Anti-MALT1 monoclonal antibody, clone NU2/521

Background: The B cell antigen receptor (BCR) and pathogen recognition receptors, such as Toll-like receptor 4 (TLR4), act in concert to control adaptive B cell responses. However, little is known about the signaling pathways that integrate BCR activation with intrinsic TLR4 stimulation. Antigen receptors initialize activation of the inducible transcription factor nuclear factor-kappa B (NF-kappa B) via recruitment of the membrane-associated guanylate kinase caspase recruitment domain protein 11 (CARD11), the adapter molecule B cell CLL/lymphoma 10 (BCL10), and the "paracaspase" mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1) into lipid rafts. Upon BCR triggering, this activation strictly depends on BCL10, but not on MALT1, leading to the hypothesis that a MALT1-independent NF-kappa B activation pathway contributes to BCR-induced NF-kappa B activation downstream of BCL10. The identity of this pathway has remained elusive. Results: Using genetic and biochemical approaches, we demonstrate that the IRAK4- and IRAK1-dependent TLR signaling branch is activated upon BCR triggering to induce partial NF-kappa B activation. BCR-induced MALT1-independent I kappa B degradation and B cell proliferation were inhibited in MALT1/IRAK4 double knockout B cells. Moreover, IRAK1 was recruited into lipid rafts upon BCR stimulation and activated following transient recruitment of IRAK4. Conclusion: We propose that the observed crosstalk between BCR and TLR signaling components may contribute to the discrimination of signals that emanate from single and dual receptor engagement to control adaptive B cell responses.

The MALT1 (Mucosa associated lymphoid tissue lymphoma translocation protein 1) paracaspase couples antigen receptors on lymphocytes to downstream signaling events. Activation of MALT1 is known to involve stimulus-dependent CBM complex formation, that is, the recruitment of BCL10-bound MALT1 to a CARD-Coiled Coil protein. Beyond this canonical, CBM-dependent mechanism of MALT1 activation, recent studies suggest that MALT1 protease activity may be triggered by alternative mechanisms. For instance, the E3-ligase TRAF6 can activate MALT1 proteolytic function and induce MALT1 auto-cleavage. However, the interplay between CBM and TRAF6 with regard to MALT1 activation has remained incompletely elucidated. Here, by generating CRISPR/Cas9-derived knock-out Jurkat T-cells, we show that TRAF6 was dispensable for CARD11/BCL10-dependent MALT1 activation upon T-cell stimulation. However, ectopically-expressed TRAF6 could induce MALT1 activity in Jurkat T-cells devoid of either CARD11 or BCL10. These data provide unequivocal evidence that TRAF6-mediated MALT1 activation does not require the upstream scaffold CARD11 or the interaction between MALT1 and BCL10. Thus, TRAF6 may be part of a previously unidentified non-canonical pathway that triggers MALT1 protease activity independently of canonical CBM signalosomes. (C) 2018 Elsevier Inc. All rights reserved.