Both BCL10 and MALT1 function as signaling regulators in antigen receptor-mediated NF-κB activation. The activation of NF-κB is often associated with BCL10 phosphorylation, but the physiological significance of BCL10 phosphorylation is still unknown. In this study, we would like to identify the phosphorylation site(s) of BCL10 and try to unravel the myth of BCL10 phosphorylation. Besides, we found that BCL10 could be cleaved when co-expressed with MALT1. Therefore, we would like to locate the MALT1-induced cleavage site of BCL10. BCL10 interacts with several CARD-containing proteins via its N-terminal CARD (caspase recruitment domain), and its C-terminal Serine/Threonine rich domain is probably the target region of phosphorylation. Overexpression of BCL10 induced BCL10 phosphorylation, NF-κB activation and the appearance of filamentous structure under microscope. BCL10L41R, a CARD mutated BCL10, lost all these BCL10-induced abilities. Neither N’BCL10GFP (1~114 a.a.) nor C’BCL10GFP (114~233 a.a.) was phosphorylated, indicating that both CARD and Serine/Threonine rich domain are required for BCL10 phosphorylation. Though not phosphorylated, N’BCL10GFP retained the ability to activate NF-κB, suggesting that BCL10 phosphorylation was not essential for NF-κB activation. The level of BCL10 phosphorylation was diminished by dominant-negative IκB through blocking NF-κB activation. BCL10 deletion constructs：D1(△134~150 a.a.),D2(△185 a.a.),D3(△218~231 a.a.),D4(△134~150 & 218~231 a.a.) and site mutated BCL10 constructs：S7A,S48A,S60,61A,S85A were generated to investigate the phosphorylation site(s) of BCL10. All the mutants maintained abilities to activate NF-κB, form filamentous structure, and be phosphorylated, though the ratio of phosphorylated form to un-phosphorylated form of these constructs were more or less different. These results implied that there were multiple phosphorylation sites on BCL10 molecule. Moreover, the phosphorylation level of BCL10 correlated to its expression level. Endogenous BCL10 was observed to be processed into a shorter molecule in Jurkat cells treated with TPA/ionomycin. MALT1 was reported to be a paracaspase and able to interact with BCL10. We then tested the possibility as whether MALT1 could mediate the cleavage. With the co-expression of individual BCL10 constructs and MALT1, cell lysates were analyzed by western blotting. Except D3 and D4, BCL10 as well as other BCL10 mutants were shown to be cleaved. Therefore, the common deleted region (218~231 a.a.) of D3 and D4 contained the probable cleavage site. Serial BCL10 deletion constructs：C1 (△218~224 a.a.), C2 (△219~225 a.a.), C3 (△220~226 a.a.), C4 (△221~227 a.a.), C5 (△222~228 a.a.), C6 (△223~229 a.a.), C7 (△224~230 a.a.) and C8 (△225~231 a.a.) were created to pinpoint the precise cleavage site. Since C1 was the only one to be cleaved, leucine 225 of BCL10 was predicted to be the best cleavage site candidate. The cleaved products had M.W. larger than authentic BCL10, indicating possible post-translational modification. In fact, CIAP (Calf Intestine Alkaline Phosphatase) treatment relieved the slower migrating property of the cleaved products. The cleaved products were often observed to be accompanied with the diminished level of phosphorylated BCL10. Meanwhile, BCL10L41RGFP and C’BCL10GFP were not good substrates for cleavage. These results supported the idea that phosphorylated BCL10 was a better substrate for MALT1-induced cleavage. MALT1 contains a death domain, two Immunoglobulin like domain and a caspase like domain. To map the domain responsible for the cleavage, five MALT1 constructs：MALT1△N (127~824 a.a.), MALT1 1~548 a.a., MALT1△498~548 a.a., MALT1△199~498 a.a. and IAP2-MALT1 (the fusion protein of IAP2 and MALT1 identified from MALT lymphoma) were tested. While MALT1△N and IAP2-MALT1 retained their abilities as wild-type MALT1, MALT1 1~548 a.a., MALT1△498~548 a.a. and MALT1△199~498 a.a. lost their abilities to induce BCL10 cleavage. Therefore, both CLD and C-terminal region of MALT1 were required for mediating the cleavage.