Figure 3.

IRF3 activation is impaired due to inhibition of the RIG-I-TRIM25 pathway. Lenti-X 293T cells were co-transfected with the IFN-β-Luc Firefly luciferase reporter plasmid and the Renilla luciferase control reporter plasmid along with the IRF3-expressing plasmid. Where indicated, co-transfections were performed, including expressing plasmids for RIG-I, TRIM25 or SARS-CoV-2 N. Cells, previously stimulated for 12 h with poly(I:C), were collected at 48 h post-transfection. The luciferase activity of the cell lysates was analyzed with the dual luciferase reporter assay system (Promega). Fold changes in IFN-β promoter activation were calculated with respect to the relative reference sample based on Firefly luciferase values normalized to Renilla luciferase activities. Results were plotted as mean values (n = 4) ± standard deviation (SD) and significance was determined by p-value (*** p < 0.005, 0.005 < ** p < 0.01). Cell lysates from reporter gene assays were analyzed by immunoblotting.