YTHDF1介导m6A在支气管上皮细胞恶性转化中的作用和调控机制研究

Lung cancer has become a serious issue of public health. Recently, RNA methylation as a hot point of epigenetics research has gained more attention. In our previous research, the level of m6A is apparently higher in lung cancer cells than that in HBE. The results of further dig in TCGA database showed that YTHDF1, the key gene of m6A, had more expression in lung cancer, which indicated the important role in the regulation of m6A in lung cancer. Analysis based on GCBI platform spotted miR-218-5p, the target gene of YTHDF1. Hence, this study is designed to explore the reaction of miR-218-5p/YTHDF1/m6A axis in the aetiology of lung cancer by the applications of malignant transformation model and tumor model of nude mice. And downstream proteins in YTHDF1 could be found through proteomics analysis. This study tries to explain the molecular mechanisms and recommendation for prevention of lung cancer in the novel point of RNA methylation.

肺癌严重威胁人类健康，RNA甲基化作为近年来表观遗传学领域的最新研究热点，在肿瘤发生发展中的作用备受关注。课题组前期研究发现，肺癌细胞中N6-腺苷甲基化（m6A）表达水平明显高于正常人支气管上皮细胞，进一步对TCGA数据库测序数据进行分析发现，m6A关键调控基因YTHDF1在肺癌组织中表达显著上升，可能在调控肺癌RNA甲基化水平中发挥重要作用，结合GCBI平台分析发现miR-218-5p可能通过对YTHDF1的靶向调控对肺癌发生产生影响。因此，本项目拟应用细胞恶性转化模型及裸鼠成瘤模型，探讨miR-218-5p/YTHDF1/m6A调控轴在肺癌发生过程中的作用，并通过蛋白质组学技术分析YTHDF1对下游蛋白的调控作用。本研究从mRNA甲基化调控的新角度出发，为进一步阐明肺癌发生的分子机制及肺癌防治提供依据。

本研究在B(a)P诱导人支气管上皮恶转细胞和A549肺癌细胞中，通过转录组测序、m6A修饰测序发现YTHDF1及其m6A修饰在B(a)P致非小细胞肺癌的发生发展中有重要的作用。多个数据库分析结果显示，YTHDF1在肺癌组织中特异性高表达，且和预后密切相关。细胞功能实验表明，YTHDF1表达受到B(a)P暴露影响，且YTHDF1可调控恶转细胞增殖。miR-139-5p和miR-145-5负向调控YTHDF1表达，影响恶性转化细胞和肺癌细胞增殖、侵袭、迁移。TMT蛋白测序发现YTHDF1可调控下游一系列功能蛋白的表达，下游蛋白主要富集于PI3K-Akt和MAPK等癌症相关通路。研究提示，YTHDF1在B(a)P暴露致肺癌的发生过程中，可能通过miR-139-5p和miR-145-5p的调控，影响下游一系列功能蛋白的表达，从而对细胞功能产生影响，促进了肺癌的发生。YTDHF1可能是非小细胞肺癌的潜在生物标志物。