Figure 6. DE proteins modulate IFN-β production and affect virus replication.

(A) H5N1 infection induces IFN-β gene expression in primary CEF cells. CEF cells were cultured and infected with H5N1 viruses, and then the total RNA extracted from the mock and virus infected cells was used for chicken IFN-β gene expression determination by qRT-PCR. The values were means ± SEs of three independent experiments. (B) The effect of DE proteins on IFN-β promoter activity induced by chicken derived MDA5. DF1 cells were transfected with 60 ng/well of pCAGGS-MDA5 (chicken), 400 ng/well of reporter plasmid (IFN-β-luc) and 10 ng/well of the pRL-TK as internal control, and 800 ng/well of indicated expression plasmids or an empty control plasmid. The cells were lysed 24 h later, and firefly luciferase and Renilla luciferase activities were determined with the Dual-Luciferase reporter assay system (Promega), according to the manufacturer’s protocol. (C,D) The effect of YWHAG and TRA2A proteins on virus replication. DF1 cells were transfected with 1 μg expression plasmid wihle transfected the pCAGGS-HA as a control group, and after 24 h, the cells were infected at a multiplicity of infection (MOI) of 0.01. At the indicated times postinfection, supernatant was collected and the viral titers were determined by plaque assay on MDCK cells.