N4 acetylcytidine (ac4C) modification mainly occurs on tRNA, rRNA, and mRNA, playing an important role in the expression of genetic information. However, it is still unclear whether microRNAs have undergone ac4C modification and their potential physiological and pathological functions. In this study, we identified that NAT10/THUMPD1 acetylates primary microRNAs (pri-miRNAs) with ac4C modification. Knockdown of NAT10 suppresses and augments the expression levels of mature miRNAs and pri-miRNAs, respectively. Molecular mechanism studies found that pri-miRNA ac4C promotes the processing of pri-miRNA into precursor miRNA (pre-miRNA) by enhancing the interaction of pri-miRNA and DGCR8, thereby increasing the biogenesis of mature miRNA. Knockdown of NAT10 attenuates the oncogenic characters of lung cancer cells by regulating miRNA production in cancers. Moreover, NAT10 is highly expressed in various clinical cancers and negatively correlated with poor prognosis. Thus, our results reveal that NAT10 plays a crucial role in cancer initiation and progression by modulating pri-miRNA ac4C to affect miRNA production, which would provide an attractive therapeutic strategy for cancers.

N4乙酰胞苷(ac4C)修饰主要发生在tRNA、rRNA和mRNA上，在遗传信息的表达中发挥重要作用。然而，microRNA是否经历了ac4C修饰及其潜在的生理和病理功能仍不清楚。在这项研究中，我们发现 NAT10/THUMPD1 通过 ac4C 修饰乙酰化初级 microRNA (pri-miRNA)。 NAT10 的敲低分别抑制和增强成熟 miRNA 和 pri-miRNA 的表达水平。分子机制研究发现，pri-miRNA ac4C通过增强pri-miRNA与DGCR8的相互作用，促进pri-miRNA加工成前体miRNA（pre-miRNA），从而增加成熟miRNA的生物合成。 NAT10 的敲低通过调节癌症中 miRNA 的产生来减弱肺癌细胞的致癌特征。此外，NAT10在多种临床癌症中高表达，与不良预后呈负相关。因此，我们的结果表明，NAT10 通过调节 pri-miRNA ac4C 影响 miRNA 的产生，在癌症的发生和进展中发挥着至关重要的作用，这将为癌症提供一种有吸引力的治疗策略。