RBM6-mediated alternative splicing facilitates the adaptation of Eurasian avian-like H1N1 swine influenza virus

The Eurasian avian-like (EA) H1N1 swine influenza virus (SIV), derived from avian influenza viruses (AIV), poses a serious threat to public health due to its capacity for cross-species transmission and pandemic emergence. The molecular determinants underlying its replication advantage over AIV remain poorly defined. Here, we identify RNA-binding motif protein 6 (RBM6) as a novel host factor that differentially regulates the replication of EA H1N1 SIV and AIV. Mechanistically, RBM6 binds to the critical M901 site of the viral M segment, thereby modulating RNA splicing. Substitution of M901C with M901T markedly reduced RBM6 binding, impaired M segment splicing, and attenuated viral replication both in vitro and in vivo. Conservation analysis revealed that M901T is common in avian strains, whereas M901C is predominantly maintained in swine strains, underscoring M901C as a determinant of swine adaptation. Complementation experiments further demonstrated that swine RBM6, but not avian RBM6, restored EA H1N1 SIV replication. Taken together, our findings uncover a previously unrecognized role of RBM6 in shaping influenza virus replication and highlight the RBM6-M901C axis may serve as potential targets for controlling influenza virus adaptation and interspecies transmission.