Screening for influenza B virus NS1-interacting host proteins and characterization of interactions with hnRNPA0 and DDX39B.

Influenza B virus (IBV) is a type of influenza virus. The NS1 protein is a powerful regulatory factor during the process of viral infection of host cells and plays an important role in viral replication, virulence, and innate immunity. Protein-protein interactions play an extremely important role throughout the entire life cycle of viral infection of host cells. Identifying host proteins that interact with IBV NS1 protein is of great significance for exploring the pathogenic mechanism of IBV and screening for new antiviral drugs. In this study, the NS1 protein was purified by immobilized metal affinity chromatography (IMAC) using nickel-charged resin and the known host interactome of IBV NS1 was expanded using Pull-down combined with mass spectrometry (LC-MS/MS). Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Protein-Protein Interaction (PPI) analyses were conducted on the candidate-interacting proteins identified in the mass spectrometry results. These identified candidate-interacting proteins are mainly involved in biological processes such as protein translation, protein folding, mRNA processing, small molecule metabolism, ribosome biogenesis, and viral processes. The heterogeneous nuclear ribonucleoprotein (hnRNPA0) and the DDX39B protein of the DEAD-box RNA helicase family were further studied. Co-IP, IFA, and BiFC all confirmed that the NS1 protein of IBV interacts with the hnRNPA0 and the DDX39B proteins. We further mapped the interaction between the NS1-RBD and NS1-ED domains of NS1 protein and the hnRNPA0-GRD domain. These data provide resources for further research on the mechanism by which NS1 protein modulates host cells.