Wang Xiao-Shuang, He Jin-Rong, Yu Shan, Yu Jia
State Key Lab of Medical Molecular Biology,Department of Biochemistry,Institute of Basic Medical Sciences,CAMS and PUMC,Beijing 100005,China.
Zhongguo Yi Xue Ke Xue Yuan Xue Bao. 2018 Jun 28;40(3):308-314. doi: 10.3881/j.issn.1000-503X.2018.03.002.
Objective To investigate the role of methyltransferase-like 3(METTL3) in the proliferation of acute myeloid leukemia (AML) cells and its mechanism. Methods METTL3 expression in AML patients was analyzed in Gene Expression Omnibus data files. METTL3 expression was inhibited by lentivirus-mediated gene transduction in MOLM13 cells,after which cell proliferation was analyzed by cell counting kit-8,N6-methyladenosine (m6A) levels of total mRNA was analyzed by ELISA,specific m6A on MYC was analyzed by gene-specific m6A RNA immunoprecipitation,and MYC expression was analyzed by RT-qPCR and Western blot analysis. Results METTL3 level was slightly increased in AML-M5 patients,and its expression was significantly higher in immature cells than in mature monocytes (t=4.504,P=0.0098). METTL3 knock-down significantly suppressed cell proliferation (P<0.001),reduced m6A level of total mRNA (t=3.606,P=0.042) and specific m6A level on MYC mRNA (P<0.01),and suppressed MYC expression (P<0.01). Conclusion METTL3 acts as an oncogene in MOLM13 cells by upregulating MYC expression.
目的 探讨甲基转移酶样3（METTL3）在急性髓系白血病（AML）细胞增殖中的作用及其机制。方法 在基因表达综合数据库数据文件中分析AML患者的METTL3表达。通过慢病毒介导的基因转导抑制MOLM13细胞中的METTL3表达，之后用细胞计数试剂盒-8分析细胞增殖，用酶联免疫吸附测定法分析总mRNA的N6-甲基腺苷（m6A）水平，用基因特异性m6A RNA免疫沉淀法分析MYC上的特异性m6A，并用逆转录定量聚合酶链反应和蛋白质免疫印迹分析来分析MYC表达。结果 AML-M5患者的METTL3水平略有升高，其在未成熟细胞中的表达明显高于成熟单核细胞（t=4.504，P=0.0098）。敲低METTL3可显著抑制细胞增殖（P<0.001），降低总mRNA的m6A水平（t=3.606，P=0.042）和MYC mRNA上的特异性m6A水平（P<0.01），并抑制MYC表达（P<0.01）。结论 METTL3通过上调MYC表达在MOLM13细胞中发挥癌基因作用。
