FIG. 4.

SARS-CoV ORF 3b, ORF 6, and N proteins inhibit activation of IRF-3. A. 293T cells were cotransfected with the p55-CIB promoter, which contains three IRF-3 binding sites, a plasmid that constitutively expresses Renilla luciferase, and plasmids expressing SARS-CoV proteins or the indicated control plasmids. Cells were infected with Sendai virus 24 h posttransfection. Cells were harvested at 24 h postinfection and analyzed for firefly and Renilla luciferase. Data were normalized using the Renilla luciferase values. Data are averages plus standard deviations for three experiments. A value of 100% represents approximately 10,000,000 firefly luciferase units(p55-CIB) and 6,000,000 Renilla luciferase units. B. 293T cells were transfected with the indicated plasmid and infected with Sendai virus for 6 h. Cells were harvested, and lysates were analyzed by Western blotting using a phospho-IRF-3 antibody. Quantitations are given below each band and are percentages of the value for the empty, infected control. C. 293T cells were transfected with the indicated plasmids for 24 h and then infected with Sendai virus. Cells were fixed 8 h postinfection and analyzed by microscopy using an antibody that recognizes the HA tag.