Tyrosine kinase LYN restricts the replication and virulence of influenza A virus by directly phosphorylating viral nucleoprotein

The continuous antigenic shift and drift of influenza A virus (IAV) result in the emergence of novel strains and drug resistance. Accordingly, it is urgent to develop novel antiviral drugs that target host factors. Here, we revealed that tyrosine kinase LYN (Lck/Yes-related novel protein tyrosine kinase) interacts directly with nucleoprotein (NP) and reduces the replication and virulence of IAV <i>in vitro</i> and <i>in vivo</i>, in a kinase-dependent manner. By directly catalyzing the tyrosine phosphorylation of NP at Y10/40/97 sites, LYN impairs the interaction of NP with viral RNA and polymerase proteins, as well as the oligomerization of NP, and thus negatively modulates the assembly of the viral ribonucleoprotein complex. The NP^Y10/40/97F mutation significantly increases the pathogenicity of IAV in mice. Furthermore, the LYN-specific agonist MLR-1023 showed promising therapeutic effects against IAV. Collectively, our findings suggest that LYN is a novel host kinase restricting IAV replication, and a promising target for anti-influenza drug development.IMPORTANCEThe nucleoprotein (NP) of influenza A virus (IAV) is a highly conserved, multifunctional, and the most abundant viral protein in infected cells, which makes NP a promising target for the development of anti-influenza drugs. Phosphorylation plays a crucial role in the stability and functionality of NP. However, few kinases have been identified that directly phosphorylate NP. In this study, LYN (Lck/Yes-related novel protein tyrosine kinase) was identified as a novel kinase that directly catalyzes the tyrosine phosphorylation of IAV NP and thus restricts the replication and virulence of IAV. The LYN-specific agonist MLR-1023 exhibited promising protective efficacy against lethal IAV infection. Our findings discovered LYN and LYN-agonist as potential drug candidates targeting IAV NP, which will facilitate the development of host-directed antiviral therapies.