Constitutive expression of MALT1 in lipid rafts of human B-cell lymphoma cells. (A) Western blotting shows MALT1, as well as CARMA1 and BCL10, to be located in both lipid raft (R) and soluble fractions (S) in DLBCL cell lines separated by sucrose gradient centrifugation. Arrowheads mark the band of the expected full-length size for each protein. The origin of the smaller band on the MALT1 blots, not observed in blots from BJAB cells, is uncertain. The multiple bands of BCL10 represent phosphorylation isoforms as they disappear after phosphatase treatment (data not shown). The partitioning of LYN, a B-cell-specific protein of the Src protein kinase family known to be constitutively present in rafts, confirms effective separation of raft and nonraft protein fractions. (B) Structure of C-terminal myc-tagged MALT1 and API2-MALT1 transgenes and pMSCV retroviral vectors used in the study. Arrowheads show positions of the most common breakpoints found in t(11;18) that give rise to the API2-MALT1 fusion gene, which is used for all of the following experiments. (C) Lysates from equivalent cell numbers of pools of BJAB cells transfected with API2-MALT1 or MALT1 were immunoblotted for myc, MALT1, and BCL10. The exogenous API2-MALT1 and MALT1 are detected by anti-myc and with the MALT1 antibody (^*). (D) Lipid raft and soluble fractions of lysates from pools of MALT1- and API2-MALT1-expressing BJAB cells were immunoblotted for myc and LYN. LTR indicates long-term repeat; BIR, baculovirus inhibitor of apoptosis repeat; DD, death domain; RING, RING domain; and IRES, internal ribosomal entry site.