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. 2020 Jul 30;11:3810. doi: 10.1038/s41467-020-17665-9

Fig. 3. SARS-CoV-2 ORF6 inhibits RIG-I-like signaling pathways.

Fig. 3

a Effects of SARS-CoV-2 ORF6 on SeV-induced IFN-β promoter activation. HEK293T cells were transfected with an IFN-β reported plasmid, along with a control plasmid or with increasing amounts plasmids expressing ORF6. Cells were infected with SeV for 12 h and assayed for luciferase activity. b Effects of SARS-CoV-2 ORF6 on poly(I:C)-induced IFN-β promoter activation. HEK293T cells were transfected as described in a. At 24 h post-infected, cells were transfected with high molecular weight poly(I:C) (poly(I:C)-HMW) for 12 h and assayed for luciferase activity. c–f Effects of ORF6 on RIG-IN, MDA5, MAVS, or IRF3-induced IFN-β promoter activation. HEK293T cells were transfected with an IFN-β reporter plasmid, along with a control plasmid or with increasing amount plasmids expressing ORF6, together with plasmids expressing RIG-IN (c), MDA5 (d), MAVS (e), or IRF3-5D (f). At 24 h post-transfection, luciferase activity was measured. g, h Effect of SARS-CoV ORF6 and SARS-CoV-2 ORF6 on MAVS and IRF3-5D-induced IFN-β promoter activation. 293T cells were transfected with IFN-β reporter plasmid, along with a control plasmid or plasmids expressing SARS-CoV ORF6 or SARS-CoV-2 ORF6, together with plasmids expressing MAVS (g) or IRF3-5D (h). At 24 h post-transfection, luciferase activity was measured. i Confocal immunofluorescence imaging of IRF3 and SARS-CoV-2 ORF6. HEK293 cells were transfected with a control plasmid or a plasmid expressing SARS-CoV-2 ORF6. At 24 h of post-infection, cells were infected with SeV. At 4 h of post-infection, cells were stained with indicated antibodies and subjected to immunofluorescence analyses. Red: IRF3 antibody signal; Green: ORF6 signal; Blue: DAPI (nuclei staining). Merge 1 and Merge 2 indicate the merged red and green channels and the merged red, green, and blue channels, respectively. Scale bar, 10 μm. All experiments were done at least twice, and one representative is shown. Error bars indicate SD of technical triplicates. *P < 0.05, **P < 0.01, and ***P < 0.001, two-tailed Student’s t-test. Source data are provided as a Source Data file.