Subcellular localization of MALT1, API2-MALT1, and BCL10 with or without LMB treatment. Subcellular localization of MALT1-flag, flag-API2-MALT1, and myc-BCL10 transiently expressed in COS7 cells (A-F): 2 × 104 COS7 cells were transfected with the following expression vectors: MALT1-flag, 0.2 μg pcDNA3-MALT1-flag without (A) or with 4 ng/mL LMB (B); flag-API2-MALT1, 0.2 μg pcDNA3-flag-API2-MALT1 without (C) or with 4 ng/mL LMB (D); myc-BCL10, a mixture of 0.02 μg pcDNA3-myc-BCL10 and 0.18 μg pcDNA3 without (E) or with 4 ng/mL LMB (F). The LMB treatment was conducted at 18 hours after transfection. At 21.5 hours after transfection, 25 μM MG132 was added only to the wells for MALT1-flag (A-B), and 24 hours after transfection, the cells were examined with an immunofluorescence microscope. Mouse anti-flag antibody was used for flag-tagged constructs; fluorescein goat anti-mouse IgG, as the second antibody for the mouse anti-flag antibody; rabbit anti-human BCL10 antibody, for myc-BCL10; and Alexa Fluor 546 goat anti-rabbit IgG, as the second antibody for the rabbit anti-BCL10 antibody. Nuclei were stained with propidium iodide except for the cells expressing myc-BCL10. Subcellular localization of endogenous MALT1 in COS7 cells (G-H): COS7 cells treated without (G) or with 4 ng/mL LMB (H) for 6 hours were examined. Rabbit anti-MALT1 polyclonal antibody was used as the first antibody and Alexa Fluor 546 goat anti-rabbit IgG as the second antibody for the rabbit anti-MALT1 polyclonal antibody.
