FIG. 5.

Effects of NSs expression on protein synthesis. (A) Metabolic labeling of BUN-infected mammalian BHK-21 and mosquito C6/36 cells. Cells were infected at a multiplicity of infection of 5 by wtBUN or BUNdelNSs 9a; at 18 h postinfection, they were labeled with 50 μCi of [³⁵S]methionine for 2 h. Cell extracts were analyzed by SDS-PAGE (18% acrylamide gel). (B) Effects of NSs on transient reporter gene expression in mammalian BSR-T7/5 and mosquito C6-IBT7/3 cells. Cells were cotransfected with phRL-CMV (Promega) (0.5 μg per well) and pT7AcNSs at various concentrations (5, 1, 0.5, 0.1, 0.05, 0.01, 0.005, or 0.001 μg per well). Renilla luciferase reporter gene activities were determined at 16 h posttransfection by using the single Renilla luciferase assay kit (Promega).