一种新的与食管鳞癌预后相关的长链非编码RNA EP1的生物学功能及其作用机制的研究

Previous work by our group using the gene chip to detect the expression level of 8990 lncRNAs in 119 pairs of esophageal squamous cell cancer and adjacent normal tissue, we found that a large number of lncRNAs are abnormal expression, showing a significantly difference expression profile in the tumor tissue, this result showed that lncRNA molecules involved in the development of esophageal squamous cell carcinoma. Further analysis of our study found a lncRNA (located on 14q23.1) which function is uncommented has significant lower expression level in esophageal squamous cell carcinoma, and its reduced expression is significantly associated with lymph node metastasis and poor prognosis of patients, so we named it EP1. This finding prompted that EP1 might regulate the transfer of esophageal squamous cell carcinoma. Our project will use Northern Blot and RACE to get the whole sequence of EP1; and then study the cell proliferation, apoptosis, movement and invasion capacity in esophageal squamous cell carcinoma cells by overexpressing EP1; based on these studies, we use RNA-Seq to detect whether EP1 overexpression can cause any downstream mRNA transcription anomaly. Through the above research，we may reveal the mechanism of the new lncRNA EP1 in esophageal squamous cell carcinoma.

本课题组前期利用基因芯片检测了119对食管鳞癌组织和癌旁正常组织8990个lncRNAs的表达水平，生物信息学分析表明，在肿瘤组织中呈现出了明显的异常lncRNA表达谱，提示大量的lncRNA参与了食管鳞癌的发生发展。进一步分析发现了一个功能未被报道的lncRNA分子（定位于14q23.1染色体负链）在食管鳞癌组织中表达发生显著下调，而且其表达降低与淋巴结转移和病人的预后差显著相关，因此我们将其命名为EP1。这些发现表明EP1可能调节了食管鳞癌细胞的转移。本项目将首先利用Northern Blot和RACE获取EP1的cDNA全长序列；通过RNA-FISH和RT-PCR明确EP1的细胞定位；进一步通过在食管鳞癌细胞中过表EP1来研究其对食管鳞癌细胞增殖、凋亡、运动和侵袭能力的影响；在此基础上通过RNA-Seq检测检测EP1的过表达引起了哪些下游mRNA的转录mRNA的转录异常。

我们对前期芯片研究发现的在食管鳞癌中显著异常表达的lncRNA进行了深入分析和初步全长获取实验，利用RACE法成功获得了1个新的lncRNA的全长序列，根据后续研究将其命名为lncRNA NMR。利用qRT-PCR对这个新的lncRNA在食管鳞癌组织中的表达进行了检测，证实了lncNMR在食管鳞癌中表达升高并与患者淋巴结转移和预后差相关。利用RNA FISH进一步确定了lncNMR在定位在食管鳞癌细胞的细胞核。继而深入研究证实了lncNMR促进了食管鳞癌细胞迁移和侵袭能力，并抑制了顺铂和紫杉醇诱导的食管鳞癌细胞凋亡。