Mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1), an intracellular signaling molecule, is widely expressed during inflammatory responses. To investigate the immune function of AjMALT1 in Apostichopus japonicus, the full length of AjMALT1 gene was cloned using transcriptome data and RACE technology. The results showed that AjMALT1 was distributed in all tissues, with higher expression found in coelomocytes and intestine. The expression of AjMALT1 was significantly upregulated in Vibrio splendidus-challenged sea cucumbers, as well as in coelomocytes exposed to inactive V. splendidus, and was positively correlated with the expression of the pro-inflammatory cytokine AjIL17 and the inflammasome component AjNLRP3. Further investigation using specific siRNA to silence AjMALT1 for 48 h revealed that the expression of AjIL17 and AjNLRP3 was reduced under V. splendidus stimulation. Additionally, histological observations showed a decrease in intestinal inflammation. Interference with AjMALT1 also led to downregulation of AjTRAF6 and AjRel expression, as well as inhibited nuclear translocation of AjRel. These findings suggest AjMALT1 exacerbates intestinal and coelomic inflammation by activating the AjTRAF6-dependent NF-κB pathway in A. japonicus.