Post-translational modification of proteins plays a critical role in plant–pathogen interactions. Here, we demonstrate in Nicotiana benthamiana that knockout of NbHAG1 promotes Chinese wheat mosaic virus (CWMV) infection, whereas NbHAG1 overexpression inhibits infection. Transcriptome sequencing indicated that a series of disease resistance-related genes were up-regulated after overexpression of NbHAG1. In addition, cleavage under targets and tagmentation (Cut&Tag)-qPCR results demonstrated that NbHAG1 may activate the transcription of its downstream disease-resistance genes by facilitating the acetylation level of H3K36ac. Therefore, we suggest that NbHAG1 is an important positive regulator of resistance to CWMV infestation.

Although RNA cytosine-5 methylation (m⁵C) is an important post-transcriptional regulatory mechanism, its contribution to plant antiviral immunity remains unclear. In this study, we identified Thiamine thiazole synthase 2 (TaTHI2) as a host mRNA target of the wheat m⁵C methyltransferase TaNSUN2 during infection by Chinese wheat mosaic virus (CWMV), a soil-borne virus that poses a major threat to wheat production. TaNSUN2 contributes to the m⁵C modification of TaTHI2 transcripts, enhancing mRNA stability and sustaining TaTHI2 accumulation. The disruption of a key m⁵C site markedly reduced methylation, weakened TaNSUN2–RNA binding, and accelerated transcript decay, leading to the compromised production of reactive oxygen species (ROS) and increased viral infection. Mechanistically, the TaNSUN2-dependent m⁵C modification stabilized TaTHI2 mRNA, thereby promoting ROS-mediated antiviral defense. Collectively, our results establish the m⁵C modification of TaTHI2 mRNA as a critical post-transcriptional control point in CWMV resistance and highlight TaNSUN2-dependent RNA methylation as an integral component of host antiviral immunity.