Epitranscriptomic Enhancement of Influenza A Virus Gene Expression and Replication

Many viral RNAs are modified by methylation of the N-6 position of adenosine (m(6)A). m(6)A is thought to regulate RNA splicing, stability, translation, and secondary structure. Influenza A virus (IAV) expresses m(6)A-modified RNAs, but the effects of m(6)A on this segmented RNA virus remain unclear. We demonstrate that global inhibition of m(6)A addition inhibits IAV gene expression and replication. In contrast, overexpression of the cellular m(6)A reader protein YTHDF2 increases IAV gene expression and replication. To address whether m(6)A residues modulate IAV RNA function in cis, we mapped m(6)A residues on the IAV plus (mRNA) and minus (vRNA) strands and used synonymous mutations to ablate m(6)A on both strands of the hemagglutinin (HA) segment. These mutations inhibited HA mRNA and protein expression while leaving other IAV mRNAs and proteins unaffected, and they also resulted in reduced IAV pathogenicity in mice. Thus, m(6)A residues in IAV transcripts enhance viral gene expression.