An Di, Wu Yuhao, Han Jingzhe, Fang Pingping, Bu Yi, Ji Guang, Deng Jinliang, Song Xueqin
The Second Hospital of Hebei Medical University, Hebei Medical University, Shijiazhuang, Hebei, China.
Department of Neurology, Affiliated Hospital of Hebei University, Baoding, Hebei, China.
Am J Med Genet B Neuropsychiatr Genet. 2025 Jul 17:e33048. doi: 10.1002/ajmg.b.33048.
We investigate the role of m6A RNA methylation in regulating transcription factor EB (TFEB) and its contribution to mitochondrial autophagy (mitophagy) dysfunction in amyotrophic lateral sclerosis (ALS). ALS cell models were used to analyse mitophagy markers and TFEB expression under METTL3 and TFEB modulation, using RT-qPCR, Western blot, MeRIP, RIP, and immunofluorescence. Elevated m6A methylation and reduced TFEB expression were observed in hSOD1-G93A models. METTL3 overexpression suppressed TFEB expression, leading to impaired mitophagy, while METTL3 knockdown alleviated these effects. MeRIP assays confirmed increased m6A modifications on TFEB mRNA, and RIP assays demonstrated direct interaction between METTL3 and TFEB mRNA. Notably, TFEB overexpression rescued mitophagy dysfunction, whereas TFEB knockdown exacerbated the impairment. METTL3-mediated m6A methylation inhibits mitophagy by downregulating TFEB expression, revealing the m6A-TFEB pathway as a promising therapeutic target for ALS.
我们研究了m6A RNA甲基化在调节转录因子EB（TFEB）中的作用及其对肌萎缩侧索硬化症（ALS）中线粒体自噬（线粒体吞噬）功能障碍的影响。使用ALS细胞模型，通过RT-qPCR、蛋白质免疫印迹、甲基化RNA免疫沉淀（MeRIP）、RNA免疫沉淀（RIP）和免疫荧光分析在METTL3和TFEB调节下的线粒体自噬标志物和TFEB表达。在hSOD1-G93A模型中观察到m6A甲基化升高和TFEB表达降低。METTL3过表达抑制TFEB表达，导致线粒体自噬受损，而METTL3敲低则减轻了这些影响。MeRIP分析证实TFEB mRNA上的m6A修饰增加，RIP分析表明METTL3与TFEB mRNA之间存在直接相互作用。值得注意的是，TFEB过表达挽救了线粒体自噬功能障碍，而TFEB敲低则加剧了这种损伤。METTL3介导的m6A甲基化通过下调TFEB表达来抑制线粒体自噬，揭示了m6A-TFEB途径作为ALS的一个有前景的治疗靶点。
