Extended Data Fig. 9: RBD mutations modulate hNEC infectivity.
From: Phenotypic evolution of SARS-CoV-2 spike during the COVID-19 pandemic
a, Replication kinetics of the SARS-CoV-2 spike mutants in human nasal epithelium (hNEC). Graphs represent data collected using an independent set of clones for each SARS-CoV-2 mutant (n = 3) as in Fig. 5c. S:B.1 (indicated by light grey area) and S:BA.1 (dashed purple line) viruses were used as reference viruses in all experiments. The copy numbers of viral RNA in the culture supernatant of infected hNECs were quantified by RT-qPCR at the indicated timepoints. Data are mean ± standard error to the mean (SEM). a, statistical significance between S:B.1 and the mutant spike virus panel across time points was determined one-way ANOVA with multiple comparisons between area under the curve of the different viruses and S:B.1 using the Holm-Ĺ Ădák post-hoc test. The calculated p-values are indicated in the figure. b, Ribbon diagram molecular structure of spike98 (based on PDB:6VSB, one RBD up conformation99) illustrating the location of the RBD-4 mutations. Left, full spike context; inset, zoom on the apex of spike. c, Top-down view of BA.1 spike trimer (PDB:7TF8117) individual protomers are colour-coded. d, Inset, as marked in (c), displaying intra-protomer interactions mediated by BA.1 RBD mutations. Side chains are shown for interacting residues with contacts denoted by dashed grey lines. Interacting residues are labelled and color-coded by protomer, as in (c). Previous cryo-electron microscopy analysis indicates that the F371, P373 and F375 mutations modulate inter-protomer RBD-RBD100, for instance by mediating interaction with the RBM of the adjacent protomer (for example H505-P373). It is likely that this will affect the conformational plasticity of the RBD (for example up/down switching), which in turn may impact ACE2 acquisition and spike stability. Reverting these residues in the context of BA.1 evidently creates incompatibilities that prevent hNEC infection; however, determining precisely how this relates to spike conformation and mechanism will require further investigation.
