Gain-of-function mutations in CARD11 promote enhanced aggregation and idiosyncratic signalosome assembly

•Gain-of-function CARD11 variants form shell-like structures (mCADS) in lymphocytes.•mCADS constitutively signal via sustained MALT1 and phospho-IKK association.•Wild type CARD11 cannot form these structures upon antigen receptor stimulation.•mCADS require BCL10 and MAGUK domain interactions for assembly.•Endogenous mCADS are detectable in DLBCL harboring similar CARD11 GOF mutations. BENTA (B cell Expansion with NF-κB and T cell Anergy) is a novel lymphoproliferative disorder caused by germline, gain-of-function (GOF) mutations in the lymphocyte-restricted scaffolding protein CARD11. Similar somatic CARD11 mutations are found in lymphoid malignancies such as diffuse large B cell lymphoma (DLBCL). Normally, antigen receptor (AgR) engagement converts CARD11 into an active conformation that nucleates a signalosome required for IκB kinase (IKK) activation and NF-κB nuclear translocation. However, GOF CARD11 mutants drive constitutive NF-κB activity without AgR stimulation. Here we show that unlike wild-type CARD11, GOF CARD11 mutants can form large, peculiar cytosolic protein aggregates we term mCADS (mutant CARD11 dependent shells). MALT1 and phospho-IKK are reliably colocalized with mCADS, indicative of active signaling. Moreover, endogenous mCADS are detectable in ABC-DLBCL lines harboring similar GOF CARD11 mutations. The unique aggregation potential of GOF CARD11 mutants may represent a novel therapeutic target for treating BENTA or DLBCL.