Abstract
N6-methyladenosine (m6A) epitranscriptomic modifications play crucial roles in regulating both host and viral gene expression. Here, we revealed a novel mechanism by which paramyxoviruses exploit host m6A machinery to simultaneously enhance viral replication and suppress host immunity. Our results demonstrated that the viral matrix protein (M) of bovine parainfluenza virus type 3 (BPIV3) binds to the methyltransferase domain of METTL3 in the nucleus and facilitates its translocation to the cytoplasm through an exportin-1-dependent pathway. This mechanism is conserved across multiple paramyxoviruses, including human parainfluenza virus type 3, Sendai virus, Nipah virus, and measles virus, suggesting an evolutionarily conserved viral strategy. The relocated METTL3 catalyzes m6A modification at specific sites within viral nucleocapsid protein (N) mRNA, significantly enhancing its stability and protein expression. Using reverse genetics, we generated recombinant viruses harbouring mutations at these m6A acceptor sites, which exhibited markedly attenuated viral replication, confirming the critical role of these epitranscriptomic marks in the viral life cycle. Rescue experiments demonstrated that the expression of exogenous N protein partially restored the viral titer and concomitant genome/antigenome synthesis in m6A site mutant, indicating that reduced N protein abundance represents a key mechanism underlying impaired viral replication. Furthermore, M protein-mediated depletion of nuclear METTL3 significantly reduces m6A modification of host IFN-β mRNA, resulting in diminished interferon expression and compromised antiviral responses. Supporting this mechanism, infection with viruses bearing nuclear export signal mutations that prevent METTL3 cytoplasmic translocation, maintained IFN-β mRNA m6A modification and resulted in significantly elevated IFN-β expression. These findings provide direct mechanistic evidence that paramyxoviruses utilize M-driven METTL3 relocalization as a sophisticated immune evasion strategy. Our study illuminates how paramyxoviruses strategically manipulate epitranscriptomic regulation to create an environment conducive to viral propagation, thereby advancing our understanding of virus-host interactions and identifying potential targets for antiviral therapeutics.