The R203M and D377Y mutations of the nucleocapsid protein promote SARS-CoV-2 infectivity by impairing RIG-I-mediated antiviral signaling

PLoS Pathog. 2025 Jan 22;21(1):e1012886. doi: 10.1371/journal.ppat.1012886. Online ahead of print.

Abstract

The viral protein mutations can modify virus-host interactions during virus evolution, and thus alter the extent of infection or pathogenicity. Studies indicate that nucleocapsid (N) protein of SARS-CoV-2 participates in viral genome assembly, intracellular signal regulation and immune interference. However, its biological function in viral evolution is not well understood. SARS-CoV-2 N protein mutations were analyzed in Delta, Omicron, and original strains. Two mutations with a methionine (M) residue at site 203 and a tyrosine (Y) residue at site 377 of the N protein were found in Delta strain but not in Omicron and original strains, and promoted SARS-CoV-2 infection therein. Those mutations, R203M and D377Y, enhanced the inhibitory impact of N protein on the impairment of RIG-I-mediated antiviral signaling, such as IRF3 phosphorylation and IFN-β activation. The viral RNA-binding activity of N protein was promoted by these mutations, effectively attenuating the recognition and interaction of RIG-I with viral RNA compared to the original or other variants. The R203M/D377Y mutations thus enhanced the suppressive activity of the N protein on RIG-I-mediated interferon induction both in vitro and in vivo, which in turn promoted viral replication. This study helps to understand the variability of SARS-CoV-2 in regulating host immunity.