell lymphoma (BCL). B-cell TRAF3 deficiency results in enhanced cell survival, elevated activation receptor signaling, and increased activity of certain transcriptional pathways regulating expression of prosurvival proteins. A recent analysis of TRAF3 protein staining of ∼300 human BCL tissue samples revealed that a higher proportion of samples expressing the oncogenic Epstein-Barr virus-encoded protein latent membrane protein 1 (LMP1) showed low/negative TRAF3 staining than predicted. LMP1, a dysregulated mimic of the CD40 receptor, binds TRAF3 more effectively than CD40. We hypothesized that LMP1 may sequester TRAF3, reducing its availability to inhibit prosurvival signaling pathways in the B cell. This hypothesis was addressed via 2 complementary approaches: (1) comparison of TRAF3-regulated activation and survival-related events with relative LMP1 expression in human BCL lines and (2) analysis of the impact upon such events in matched pairs of mouse BCL lines, both parental cells and subclones transfected with inducible LMP1, either wild-type LMP1 or a mutant LMP1 with defective TRAF3 binding. Results from both approaches showed that LMP1-expressing B cells display a phenotype highly similar to that of B cells lacking TRAF3 genes, indicating that LMP1 can render B cells functionally TRAF3 deficient without TRAF3 gene mutations, a finding of significant relevance to selecting pathway-targeted therapies for B-cell malignancies.