OTUB1通过DDX5促进结直肠癌细胞EMT的机制研究

Metastasis is the main cause of colorectal cancer treatment failure. Epithelial - mesenchymal transition (EMT) is the initial stage of cancer metastasis. Exploring the mechanism of EMT is very important to block cancer cell metastasis. Our preliminary study showed that the deubiquitinating enzyme OTUB1 could facilitate EMT, and promote the metastasis of colorectal cancer. But its mechanism was unknown. Bioinformatics analysis revealed that coactivator DDX5 was deubiquitinated by OTUB1 which was probably promote the EMT and then induced colorectal cancer metastasis. Therefore this topic will deeply explore the molecular mechanism which OTUB1 promotes EMT of colorectal cancer cells by DDX5 deubiquitination by immunohistochemical, immune coprecipitation and protein ubiquitination. This study will reveal the process of colorectal cancer metastasis, and find the new molecular mechanisms of EMT.

结直肠癌转移是治疗失败的最主要原因，上皮-间质转化（EMT）是癌细胞转移的起步阶段，探讨EMT的调控机制对于阻断癌细胞转移具有重要意义。我们的前期研究发现去泛素化酶OTUB1可调控EMT，并促进结直肠癌转移，其机制有待深入研究。生物信息学分析揭示OTUB1可去泛素化转录共激活子DDX5，其很可能促进EMT进程，进而诱导结直肠癌细胞转移。因此本课题拟通过免疫组化、免疫共沉淀、蛋白质泛素化等实验技术深入探索OTUB1去泛素化DDX5促进结直肠癌细胞EMT的分子机制。本研究将有助于揭示结直肠癌转移的过程，并寻找EMT调控新的分子机制。

结直肠癌是一种常见的恶性肿瘤，也是导致癌症患者死亡的主要原因之一。我们的前期研究发现OTUB1在结直肠癌组织中高表达，并通过调控EMT相关分子的表达促进结直肠癌转移，但是OTUB1异常表达的机制尚不清楚。我们通过生物信息学分析发现转录因子ERRα在结直肠癌组织中表达与OTUB1正相关，ERRα高表达患者预后差，因此我们推测ERRα可能调控OTUB1表达。首先，我们在结直肠癌细胞HCT116、Caco2细胞中过表达结或敲降ERRα，OTUB1在mRNA和蛋白水平均表达升高或降低。我们将OTUB1启动子区插入PGL3 Basic质粒，通过双荧光素酶检测发现，ERRα能够调控OTUB1启动子。按照预测的结合位点将OTUB1启动子区分段并突变启动子区，通过双荧光素酶实验发现ERRα和OTUB1启动区ERRE-S4 815/823 bp结合，同时通过CHIP验证了以上结果。我们发现过表达及敲降ERRα能够增加或降低结直肠癌细胞的迁移能力，降低或者过表达OTUB1能够逆转这种能力。我们进一步研究发现ERRα及OTUB1可能通过Vimentin促进结直肠癌转移。我们的研究进一步丰富了结直肠癌转移的机制，ERRα-OTUB1-Vimentin轴可能成为结直肠癌治疗的新靶点。