Extended Data Fig. 11: Recombinant MERS-CoV with a single amino acid change at the caspase-6 cleavage motif of N protein.
From: Coronaviruses exploit a host cysteine-aspartic protease for replication
a rMERS-CoV/TKKA is constructed by introducing a D to A change at the TKKD motif of the MERS-CoV N gene with red recombineering. b Huh7 cells were infected with rMERS-CoV/TKKA or rMERS-CoV/TKKD at 1MOI and cell lysates were harvested at 1, 24, and 48 hpi for Western blot analysis of N protein. The experiment was repeated two times independently with similar results. c A549, Huh7, and VeroE6 cells were infected with rMERS-CoV/TKKA or rMERS-CoV/TKKD at 1MOI (A549 and Huh7) or 0.5MOI (VeroE6). MERS-CoV N gene copies in the cell lysate samples at the indicated time points were quantified with RT-qPCR (n = 3). d Huh7 and A549 cells were infected with rMERS-CoV/TKKA or rMERS-CoV/TKKD at 1MOI and treated with z-VEID-fmk at the indicated concentrations. MERS-CoV N gene copies were quantified with RT-qPCR (n = 3). e hDPP4 KI mice were intranasally inoculated with 3.5x104PFU rMERS-CoV/TKKA or rMERS-CoV/TKKD. Mouse lung samples were harvested at day 2 and day 4 post infection. Representative images of H&E stained lungs from rMERS-CoV/TKKA- or rMERS-CoV/TKKD-infected hDPP4 KI mice were shown. The experiment was repeated two times independently with similar results. Data represented mean and standard deviations from the indicated number of biological repeats. Statistical significance between groups was determined with two-way ANOVA (c, d). Scale bar in (e) represented 500 μm.
