N6-Methyladenosine modification of hepatitis B and C viral RNAs attenuates host innate immunity via RIG-I signaling

N-6-Methyladenosine (m(6)A), the methylation of the adenosine base at the nitrogen 6 position, is the most common epitranscriptomic modification of mRNA that affects a wide variety of biological functions. We have previously reported that hepatitis B viral RNAs are m(6)A-modified, displaying a dual functional role in the viral life cycle. Here, we show that cellular m(6)A machinery regulates host innate immunity against hepatitis B and C viral infections by inducing m(6)A modification of viral transcripts. The depletion of the m(6)A writer enzymes (METTL3 and METTL14) leads to an increase in viral RNA recognition by retinoic acid-inducible gene I (RIG-I), thereby stimulating type I interferon production. This is reversed in cells in which m(6)A METTL3 and METTL14 are overexpressed. The m(6)A modification of viral RNAs renders RIG-I signaling less effective, whereas single nucleotide mutation of m(6)A consensus motif of viral RNAs enhances RIG-I sensing activity. Importantly, m(6)A reader proteins (YTHDF2 and YTHDF3) inhibit RIG-I-transduced signaling activated by viral RNAs by occupying m(6)A-modified RNAs and inhibiting RIG-I recognition. Collectively, our results provide new insights into the mechanism of immune evasion via m(6)A modification of viral RNAs.