Figure 1.

SARS-CoV-2 M^proinhibits interferon-stimulated gene production.A, HEK-293T cells were transfected with pCDNA3.1 empty vector or the pCDNA3.1-M^pro-HA plasmid. Twenty-four hours posttransfection, cells were stimulated with 1000 U/ml IFNα for 8 h as indicated. Isolated RNA was analyzed by RT-PCR for the ISG15, ISG56, IFIT3, or OAS1 relative mRNA level and normalized to GAPDH mRNA transcription. B, HEK-239T cells, Vero E6 cells, or A549 cells were transfected with various concentrations of pCDNA3.1-M^pro-HA plasmid, along with pRL-TK plasmid (10 ng) and pISRE-Luc (50 ng) or pISG15-Luc. Twenty-four hours posttransfection, cells were stimulated with or without IFNα (1000 U/ml) for 12 h, followed by a dual-luciferase assay. C, schematic representation of SARS-CoV-2 M^pro (PDB code: 7JYC) with the conserved catalytic residues His41 and Cys145. D, HEK-293T cells were transfected with pISRE-Luc plasmid (50 ng), pRL-TK plasmid (10 ng), along with SARS-CoV-2 M^pro expression constructs (200 ng) or its inactive double mutants H41A and C145A. Twenty-four hours posttransfection, a dual-luciferase assay was performed after treatment IFNα (1000 U/ml) for another 8 h. Western blotting for the expression levels of M^pro were shown below the graph. All presented results represent the means and standard deviations of data from three independent experiments. Statistical significance was calculated using unpaired, two-tailed Student’s t test. Data significance is shown as indicated. ns, not significant; ∗P < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. ∗∗∗∗p < 0.0001. ISG, interferon-stimulated gene; Mpro, main protease; OAS, 2′,5′-oligoadenylate synthetase; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.