The latent infection by herpes virus type 1 (HSV-1) may be lifelong in trigeminal ganglia and a suspected cause of Alzheimer's Disease (AD) and Amyotrophic lateral sclerosis (ALS). Whether and how N6-methyladenosine (m6A) modification of viral RNAs affects virus infection are poorly understood. Here, we report that HSV-1 infection enhanced the expression of m6A writers (METTL3, METTL14) and readers (YTHDF1/2/3) at the early infection stage and decreased their expression later on, while suppressed the erasers' (FTO, ALBKH5) expression immediately upon infection to facilitate viral replication. Inhibiting m6A modification by 3-deazaadenosine (DAA) significantly decreased viral replication and reduced viral reproduction over 1000 folds. More interestingly, depleting the writers and readers by siRNAs inhibited virus replication and reproduction; whereas depleting the erasers promoted viral replication and reproduction. Silencing YTHDF3 strikingly decreased viral replication by up to 90%, leading to reduction of up to 10-fold viral replication and over 100-fold virus reproduction, respectively. Depletion of m6A initiator METTL3 (by 60%–70%) by siRNA correlatedly decreased viral replication 60%–70%, and reduced virus yield over 30-fold. Consistently, ectopic expression of METTL3 largely increased virus yield. METTL3 knockdown suppressed the HSV-1 intermediate early and early genes (ICP0, ICP8 and UL23) and late genes (VP16, UL44, UL49 and ICP47); while ectopic expression of METTL3 upregulated these gene expression. Results from our study shed the lights on the importance for m6A modification to initiate HSV-1 early replication. The components of m6A modification machinery, particularly m6A initiator METTL3 and reader YTHDF3, would be potential important targets for combating HSV-1 infections.

1 型疱疹病毒 (HSV-1) 的潜伏感染可能在三叉神经节中持续存在，并且可能导致阿尔茨海默病 (AD) 和肌萎缩侧索硬化症 (ALS)。对病毒 RNA 的 N6-甲基腺苷 (m6A) 修饰是否以及如何影响病毒感染知之甚少。在这里，我们报告 HSV-1 感染在感染早期增强了 m6A 写入者（METTL3，METTL14）和读取者（YTHDF1/2/3）的表达，随后降低了它们的表达，同时抑制了擦除器（FTO，ALBKH5 ) 感染后立即表达以促进病毒复制。通过 3-deazaadenosine (DAA) 抑制 m6A 修饰可显着降低病毒复制并将病毒复制减少 1000 倍以上。更有趣的是，通过 siRNA 耗尽作者和读者，抑制了病毒的复制和繁殖；而耗尽橡皮擦则促进了病毒的复制和繁殖。沉默 YTHDF3 显着降低了高达 90% 的病毒复制，分别导致病毒复制减少了 10 倍和病毒复制减少了 100 倍以上。siRNA 消耗 m6A 引发剂 METTL3（减少 60%–70%）使病毒复制减少 60%–70%，并使病毒产量减少 30 倍以上。一致地，METTL3 的异位表达大大增加了病毒产量。METTL3 敲低抑制了 HSV-1 中间早期和早期基因（ICP0、ICP8 和 UL23）和晚期基因（VP16、UL44、UL49 和 ICP47）；而 METTL3 的异位表达上调了这些基因的表达。我们的研究结果揭示了 m6A 修饰对启动 HSV-1 早期复制的重要性。m6A 修饰机制的组件，特别是 m6A 引发剂 METTL3 和阅读器 YTHDF3，将成为对抗 HSV-1 感染的潜在重要目标。