FIG. 5.

Ni and CE(NiN) infections prevent nuclear translocation and dimerization of IRF-3. (A) SYM-I cells were inoculated, in suspension, with each virus strain at an MOI of 2 FFU/cells and seeded. After 24 h, cells were transfected with the GFP-IRF-3 expression plasmid (green). At 24 h posttransfection, the cells were fixed and stained with an anti-N monoclonal antibody (red). (B) Assessment of the rate of GFP-IRF-3 nuclear translocation in GFP-IRF-3-expressing and virus-infected calls. Each value is the average (± the SD) of three independent experiments in which 100 cells were counted. *, Significant difference (P < 0.01). (C) Subcellular localization of endogenous IRF-3 in SYM-I cells infected with each strain. SYM-I cells were mock infected or infected with Ni, Ni-CE, or CE(NiN) strains at an MOI of 2. After 24 h, cells were fixed and examined by double immunofluorescence staining using anti-IRF-3 polyclonal antibody (green) and anti-N monoclonal antibody (red). (D) Extracts from SYM-I cells infected with Ni, Ni-CE, and CE(NiN) strains for 24 h were analyzed by native gel electrophoresis, followed by Western blotting, to detect IRF-3. N protein and tubulin of same samples were detected by SDS-PAGE, followed by Western blotting.