MiR-33a suppresses proliferation of NSCLC cells via targeting METTL3 mRNA
MiR-33a suppresses proliferation of NSCLC cells via targeting METTL3 mRNA
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DOI：
10.1016/j.bbrc.2016.11.077
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发表时间：
2017-01-22
影响因子：
3.1
通讯作者：
Gao, Yushun
中科院分区：
文献类型：
--
作者：
Du, Minjun;Zhang, Yanjiao;Gao, Yushun
甲基转移酶样3（Methyltransferase like 3，简称MTL 3）最初被认为是一种甲基转移酶，负责N-6-甲基腺苷（m（6）A）的甲基化。胃L3可促进几种关键癌蛋白的表达，其高表达增强人肺癌细胞的增殖、存活和侵袭。然而，在非小细胞肺癌（NSCLC）中，胃L3是如何调节的还很少了解。在本研究中，通过计算机模拟预测，miR-33 a被怀疑靶向于胃L3 mRNA的3 '-非翻译区（3' UTR）。此外，胃L3在NSCLC组织中的表达高于癌旁组织，且胃L3与miR 33 a的表达呈正相关。荧光素酶报告基因检测表明，miR-33 a可直接靶向胃癌L3 mRNA的3 'UTR。此外，我们发现，miR-33 a可以减少胃L3在mRNA和蛋白质水平的表达，通过逆转录-聚合酶链反应（RT-PCR）和Western blot分析。在功能上，miR-33 a可以降低A549和NCI-H460细胞的增殖。相反，使用4，5-二甲基-2-噻唑基）-2，5-二苯基-2-H-溴化四唑（MIT）和5-乙炔基-2-脱氧尿苷（EdU）测定，通过抗miR-33 a抑制miR-33 a可以挽救该结果。类似地，miR-33 a可以减少A549细胞的细胞锚定非依赖性生长。此外，通过Western blot分析检测miR-33 a对胃L3下游基因的负面影响。因此，我们得出结论，miR 33 a可以通过靶向胃L3 mRNA的3 'UTR来减弱NSCLC细胞的增殖。我们的发现为microRNA调控胃L3的机制提供了新的见解，并支持胃L3作为NSCLC的治疗靶点。(C)2016 Elsevier Inc. All rights reserved.
Methyltransferase like 3 (METTL3) was incipiently known as a methyltransferase which was responsible for N-6-methyladenosine (m(6)A) methylation. METTL3 can promote the expression of several crucial oncoproteins and its high expression enhanced proliferation, survival, and invasion of human lung cancer cells. However, how METTL3 was regulated is seldom understood in non-small-cell lung carcinoma (NSCLC). In the present study, miR-33a was suspicious to target to the 3'-untranslated region (3'UTR) of METTL3 mRNA via in silico prediction. Besides, the expressions of METTL3 were higher in NSCLC tissues than those in adjacent tissues, and METTL3 expressions were positively related to the expressions of miR33a in NSCLC tissues which confirmed by quantitative real-time polymerase chain reaction (qRT-PCR). MiR-33a can directly target to the 3'UTR of METTL3 mRNA which examined by luciferase reporter gene assay. Moreover, we found that miR-33a can reduce the expression of METTL3 at both mRNA and protein levels using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. Functionally, miR-33a can reduce the proliferation of A549 and NCI-H460 cells. Conversely, inhibition of miR-33a by anti-miR-33a can rescue that using 4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MIT) and 5-ethynyl-2-deoxyuridine (EdU) assay. Similarly, miR-33a can reduce cellular anchorage-independent growth of A549 cells. Additionally, the negative influences of miR-33a on the downstream genes of METTL3 were examined by Western blot analysis. Thus, we concluded that miR33a can attenuate NSCLC cells proliferation via targeting to the 3'UTR of METTL3 mRNA. Our findings provide new insights into the mechanism of METTL3 regulation by micro RNA, and supports METTL3 as a therapeutic target in NSCLC. (C) 2016 Elsevier Inc. All rights reserved.