Screening and identification of host factors interacting with the nucleocapsid protein of SARS-CoV-2 omicron variant using the yeast two-hybrid system.

Background: The nucleocapsid protein (N protein) of SARS-CoV-2 is highly conserved in viral evolution and serves as the primary structural protein in viral infection, being the most abundant in viral particles. The N protein is highly immunogenic and plays a key role in the processes of viral infection and replication, making it of significant research value in both basic studies and clinical applications.

Results: To further investigate the functions of SARS-CoV-2 N protein, the Matchmaker Gold Yeast Two-Hybrid System was used to identify potential interacting partners of the N protein in human peripheral blood mononuclear cells (PBMCs). Through this approach, we identified 11 host proteins that might interact with the SARS-CoV-2 N protein. We further validated the interaction between the N protein and two host proteins, RNF2 and ARL15, which showed the highest positive clone rates at the cellular level. We also predicted the critical amino acid residues mediating the interaction of the N protein with RNF2 or ARL15. Additionally, we explored the impact of these two host proteins on coronavirus replication. Functional analysis of all the 11 host proteins revealed their involvement in ribosome biogenesis, antigen processing and presentation, as well as various signaling pathways such as JAK-STAT.

Conclusions: This study further enriches the understanding of interactions between the SARS-CoV-2 N protein and host, providing important theoretical insights for the deeper understanding of viral pathogenesis and the development of antiviral strategies.