Fig. 3: The efficient furin CS-dependent entry of SARS-CoV-2 is due to TMPRSS2 and allows for subsequent escape from IFITM3.
From: The furin cleavage site in the SARS-CoV-2 spike protein is required for transmission in ferrets
a, Relative lentiviral PV entry into 293T cells expressing ACE2-FLAG with or without co-expression of TMPRSS2. Entry into cells not transfected with TMPRSS2 was normalized to 1. Assays were performed in triplicate and are plotted as mean + s.d. Data shown are a representative replicate (n = 4). Statistics were determined by multiple t-tests. ****P ≤ 0.0001. b,c, Replication kinetics of SARS-CoV-2 WT and ΔCS viruses in Vero E6 (b) and Vero E6/TMPRSS2 (c) cells at an MOI of 0.1. Assays were performed in triplicate and are plotted as mean + s.d. Statistics were determined by one-way ANOVA with multiple comparisons on log-transformed data. d–f, Relative PV entry into 293T-ACE2 (d), Caco-2 (e) or Calu-3 (f) cells pretreated with amphoB (pink bars). Entry into untreated cells was normalized to 1 (black bars). Assays were performed in triplicate and are plotted as mean + s.d. Data shown are a representative replicate (n = 4). Statistics were determined by multiple two-tailed t-tests. *0.05 ≥ P > 0.01; **0.01 ≥ P > 0.001. g, Relative PV entry into 293T cells overexpressing ACE2-FLAG and TMPRSS2, with or without IFITM3. Entry into cells not transfected with IFITM3 was normalized to 1 (black bars). Assays were performed in triplicate and are plotted as mean + s.d. Data shown are a representative replicate (n = 4). Statistics were determined by multiple two-tailed t-tests. **0.01 ≥ P > 0.001; ***0.001 ≥ P > 0.0001; ****P ≤ 0.0001. h,i, Replication kinetics of SARS-CoV-2 WT and ΔCS viruses in Calu-3 (h) or HAE (i) cells. Cells were pretreated with control media or media containing amphoB for 1 h then infected at an MOI of 0.05 (Calu-3) or 0.1 (HAE). Assays were performed in triplicate and are plotted as mean + s.d. The HAE assay was performed with three separate donors, with data from a representative donor shown. Statistics were determined by one-way ANOVA with multiple comparisons on log-transformed data. Black P values indicate statistical significance between no drug controls of WT and ΔCS, while coloured P values indicate significance between no drug control or amphoB. Vehicle controls were the same as from d. Dotted lines in b, c, h and i indicate limits of detection.
