N(6)-methyladenosine-binding protein YTHDF1 suppresses EBV replication and promotes EBV RNA decay

N⁶ -methyladenosine (m⁶ A) modification of mRNA mediates diverse cellular and viral functions. Infection with Epstein-Barr virus (EBV) is causally associated with nasopharyngeal carcinoma (NPC), 10% of gastric carcinoma, and various B-cell lymphomas, in which the viral latent and lytic phases both play vital roles. Here, we show that EBV transcripts exhibit differential m⁶ A modification in human NPC biopsies, patient-derived xenograft tissues, and cells at different EBV infection stages. m⁶ A-modified EBV transcripts are recognized and destabilized by the YTHDF1 protein, which leads to the m⁶ A-dependent suppression of EBV infection and replication. Mechanistically, YTHDF1 hastens viral RNA decapping and mediates RNA decay by recruiting RNA degradation complexes, including ZAP, DDX17, and DCP2, thereby post-transcriptionally downregulating the expression of EBV genes. Taken together, our results reveal the critical roles of m⁶ A modifications and their reader YTHDF1 in EBV replication. These findings contribute novel targets for the treatment of EBV-associated cancers.