RBBP6调控NF-κB促进结肠癌细胞EMT的作用及机制研究

课题组前期通过基因芯片、QPCR和组织芯片发现RBBP6与结肠癌转移相关。上皮间质转化(EMT)是癌转移的关键步骤，E-cad丢失是其重要标志。我们在RBBP6高表达组织中观察到E-cad丢失，提示RBBP6参与EMT发生。泛素化作用激活NF-κB是调控EMT的重要途径；我们沉默泛素化酶RBBP6后NF-κB亚单位p65入核减少，而RBBP6高表达组织中p65上调。据此分析：RBBP6通过泛素化激活NF-κB促进EMT进而诱发癌转移，结肠癌转移可能存在RBBP6-NF-κB-EMT信号途径，有待进一步证实。本课题通过①基因沉默和过表达观察RBBP6在结肠癌转移中的作用；②标志物检测验证RBBP6是否参与EMT；③免疫沉淀和激光共聚焦探讨RBBP6调控NF-κB的作用位点；④蛋白酶阻滞剂MG132进行干预研究。可望阐明RBBP6调控NF-κB促进EMT的分子机制，为结肠癌防治提供潜在靶点。

By Our previous study using microarray, QPCR and colon cancer tissue microarray, we found RBBP6 was related to metastasis. Epithelial-mesenchymal transition (EMT) is a key step in cancer metastasis, and E-cad loss is a significant symbol during this process. We observed E-cad loss in RBBP6 high expression tissue, which suggests that RBBP6 is involved in EMT. Activating NF-κB by ubiquitination is an important way of regulating EMT. After silencing ubiquitination enzyme RBBP6, subunit p65 of NF-κB decreased in the nucleus. And p65 was up-regulated in RBBP6 high expression tissue. Above experiments display that ubiquitination enzyme RBBP6 can activate NF-κB to stimulate EMT and then promote metastasis. The RBBP6-NF-κB-EMT pathway may be existed in colon cancer metastasis. It needs to be confirmed. Thus in this study we will observe the effect of RBBP6 in colon cancer metastasis by gene silencing and over-expression, verify whether RBBP6 participate in EMT by markers detecting, explore the binding sites of RBBP6 on NF-κB by co-immunoprecipitation and confocal laser scanning, and study the effect of proteasome blocker MG132 on RBBP6. Our purpose is to clarify the molecular mechanisms about RBBP6 activating NF-κB to promote EMT, provide potential target for prevention and treatment of colon cancer.