Nucleocapsid protein captures DDX5 and RNMT facilitating viral RNA synthesis and viral protein translation for coronavirus replication.

Coronaviruses (CoVs) hijack host RNA-binding proteins (RBPs) to facilitate their replication, but the viral proteins and host RBDs that participate in the synthesis of viral RNA and protein are unclear. In this study, we revealed that DEAD-box helicase (DDX5) and staphylococcal nuclease domain-containing protein (SND1) facilitate viral RNA synthesis and that RNA guanine-7 methyltransferase (RNMT) enhances viral protein translation to promote viral replication via coronaviral subgenomic RNA-protein interactomes. DDX5 and SND1 positively regulate PEDV replication by promoting viral RNA synthesis via the binding of DDX5 to positive-sense viral RNA, whereas SND1 specifically detects negative-sense viral RNA. The interaction of DDX5/SND1 and N/nsp9/nsp12 promotes the formation of replication-transcription complexes for viral RNA synthesis to facilitate viral replication. We found that RNMT captures the host protein translation system to cyclize viral mRNA to assist in viral protein translation to promote viral replication. We also found that DDX5 broadly interacts with the N protein of CoVs and promotes the RNA synthesis of bovine coronavirus and porcine delta-coronavirus to promote viral replication. These results indicate that CoVs use host proteins to assist in the synthesis of viral RNA and protein to facilitate viral replication. IMPORTANCE: The synthesis of viral RNA and proteins is a crucial process in the life cycle of CoVs. Our observations indicate that DDX5 and SND1 facilitate the assembly of viral replication-transcription complexes and enhance viral RNA synthesis, with DDX5 binding to positive-sense RNA and SND1 binding to negative-sense RNA. Meanwhile, RNMT promotes viral protein translation by hijacking the host translation machinery and mediating the circularization of viral mRNA. These findings offer new insights into the mechanisms through which coronaviruses exploit both viral and host proteins to synthesize viral RNA and proteins.