N4-acetylcytidine regulates the replication and pathogenicity of enterovirus 71

Chemical modifications are important for RNA function and metabolism. N4-acetylcytidine (ac4C) is critical for the translation and stability of mRNA. Although ac4C is found in RNA viruses, the detailed mechanisms through which ac4C affects viral replication are unclear. Here, we reported that the 5' untranslated region of the enterovirus 71 (EV71) genome was ac4C modified by the host acetyltransferase NAT10. Inhibition of NAT10 and mutation of the ac4C sites within the internal ribosomal entry site (IRES) suppressed EV71 replication. ac4C enhanced viral RNA translation via selective recruitment of PCBP2 to the IRES and boosted RNA stability. Additionally, ac4C increased the binding of RNA-dependent RNA polymerase (3D) to viral RNA. Notably, ac4C-deficient mutant EV71 showed reduced pathogenicity in vivo. Our findings highlighted the essential role of ac4C in EV71 infection and provided insights into potential antiviral treatments.