FIG 8.

Activation of MAPK/ERK pathway is not related to the inhibition of IFN-α/β signaling by BTV-NS3. (A) HEK-293T cells were cotransfected with IFN-β-pGL3 plasmid that contains the firefly luciferase reporter gene downstream of an IFN-β-specific promoter sequence, pRL-CMV reference plasmid, and pCI-neo-3×FLAG expression vectors encoding 3×FLAG alone or fused to NΔRIG-I (N-terminal CARDs of RIG-I) and BTV-NS3_FL, BTV-NS3A, or fragments. After 48 h, the relative luciferase activity was determined. ***, Differences observed between NS3_FL, NS3A, NS3_1–182, NS3_118–229, or NS3_Δ118–182 and the corresponding control vector pCI-neo-3×FLAG were statistically significant P < 0.0005; ns, nonsignificant differences between NS3_1–117 and the corresponding control vector pCI-neo-3×FLAG. (B) Same experiments as in panel A but, at 24 h posttransfection, the cells were serum starved for 6 h and then treated with 20 μM U0126, as indicated. After 24 h, the relative luciferase activity was determined. All experiments were performed in triplicates, and data represent means ± the SD. ND, not determined.