N⁶-methyladenosine (m⁶A) is an abundant internal RNA modification, influencing transcript fate and function in uninfected and virus-infected cells. Installation of m⁶A by the nuclear RNA methyltransferase METTL3 occurs cotranscriptionally; however, the genomes of some cytoplasmic RNA viruses are also m⁶A-modified. How the cellular m⁶A modification machinery impacts coronavirus replication, which occurs exclusively in the cytoplasm, is unknown. Here we show that replication of SARS-CoV-2, the agent responsible for the COVID-19 pandemic, and a seasonal human β-coronavirus HCoV-OC43, can be suppressed by depletion of METTL3 or cytoplasmic m⁶A reader proteins YTHDF1 and YTHDF3 and by a highly specific small molecule METTL3 inhibitor. Reduction of infectious titer correlates with decreased synthesis of viral RNAs and the essential nucleocapsid (N) protein. Sites of m⁶A modification on genomic and subgenomic RNAs of both viruses were mapped by methylated RNA immunoprecipitation sequencing (meRIP-seq). Levels of host factors involved in m⁶A installation, removal, and recognition were unchanged by HCoV-OC43 infection; however, nuclear localization of METTL3 and cytoplasmic m⁶A readers YTHDF1 and YTHDF2 increased. This establishes that coronavirus RNAs are m⁶A-modified and host m⁶A pathway components control β-coronavirus replication. Moreover, it illustrates the therapeutic potential of targeting the m⁶A pathway to restrict coronavirus reproduction.

中文翻译：

---

靶向 m6A RNA 修饰途径可阻断 SARS-CoV-2 和 HCoV-OC43 复制

N ⁶ -甲基腺苷 (m ⁶ A) 是一种丰富的内部 RNA 修饰，影响未感染和病毒感染细胞中的转录物命运和功能。核 RNA 甲基转移酶 METTL3 对m ^{6 A 的安装是同步发生的；}然而，一些细胞质 RNA 病毒的基因组也是 m ⁶ A 修饰的。细胞 m ⁶ A 修饰机制如何影响仅发生在细胞质中的冠状病毒复制尚不清楚。在这里，我们表明 SARS-CoV-2（导致 COVID-19 大流行的病原体）和季节性人类 β-冠状病毒 HCoV-OC43 的复制可以通过 METTL3 或细胞质 m 6 的耗尽来^抑制阅读器蛋白 YTHDF1 和 YTHDF3 以及高度特异性的小分子 METTL3 抑制剂。感染滴度的降低与病毒 RNA 和必需核衣壳 (N) 蛋白的合成减少有关。^{通过甲基化 RNA 免疫沉淀测序 (meRIP-seq) 对两种病毒的基因组和亚基因组 RNA 的 m 6} A修饰进行了定位。^{HCoV-OC43 感染未改变 m 6} A 安装、移除和识别中涉及的宿主因素水平；^{然而，METTL3 和细胞质 m 6} A 读取器 YTHDF1 和 YTHDF2的核定位增加。这表明冠状病毒 RNA 是 m ⁶ A 修饰的并且宿主是 m ⁶A 通路成分控制 β-冠状病毒复制。^{此外，它还说明了靶向 m 6} A 通路以限制冠状病毒繁殖的治疗潜力。