甲基转移酶样因子3通过调控GLUT4-mTORC1轴影响食管鳞状细胞癌细胞的糖酵解及增殖  

Methyltransferase-like 3 affects glycolysis and proliferation of esophageal squamous cell carcinoma cells by regulating the GLUT4-mTORC1 axis

作　　者：周锡 钟晓武 高川力[1] 李青容 程吉兵 马强[2,3] 郭晓兰[1,2] ZHOU Xi;ZHONG Xiaowu;GAO Chuanli;LI Qingrong;CHENG Jibing;MA Qiang;GUO Xiaolan(Department of Laboratory Medicine,North Sichuan Medical College,Nanchong 637000,Sichuan,China;Department of Clinical Laboratory,North Sichuan Medical College,Nanchong 637000,Sichuan,China;Translational Medicine Research Center,North Sichuan Medical College,Nanchong 637000,Sichuan,China)

机构地区：[1]川北医学院检验系,四川南充637000 [2]川北医学院检验科,四川南充637000 [3]川北医学院转化医学研究中心,四川南充637000

出　　处：《中国肿瘤生物治疗杂志》2022年第12期1076-1086,共11页Chinese Journal of Cancer Biotherapy

基　　金：四川省应用基础研究计划项目(No.2021YJ0202)。

摘　　要：目的:探讨甲基转移酶样因子3(METTL3)在食管鳞状细胞癌(ESCC)组织和细胞中的表达水平及其对ESCC细胞糖酵解和增殖能力的影响和潜在的分子机制。方法:基于TCGA数据库分析METTL3在ESCC细胞中的表达及可能的富集通路。收集2021年1月至2021年6月间在北川医学院附属医院外科手术切除的34例ESCC组织及相应癌旁组织,采用免疫组化法验证ESCC组织中METTL3的表达。采用CCK-8法和平板克隆形成实验检测干扰METTL3后ESCC细胞增殖能力的变化,利用比色法检测干扰METTL3后ESCC细胞总RNA中m6A的表达水平,采用甲基化RNA免疫沉淀定量PCR(MeRIP-qPCR)检测METTL3对葡萄糖转运蛋白4(GLUT4)基因mRNA的m6A修饰水平的影响,采用WB和qPCR等技术探索METTL3参与ESCC细胞糖酵解的生物学机制。结果:METTL3在ESCC组织以及细胞中均呈高表达(均P<0.001)。干扰METTL3表达后,ESCC细胞的增殖能力明显减弱、细胞内总RNA的m6A修饰水平显著降低(均P<0.001)。此外,干扰METTL3可显著抑制KYSE150和TE-1细胞中GLUT4基因mRNA的m6A修饰水平(均P<0.01),并通过下调GLUT4的表达抑制葡萄糖的摄取以及乳酸的释放(均P<0.01),最终下调mTORC1通路活性并抑制ESCC细胞的增殖;在干扰METTL3的ESCC细胞同时联合运用mTORC1通路抑制剂显示有协同的抗癌作用。结论:METTL3介导的m6A修饰通过调控GLUT4-mTORC1信号轴影响ESCC细胞的糖酵解及增殖。Objective: To investigate the expression of methyltransferase-like 3(METTL3) in esophageal squamous cell carcinoma(ESCC) tissue and cells and its effect on the glycolysis proliferation of ESCC cells as well as the potential molecular mechanisms.Methods: Based on the TCGA database, the expression of METTL3 and its possible enrichment pathway in ESCC were elucidated.Thirty-four ESCC tissues and corresponding paracancerous tissues were collected from surgical resections at the Affiliated Hospital of Beichuan Medical College between January 2021 and June 2021, the expression of METTL3 in ESCC tissues was verified by immunohistochemistry. The CCK-8 and colony formation assay were used to evaluate the change in the proliferation ability of ESCC cells after METTL3 knockdown. The expression level of m~6A in total RNA of ESCC cells after METTL3 knockdown was detected by colorimetric method. Methylated RNA immunoprecipitation qPCR(MeRIP-qPCR) was used to detect the effect of METTL3 on the m~6A modification level of glucose transporter 4(GLUT4) mRNA. The biological mechanisms of METTL3 in the glycolysis metabolism of ESCC were evaluated using WB and q PCR. Results: The expression level of METTL3 was significantly increased in ESCC tissues as well as cell lines(all P<0.001). After the knockdown of METTL3, the proliferation ability of ESCC cells was significantly reduced,and the m~6A modification level of total RNA was significantly reduced(all P<0.001). In addition, knockdown of METTL3 significantly inhibited the m~6A modification level of GLUT4 mRNA in KYSE150 and TE-1 cells(all P<0.01), inhibited glucose uptake and lactate release by down-regulating GLUT4 expression(all P<0.01), and finally down-regulated the activity of mTORC1 pathway and inhibited the proliferation of ESCC cells. Moreover, a synergetic effect was found in METTL3-depleted ESCC cells combined with mTORC1 pathway inhibitor. Conclusion: METTL3-mediated m~6A modification promotes glycolysis and proliferation of ESCC cells by regulating the GLUT4-mTORC1 signal

关 键 词：食管鳞状细胞癌 甲基转移酶样因子3 葡萄糖转运蛋白4 糖酵解 mTORC1通路 

分 类 号：R735.1[医药卫生—肿瘤] R730.2[医药卫生—临床医学]