N^(6)-甲基腺苷(m6A)转移酶METTL3和m6A调控LINC01465在肝细胞癌增殖转移中的作用机制  

Mechanism of N^(6)-methyladenosine(m6A)transferase METTL3 and m6A regulation of LINC01465 in the proliferation and metastasis of hepatocellular carcinoma

作　　者：刘肸琳 龙朴泽 黄云美[3] 罗春英[3,4] Liu Xilin;Long Puze;Huang Yunmei;Luo Chunying(Department of Graduate School,Youjiang Medical University for Nationalities,Baise Guangxi 533000,China;Department of Pathology of Dongfeng Hospital,Hubei University of Medicine,Shiyan Hubei 442008,China;Department of Pathology,Affiliated Hospital of Youjiang Medical University for Nationalities,Baise Guangxi 533000,China;The Key Laboratory of Molecular Pathology(For Hepatobiliary Diseases)of Guangxi,Baise Guangxi 533000,China)

机构地区：[1]右江民族医学院研究生学院,广西百色533000 [2]湖北医药学院附属东风医院病理科,湖北十堰442008 [3]右江民族医学院附属医院病理科,广西百色533000 [4]广西肝胆疾病分子病理重点实验室,广西百色533000

出　　处：《遵义医科大学学报》2024年第2期135-144,共10页Journal of Zunyi Medical University

基　　金：广西自然科学基金资助项目(NO:2017GXNSFGA198002);百色市区域多发病联合专项计划项目(NO:20224104)。

摘　　要：目的探讨甲基转移酶样3(METTL3)在肝癌细胞中调控m6A水平,影响长链非编码(lncRNA)LINC01465的表达,促进肝癌细胞增殖转移的机制。方法利用人类癌症基因组图谱(TCGA)数据库分析METTL3在肝癌组织和正常肝组织中的表达差异及与患者预后的关系;用Western blot、RT-qPCR分析验证METTL3在细胞、组织中的表达;用m6A比色法验证肝癌细胞及正常肝细胞中的m6A水平;敲低METTL3后进行CCK-8、克隆形成、Transwell迁移、侵袭实验,研究METTL3对肝癌增殖转移的调控作用;转录组RNA m6A甲基化测序确定METTL3的下游调控基因LINC01465。RT-qPCR验证沉默METTL3后LINC01465的表达量,以及LINC01465在肝细胞癌中的表达水平。结果METTL3在肝癌组织和细胞中明显高表达且与患者预后较差相关;高表达的METTL3促进HCC细胞增殖、迁移和侵袭;METTL3影响LINC01465的m6A水平及其表达量来促进肝细胞癌的发展进程。结论METTL3可通过m6A依赖的方式来影响LINC01465的表达水平,促进肝癌细胞增殖转移,METTL3可能成为肝癌预后及治疗的靶向标志物。Objective To investigate the mechanism of methyltransferase like 3(METTL3)in hepatocellular carcinoma cells to regulate the level of m6A,affect the expression of long-chain non-coding(lncRNA)LINC01465,and promote the proliferation and metastasis of hepatocellular carcinoma cells.Methods The Cancer Genome Atlas(TCGA)database was used to analyze the expression differences of METTL3 in hepatocellular carcinoma tissues and normal liver tissues and explain its relationship with patients’prognosis;the expression of METTL3 in cells and tissues was verified by western blot and RT-qPCR analyses;the levels of m6A were verified by the m6A colorimetric assay in hepatocellular carcinoma cells and normal hepatocytes;and after the knockdown of METTL3,the CCK-8,clone formation,Transwell migration,and invasion assays was used to study the regulatory role of METTL3 on the proliferation and metastasis of hepatocellular carcinoma;transcriptome RNA m6A methylation sequencing was performed to determine LINC01465,the downstream regulatory gene of METTL3.RT-qPCR was performed to verify the expression of LINC01465 after silencing of METTL3 and the expression level of LINC01465 in hepatocellular carcinoma.Results METTL3 was significantly highly expressed in hepatocellular carcinoma tissues and cells and was associated with poorer prognosis of patients;highly expressed METTL3 promoted the proliferation,migration and invasion of HCC cells;METTL3 affected the m6A level of LINC01465 and its expression,promoting the developmental process of hepatocellular carcinoma.Conclusion METTL3 can affect the expression level of LINC01465 in an m6A-dependent manner to promote the proliferation and metastasis ability of hepatocellular carcinoma cells,suggesting that METTL3 may become a target marker for the prognosis and treatment of hepatocellular carcinoma.

关 键 词：甲基转移酶样3 m6A甲基化 LINC01465 肝细胞癌 

分 类 号：R735.7[医药卫生—肿瘤]