Identification of the SARS-CoV-2 genome packaging signal in the nsp12-coding region

For SARS-CoV-2, the cis-acting packaging signal is thought to be recognized by the nucleocapsid (N) protein, but its identity and mechanistic basis for selective recognition remain undefined. Here we identify the packaging signal within the nsp12 polymerase-coding region.

CLIP-seq maps N-bound sites and, together with virus-like particle assays, pinpoints a conserved segment with strong packaging activity. An orthogonal defective-interfering RNA approach confirms its role in genome selection. We further delineate two critical subregions, α and β, that engage the N C-terminal domain.