中国科学院昆明动物研究所机构知识库

泛素化是一种重要的蛋白质翻译后修饰，能调控细胞的多种生理功能。蛋白质泛素化体系由三个成分构成：E1泛素活化酶、E2泛素结合酶和E3泛素连接酶。E3泛素连接酶负责底物的识别，从而决定了泛素化过程的特异性，主要分为两个亚家族：有HECT结构域的HECT家族和有RING指环结构域的RING家族。该课题所研究的HECTD3是HECT家族的一个E3泛素连接酶，我们实验室前期研究发现HECTD3能够泛素化修饰MALT1，增加MALT1蛋白的稳定性，从而调节肿瘤细胞的耐药和存活。有趣的是，HECTD3的酶活突变体C823A在细胞内也能泛素化修饰MALT1，在体外却不可以。我们猜测细胞内还存在其他的E3泛素连接酶协助HECTD3突变体泛素化修饰MALT1。TRAF6是RING家族的E3泛素连接酶，文献报道TRAF6能够泛素化修饰MALT1，促进NF-κB信号通路的活化。所以，我们假设帮助HECTD3泛素化修饰MALT1的E3泛素连接酶是TRAF6。通过该课题的研究，我们首先发现HECTD3与包括TRAF6在内的多个TRAF有直接的相互作用，且TRAF6的TRAF-C结构域与HECTD3的109-393区段有相互作用；敲低TRAF6能明显减弱HECTD3对MALT1的泛素化修饰，说明TRAF6可能介导HECTD3泛素化修饰MALT1；稳定敲低TRAF6对HeLa肿瘤细胞的生长没有影响，也不能促进肿瘤细胞对化疗药物诱导的凋亡的敏感性。HECTD3和TRAF6协同泛素化修饰MALT1的功能还需要进一步研究。

Ubiquitination is one of the most important protein post-translational modifications, which maintains cell homostasis and regulates almost all cell signaling pathways. The ubiquitination system is compoused of three enzymes, E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme and E3 ubiquitin ligase. E3 ligase is responsible for recognizing substrate proteins and therefore determines the specificity of ubiquitination reaction. E3 ligases are divided into two subfamilies: HECT domain-containing E3s and RING finger domain containing E3s. HECTD3 belongs to HECT E3 ligase subfamily. According to our previous study, HECTD3 can increase MALT1 ubiquitination level, stabilize MALT1 protein and promote cancer cell drug-resistance and survival. Unexpectedly, we found enzyme activity lost mutant of HECTD3(HECTD3 C823A) also could promote MALT1 ubiquitination in vivo, but not in vitro, so we speculated that there might be another E3 ligase which facilitates HECTD3 to ubiquitinate MALT1 in vivo. TRAF6 belongs to RING finger E3 ligase subfamily. TRAF6 can increase MALT1 ubiquitination and activate NF-κB signaling pathway. Therefore, we hypothesize the E3 ligase which facilitates HECTD3 ubiquitination MALT1 in cells is TRAF6. In this study, we found HECTD3 interacts with multi-TRAFs including TRAF6 directly, the TRAF-C domain of TRAF6 and 109-393 domain of HECTD3 are responsible for the interaction. TRAF6 might mediate ubiquitination of MALT1 by HECTD3, depletion of TRAF6 can decrease MALT1 ubiquitination level by HECTD3. On the other hand, depletion of TRAF6 did not affect cancer cell proliferation or sensitivity to chemotherapeutic drugs. Therefore, the functions of ubiquitinated MALT1 by HECTD3 and TRAF6 synergistically need more exploration.