Molecular mechanisms of TRAF6 function in signaling pathways of the oncogenic viral mimic of CD40, LMP1

Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) plays important roles in EBV-mediated B cell transformation, development of EBV-associated malignancies, and exacerbation of certain autoimmune conditions. LMP1 functionally mimics tumor necrosis factor receptor (TNFR) superfamily member CD40, but LMP1 signals are amplified and sustained compared to those induced by CD40. CD40 and LMP1 rely on TNFR-associated factors (TRAFs) to mediate signaling, but use TRAFs differently. TRAF6 is important for CD40 signaling, and was implicated in LMP1 signaling in non-B cells. Here, we addressed the hypothesis that TRAF6 is a critical regulator of a subset of LMP1 signals in B cells. We found that TRAF6 was required for LMP1-mediated kinase activation and costimulatory molecule upregulation, and associated with the LMP1 TRAF1/2/3/5 binding site (TBS). Additionally, TRAF6 and the TBS contributed to LMP1-induced autoreactivity and antibody (Ab) production in vivo. Finally, in contrast to CD40, LMP1 required the TRAF6 receptor-binding domain to mediate TRAF6-dependent pathways. Thus, TRAF6 is critical for LMP1 signaling and requires LMP1 interaction to propagate signals. Importantly, TRAF6 associates with LMP1 in a manner distinct from CD40, raising the possibility of disrupting LMP1 functions while leaving normal CD40 signaling intact. We next investigated roles of the kinase TAK1 in TRAF6-dependent LMP1 functions. TAK1 was required for CD40and LMP1-mediated JNK activation in B cells, leading to IL-6 and Ab production. Understanding mechanisms of CD40 and LMP1 signaling provides important insights into normal regulatory control of CD40 functions and how LMP1-mediated pathogenesis escapes or subverts these regulatory mechanisms. LMP1 itself may be a difficult therapeutic target, because it lacks an extracellular domain and is continually