BACKGROUND The importance of mRNA methylation erased by ALKBH5 in mRNA biogenesis, decay, and translation control is an emerging research focus. Ectopically activated YAP is associated with the development of many human cancers. However, the mechanism whereby ALKBH5 regulates YAP expression and activity to inhibit NSCLC tumor growth and metastasis is not clear. METHODS Protein and transcript interactions were analyzed in normal lung cell and NSCLC cells. Gene expression was evaluated by qPCR and reporter assays. Protein levels were determined by immunochemical approaches. Nucleic acid interactions and status were analyzed by immunoprecipitation. Cell behavior was analyzed by standard biochemical tests. The m6A modification was analyzed by MeRIP. RESULTS Our results show that YAP expression is negatively correlated with ALKBH5 expression and plays an opposite role in the regulation of cellular proliferation, invasion, migration, and EMT of NSCLC cells. ALKBH5 reduced m6A modification of YAP. YTHDF3 combined YAP pre-mRNA depending on m6A modification. YTHDF1 and YTHDF2 competitively interacted with YTHDF3 in an m6A-independent manner to regulate YAP expression. YTHDF2 facilitated YAP mRNA decay via the AGO2 system, whereas YTHDF1 promoted YAP mRNA translation by interacting with eIF3a; both these activities are regulated by m6A modification. Furthermore, ALKBH5 decreased YAP activity by regulating miR-107/LATS2 axis in an HuR-dependent manner. Further, ALKBH5 inhibited tumor growth and metastasis in vivo by reducing the expression and activity of YAP. CONCLUSIONS The presented findings suggest m6A demethylase ALKBH5 inhibits tumor growth and metastasis by reducing YTHDFs-mediated YAP expression and inhibiting miR-107/LATS2-mediated YAP activity in NSCLC. Moreover, effective inhibition of m6A modification of ALKBH5 might constitute a potential treatment strategy for lung cancer.

中文翻译：

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m6A 去甲基化酶 ALKBH5 通过降低 YTHDFs 介导的 YAP 表达和抑制 miR-107/LATS2 介导的 NSCLC 中的 YAP 活性来抑制肿瘤生长和转移。

背景技术 ALKBH5 消除的 mRNA 甲基化在 mRNA 生物发生、衰变和翻译控制中的重要性是一个新兴的研究焦点。异位激活的 YAP 与许多人类癌症的发展有关。然而，ALKBH5 调节 YAP 表达和活性以抑制 NSCLC 肿瘤生长和转移的机制尚不清楚。方法 在正常肺细胞和 NSCLC 细胞中分析蛋白质和转录物的相互作用。通过 qPCR 和报告基因分析评估基因表达。通过免疫化学方法测定蛋白质水平。通过免疫沉淀分析核酸相互作用和状态。通过标准生化测试分析细胞行为。通过 MeRIP 分析 m6A 修饰。结果我们的研究结果表明，YAP的表达与ALKBH5的表达呈负相关，并且在NSCLC细胞的细胞增殖、侵袭、迁移和EMT的调节中起相反的作用。ALKBH5 减少了 YAP 的 m6A 修饰。YTHDF3 根据 m6A 修饰结合了 YAP 前 mRNA。YTHDF1 和 YTHDF2 以不依赖 m6A 的方式与 YTHDF3 竞争性相互作用以调节 YAP 表达。YTHDF2 通过 AGO2 系统促进 YAP mRNA 衰变，而 YTHDF1 通过与 eIF3a 相互作用促进 YAP mRNA 翻译；这两项活动都受 m6A 修饰的调节。此外，ALKBH5 通过以 HuR 依赖性方式调节 miR-107/LATS2 轴来降低 YAP 活性。此外，ALKBH5 通过降低 YAP 的表达和活性来抑制体内肿瘤生长和转移。结论 所提出的研究结果表明 m6A 去甲基化酶 ALKBH5 通过降低 YTHDFs 介导的 YAP 表达和抑制 miR-107/LATS2 介导的 YAP 活性在 NSCLC 中抑制肿瘤生长和转移。此外，有效抑制 ALKBH5 的 m6A 修饰可能构成肺癌的潜在治疗策略。