A novel TRAF6 binding site in MALT1 defines distinct mechanisms of NF-κB activation by API2•MALT1 fusions

The recurrent translocation t(11;18)(q21; q21) associated with mucosa-associated lymphoid tissue (MALT) lymphoma results in the expression of an API2 center dot MALT1 fusion protein that constitutively activates NF-kappa B. The first baculovirus IAP repeat (BIR) domain of API2 and the C terminus of MALT1, which contains its caspase-like domain, are present in all reported fusion variants and interact with TRAF2 and TRAF6, respectively, suggesting their contribution to NF-kappa B signaling by API2 center dot MALT1. Also, the involvement of BCL10 has been suggested via binding to BIR1 of API2 and via its interaction with the immunoglobulin domains of MALT1, present in half of the fusion variants. However, conflicting reports exist concerning their roles in API2 center dot MALT1-induced NF-kappa B signaling. In this report, streptavidin pulldowns of biotinylated API2 center dot MALT1 fusion variants showed that none of the fusion variants interacted with endogenous BCL10; its role in NF-kappa B signaling by API2 center dot MALT1 was further questioned by RNA interference experiments. In contrast, TRAF6 was essential for NF-kappa B activation by all fusion variants, and we identified a novel TRAF6 binding site in the second immunoglobulin domain of MALT1, which enhanced NF-kappa B activation when present in the fusion protein. Furthermore, inclusion of both immunoglobulin domains in API2 center dot MALT1 further enhanced NF-kappa B signaling via intramolecular TRAF6 activation. Finally, binding of TRAF2 to BIR1 contributed to NF-kappa B activation by API2 center dot MALT1, although additional mechanisms involving BIR1-mediated raft association are also important. Taken together, these data reveal distinct mechanisms of NF-kappa B activation by the different API2 center dot MALT1 fusion variants with an essential role for TRAF6.