Targeting the m6A RNA modification pathway blocks SARS-CoV-2 and HCoV-OC43 replication

N6-methyladenosine (m6A) is an abundant internal RNA modification that influences both the fate and function of cellular mRNAs. It is installed co-transcriptionally onto pre-mRNAs by the methyltransferase METTL3 before release from chromatin. Paradoxically, the RNA genomes of several viruses that replicate in the cytoplasm are also m6A-modified, suggesting that installation can occur in both the nuclear and cytoplasmic compartments. Here we show that replication of both SARS-CoV-2, the agent responsible for the COVID-19 pandemic, and a seasonal human ?-coronavirus HCoV-OC43, can be suppressed by treatment with a highly-specific small molecule inhibitor of METTL3 or by depletion of METTL3 or cytoplasmic m6A reader proteins YTHDF1 and YTHDF3. Reduction in of infectious titer correlates with a decrease in synthesis of viral RNAs and expression of the essential nucleocapsid (N) protein. Installation of m6A on the RNAs of both viruses was mapped by methylated RNA immunoprecipitation (meRIP) sequencing and by nanopore sequencing. Although the levels of host factors involved in installation, removal, and recognition of m6A were not changed by HCoV-OC43 infection of lung fibroblasts, we show that METTL3 and cytoplasmic m6A readers YTHDF1 and YTHDF2 accumulate in the nucleus. Overall, this work establishes a positive role for m6A in control of ?-coronavirus replication that might lead to new therapeutic strategies.