Twenty natural amino acid substitution screening at the last residue 121 of influenza A virus NS2 protein reveals the critical role of NS2 in promoting virus genome replication by coordinating with viral polymerase.

adaptive mutations PA-K19E/PB1-S713N regulate virus genome replication by promoting FluPol dimerization. This work highlights the coordination between NS2 and FluPol in fulfilling efficient genome replication. It further advances our understanding of the regulation of viral RNA synthesis for influenza A virus.