Fig 7. Individually expressed viral proteins block transcription and translation.
(A) Huh7 cells were transfected with indicated plasmids and lysates were collected at 18h post transfection. Western blots for HA-tag were performed as described in Materials and Methods. (B and C) Huh7 cells were transfected with plasmids coding for indicated viral proteins and labeled with 100μCi/ml of [35S] Cys/Met for 2h at 8-24h post transfection. Lysates were collected and resolved on SDS-PAGE gels and visualized as described in Materials and Methods. (B) Representative image of nsP and capsid induced shutoff compared to GFP control. (C) Densitometry was performed to quantify the extent of shutoff following transfection of indicated plasmids. ****, P<0.0001; **, P<0.01 using t-test and One-way ANOVA. Ns = not significant. Data is the average of four independent experiments. All error bars are standard deviations. (D) Huh7 cells were transfected with plasmids coding for indicated viral proteins and lysates were collected in triplicate at 12h post transfection. RT PCR for human gamma actin intron #3 was performed as described in Materials and Methods. ****, P<0.0001; ***, P<0.01; **, P<0.02 using t-test. Ns = not significant. Data is representative of four independent experiments. All error bars are standard deviations.