Rock2促进p53非泛素化降解调控肝癌EMT的机制研究

研究证明肿瘤抑制基因p53缺失或突变与包括肝癌在内肿瘤的EMT发生密切相关。但值得关注的是，约50%的肿瘤p53并无缺失或突变，其仍能发生EMT以利于侵袭转移，具体机制尚不清楚。我们研究已证明Rock2和FAT10在肝癌中高表达并促进其侵袭转移。前期我们通过人肝癌细胞裸鼠成瘤模型、基因芯片和分子生物学等实验发现降低Rock2：①p53+/+肝癌细胞肺部转移明显减少，EMT相关分子标记物明显改变；FAT10表达降低，p53泛素化水平升高但其蛋白表达却增加；敲除FAT10后Rock2则对p53泛素化和表达无影响；②p53-/-肝癌细胞中EMT和肺部转移却无明显变化。因此，我们提出Rock2经FAT10促进p53非泛素化降解从而调控肝癌EMT的科学假说。为验证假说，本课题拟进一步采用动物活体成像、基因敲除、免疫共沉淀等技术从动物、细胞和分子等水平探讨其具体机制，为肝癌侵袭转移的防治提供理论依据。

It has been proved that the deletion or mutation of tumor suppressor gene p53 is closely related to tumor cell EMT, including hepatocellular carcinoma (HCC). However, it should be noted that about 50% of p53 has not been deleted or mutated in tumor, and it could still trigger epithelial-mesenchymal transition (EMT) and lead to the occurrence of tumor invasion and metastasis, but the specific mechanism remains unclear. Our study had demonstrated that Rock2 and FAT10 were highly expressed in HCC, and promoted invasion and metastasis of HCC. In our previous study, through the model of human hepatocellular carcinoma in nude mice, microarray technology and molecular biology experiments, some phenomenon were found when Rock2 expression was decreased. Frist, stable knockdown of Rock2 significantly reduced the lung metastasis, and significantly changed EMT-related molecular markers in p53+/+ liver cancer cells. Further study showed that down-regulation of Rock2 expression decreased FAT10 expression, and increased the ubiquitination level of p53, but the protein expression of p53 was increased. However, after knocking out FAT10, Rock2 had no effect on both the ubiquitination level and expression of p53. Second, stable knockdown of Rock2 did not influence the lung metastasis and EMT in p53-/- liver cancer cells. Therefore, we put forward the hypothesis: Rock2 could promote the ubiquitination-independent degradation of p53 by FAT10 to regulate EMT of HCC cells. In order to verify the hypothesis, in this study, its specific mechanisms will be further explored at the animal, cellular and molecular levels by using animal imaging technology, gene knockout technology, and Co-immunoprecipitation assay. These provide a new theoretical foundation for prevention and treatment of HCC invasion and metastasis.