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. 2021 Nov 15;12:6602. doi: 10.1038/s41467-021-26910-8

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — a and b Effect of SARS-CoV-2 proteins on the IFNγ-mediated IRF1 GAS or NLRC5 promoter activity. Schematic images illustrate the reporter assay systems for monitoring the promoter activity of IRF1 GAS and NLRC5. HEK293T cells were transfected with IRF1 3 × GAS- (a) or NLRC5 promoter-containing reporter construct (b), along with an empty control plasmid or with plasmids expressing the indicated SARS-CoV-2 proteins. At 24 h after transfection, cells were treated with IFNγ for 9 h, and luciferase activity was measured. The results are from three independent experiments. c Quantitative real-time PCR analysis of NLRC5 gene expression level in IFNγ (100 U/ml, 9 h) treated HEK293T cells expressing the indicated SARS-CoV-2 proteins. The results are from three independent experiments. d Effect of SARS-CoV-2 ORF6 on the IRF1-mediated HLA promoter activity. HEK293T cells were transfected with the indicated HLA reporter constructs, along with a IRF1 expression vector and the plasmid expressing empty or SARS-CoV-2 ORF6. At 36 h after transfection, cells were collected to measure the luciferase activity. The results are from three independent experiments. e Immunofluorescence analysis of the endogenous IRF1 cellular localization in IFNγ (100 U/ml, 6 h) treated HEK293T cells expressing the indicated SARS-CoV-2 proteins. Images were obtained using a confocal microscope. The scale bar indicates 50 microns. Suppressed expression and nuclear import of IRF1 is indicated with white arrows. A quantitative comparison of the nuclear IRF1 signal intensity (%) is shown with a bar graph using ImageJ.