Spike and nsp6 are key determinants of SARS-CoV-2 Omicron BA.1 attenuation.

The SARS-CoV-2 Omicron variant is more immune-evasive and less virulent than other major viral variants recognized to date 1-12 . Omicron spike (S), with an unusually large number of mutations, is considered the major driver of these phenotypes. We generated chimeric recombinant SARS-CoV-2 encoding the S gene of Omicron (BA.1 lineage) in the backbone of an ancestral SARS-CoV-2 isolate and compared this virus with the naturally circulating Omicron variant. The Omicron S-bearing virus robustly escaped vaccine-induced humoral immunity, mainly due to mutations in the receptor-binding motif (RBM), yet unlike naturally occurring Omicron, efficiently replicated in cell lines and primary-like distal lung cells. Similarly, in K18-hACE2 mice, although Omicron S-carrying virus caused less severe disease compared to the ancestral virus, it failed to achieve the attenuation level of Omicron. Further investigation showed that mutating nsp6 in addition to S was sufficient to recapitulate the attenuated phenotype of Omicron. This indicates that while the vaccine escape of Omicron is driven by mutations in S, the pathogenicity of Omicron is determined by mutations both in and outside of S.