Fig 1: Effects of endogenous CHMP4B knockdown on rSG10 virion release from cells infected at an MOI of 1 or 0.01. (A) Cell viability upon transfection with each indicated siRNA was evaluated via a CCK-8 assay. The value of the si-NC-transfected sample was set to 1. The values of si-CHMP4B- and si-CHMP4C-transfected samples were obtained by comparison with the si-NC-transfected sample. The significance of each difference was calculated by one-way ANOVA. (B) Vero cells were transfected with the indicated siRNA and harvested at 24 hpt for RNA isolation; the resulting RNA was subjected to RT-qPCR to evaluate the percentage of remaining mRNA for CHMP4B or CHMP4C after knockdown. The significance of each difference was calculated with one-way ANOVA. (C) At 48 hpt with the siRNA described above, Vero cells were harvested, and the WCLs were analyzed by Western blotting conducted using specific antibodies against endogenous CHMP4B. (D to F) After transfection with the indicated siRNAs, Vero cells were infected with the indicated rNDV at an MOI of 1 (D) or 0.01 (E and F). At 24 hpi, the extracellular viruses were collected from the infected cells and their titers were determined by titration on corresponding cells. The significance of each difference was calculated with one-way ANOVA. (G to I) The extracellular viruses described for panels D to F were also pelleted through ultracentrifugation. Equal amounts of virions and WCLs were subjected to Western blotting and the budding efficiencies were calculated as described above. The significance of each difference was calculated with one-way ANOVA. The values ± SD from triplicate experiments are shown.