Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan 430000, China.
Int J Mol Sci. 2022 Jun 9;23(12):6451. doi: 10.3390/ijms23126451.
Gastric cancer (GC) is the fifth most common cancer and the third deadliest cancer in the world, and the occurrence and development of GC are influenced by epigenetics. Methyltransferase-like 3 (METTL3) is a prominent RNA n6-adenosine methyltransferase (m6A) that plays an important role in tumor growth by controlling the work of RNA. This study aimed to reveal the biological function and molecular mechanism of METTL3 in GC. The expression level of METTL3 in GC tissues and cells was detected by qPCR, Western blot and immunohistochemistry, and the expression level and prognosis of METTL3 were predicted in public databases. CCK-8, colony formation, transwell and wound healing assays were used to study the effect of METTL3 on GC cell proliferation and migration. In addition, the enrichment effect of METTL3 on DEK mRNA was detected by the RIP experiment, the m6A modification effect of METTL3 on DEK was verified by the MeRIP experiment and the mRNA half-life of DEK when METTL3 was overexpressed was detected. The dot blot assay detects m6A modification at the mRNA level. The effect of METTL3 on cell migration ability in vivo was examined by tail vein injection of luciferase-labeled cells. The experimental results showed that METTL3 was highly expressed in GC tissues and cells, and the high expression of METTL3 was associated with a poor prognosis. In addition, the m6A modification level of mRNA was higher in GC tissues and GC cell lines. Overexpression of METTL3 in MGC80-3 cells and AGS promoted cell proliferation and migration, while the knockdown of METTL3 inhibited cell proliferation and migration. The results of in vitro rescue experiments showed that the knockdown of DEK reversed the promoting effects of METTL3 on cell proliferation and migration. In vivo experiments showed that the knockdown of DEK reversed the increase in lung metastases caused by the overexpression of METTL3 in mice. Mechanistically, the results of the RIP experiment showed that METTL3 could enrich DEK mRNA, and the results of the MePIP and RNA half-life experiments indicated that METTL3 binds to the 3'UTR of DEK, participates in the m6A modification of DEK and promotes the stability of DEK mRNA. Ultimately, we concluded that METTL3 promotes GC cell proliferation and migration by stabilizing DEK mRNA expression. Therefore, METTL3 is a potential biomarker for GC prognosis and a therapeutic target.
胃癌（GC）是全球第五大常见癌症和第三大癌症死亡原因，其发生和发展受表观遗传学影响。甲基转移酶样 3（METTL3）是一种重要的 RNA N6-腺苷甲基转移酶（m6A），通过控制 RNA 的作用在肿瘤生长中发挥重要作用。本研究旨在揭示 METTL3 在 GC 中的生物学功能和分子机制。通过 qPCR、Western blot 和免疫组化检测 GC 组织和细胞中 METTL3 的表达水平，并通过公共数据库预测 METTL3 的表达水平和预后。CCK-8、集落形成、Transwell 和划痕愈合实验用于研究 METTL3 对 GC 细胞增殖和迁移的影响。此外，通过 RIP 实验检测 METTL3 对 DEK mRNA 的富集效应，通过 MeRIP 实验验证 METTL3 对 DEK 的 m6A 修饰作用，并检测当 METTL3 过表达时 DEK 的 mRNA 半衰期。点印迹实验检测 mRNA 水平的 m6A 修饰。通过尾静脉注射荧光素标记的细胞检测 METTL3 对细胞迁移能力的体内影响。实验结果表明，METTL3 在 GC 组织和细胞中高表达，且 METTL3 高表达与预后不良相关。此外，GC 组织和 GC 细胞系中 mRNA 的 m6A 修饰水平较高。在 MGC80-3 细胞和 AGS 中过表达 METTL3 促进细胞增殖和迁移，而敲低 METTL3 则抑制细胞增殖和迁移。体外挽救实验结果表明，敲低 DEK 可逆转 METTL3 对细胞增殖和迁移的促进作用。体内实验表明，敲低 DEK 可逆转 METTL3 过表达引起的小鼠肺转移增加。机制上，RIP 实验结果表明 METTL3 可富集 DEK mRNA，MePIP 和 RNA 半衰期实验结果表明 METTL3 结合 DEK 的 3'UTR，参与 DEK 的 m6A 修饰并促进 DEK mRNA 的稳定性。最终，我们得出结论，METTL3 通过稳定 DEK mRNA 表达促进 GC 细胞增殖和迁移。因此，METTL3 是 GC 预后的潜在生物标志物和治疗靶点。
