hMPV M2-2 protein modulates host mRNA alternative splicing to favor virus replication

Human metapneumovirus (hMPV) is a widespread and prevalent respiratory pathogen, yet the mechanisms by which hMPV modulates host gene expression to facilitate replication remain incompletely characterized. Here, we systematically profiled hMPV–host protein–protein interactions by affinity purification-mass spectrometry and assessed their impact on host mRNA splicing using RNA sequencing, complemented by genetic perturbation, protein interaction assays, and fluorescence microscopy–based subcellular localization and colocalization analyses. Among the viral proteins analyzed, the regulatory protein M2-2 was found to directly interact with multiple splicing factors. Transcriptomic analysis revealed that M2-2 triggers extensive changes in alternative splicing across host genes, notably affecting STRAP and TANC2 , both of which exhibit distinctsplicing patterns during hMPV infection. Knockdown of M2-2 using short hairpin RNA (shRNA) during hMPV infection reversed M2-2–driven changes in the splicing patterns of STRAP and TANC2 , confirming its regulatory role. Functional knockout of STRAP or TANC2 significantly impaired viral replication, indicating that the M2-2–favored isoforms are required for efficient viral propagation. Collectively, our findings demonstrate a novel mechanism by which hMPV M2-2 hijacks the host splicing machinery to reprogram key factors that facilitate viral replication. This extends the functional repertoire of M2-2 and highlights the virus–spliceosome interface as a potential therapeutic target for anti-hMPV intervention. • Human metapneumovirus M2-2 recruits heterogeneous nuclear ribonucleoproteins (hnRNPs) to hijack the host splicing machinery. • hMPV M2-2 reprograms the alternative splicing of the cellular genes STRAP and TANC2 to favor viral replication. • Targeting the M2-2–spliceosome axis represents a promising therapeutic strategy against hMPV.