Positions C19 and C20 in the loop of SARS-CoV-2 SL1 are required to escape NSP1-mediated repression. (A) Structures of SARS-CoV-2, bat CoV, SARS-CoV, and MERS-CoV SL1, predicted by mfold (Zuker 2003). The elements of SL1 are labeled: loop, two sections of stem 1 (stem1a and stem 1b), separated by bulge. (B) Mutagenesis of the loop shows the requirement of C19 and C20 for the derepressor function of SL1 in reporter assay. HEK293T cells were cotransfected with SL1-RL reporter or indicated SL1 mutants and the constructs encoding SARS-CoV-2 NSP1, either WT or indicated mutants. The introduced mutations (18U > A, 19C > G, 20C > G, 21C > G, [19C > G; 20C > G], [19C > G; 20C > G; 21C > G], [18U > A; 19C > G; 20C > G; 21C > G]) and resulting sequences of the loop are shown above the plots. The mutated residues are shown in red. The results are presented as in Figure 1C. Values are shown as a percentage of luciferase produced in the presence of the vector for each reporter. Values represent means ± SD from at least three experiments. P-values are calculated and presented as in Figure 1.
