E3泛素连接酶TRIM44泛素化降解肿瘤转移抑制蛋白CD82诱导食管癌侵袭转移研究

The invasion and metastasis of esophageal carcinoma are the main factors affecting the patient’s prognosis. Abnormal expression of transmembrane 4 superfamily (TM4SF) protein has been shown to participate in a variety of tumor invasion and metastasis, our previous study showed high expression of TM4SF5 and CD81 promote the invasion and metastasis of esophageal cancer (articles have been published), while the TM4SF CD82 was identified as an important tumor metastasis suppressor gene, the low expression of CD82 was reported to be involved in the invasion and metastasis of esophageal cancer. Recently, we tried to elaborate the function of CD82 through scanning the interacting proteins by co-immunoprecipitation combined with mass spectrometry, we found that the E3 ubiquitination TRIM44 is the CD82 interacting protein, which was further supported by the reciprocal co-immunoprecipitation, which indicated that the CD82 may TRIM44 ubiquitinated substrates. Then, the expression of TRIM44 was showed to be increased in esophageal carcinoma by immunohistochemistry. Thus, we speculated that TRIM44 promote esophageal carcinoma invasion and metastasis through by ubiquitinating and degrading tumor metastasis suppressor protein CD82. Here, we intends to firstly demonstrate the relationship between the level of TRIM44 and biological functions of esophageal carcinoma cell, and validate the interaction of TRIM44 and CD82; then try to verify the effect of TRIM44 on the CD82 expression; thirdly, we will detect TRIM44 and CD82 expression in esophageal carcinoma tissues, and analyze the relationship between them and their clinical significance in esophageal carcinoma; finally, the role of TRIM44 in esophageal carcinoma was verified in vivo. Thus, provide a theoretical basis in inhibiting the invasion and metastasis of esophageal carcinoma through the ubiquitin proteasome pathway.

侵袭转移是影响食管癌患者预后的主因。研究证实异常四跨膜蛋白表达参与肿瘤侵袭转移，我们既往发现高四跨膜蛋白TM4SF5与CD81表达促进食管癌侵袭转移（论文已发表），而CD82被视作重要的肿瘤侵袭转移抑制基因，其低表达参与食管癌侵袭转移。近期我们用免疫共沉淀联合质谱研究CD82的相互作用蛋白时，首次发现泛素E3连接酶TRIM44与CD82间存在相互作用，且相互沉淀也证实，这提示CD82可能为TRIM44泛素化底物。进而免疫组化等示TRIM44表达在食管癌中上调。故推测TRIM44通过泛素化降解CD82参与食管癌进展。本课题拟细胞与组织中研究TRIM44表达，细胞水平研究其与食管癌生物学特性关系；细胞中阐明TRIM44与CD82间相互作用及两者表达的关系、前者通过CD82参与食管癌进展；组织中明确TRIM44与CD82表达相关性及与食管癌临床病理和预后关系；最后裸鼠中验证。为临床干预提供新靶点。

本课题通过在细胞中调节TRIM44表达研究其对食管癌侵袭转移影响及机制，并在组织水平结合随访研究其临床意义。TRIM44在食管癌中表达高于相应癌旁组织，其中mRNA（2.42 ± 0.52 vs 0.99 ± 0.25）和蛋白质（1.01 ± 0.27 vs 0.30 ± 0.13），高表达TRIM44的患者表现出分化差（P=1.39×10-5）、TNM分期晚（P=3.87×10-4），而且，组织高表达TRIM44患者预后差（P=2.80×10-5）。细胞实验发现：高表达TRIM44诱导细胞上皮-间质转化增强食管癌细胞的迁移和侵袭。TRIM44显著增强食管癌细胞的增殖且TRIM44与Ki67的表达呈正相关。此外，TRIM44参与AKT/mTOR信号通路及其下游靶点，如STAT3磷酸化。因此，TRIM44的高表达通过AKT/mTOR途径促进HEC的进展，TRIM44可能是HEC患者根治性切除后的一个新的预后指标。