Figure 6.

The PKR mutant K296H increases the viral protein accumulation caused by the dsRNA binding domain. (A) Schematic representation of the PKR mutants. (B) Plasmids encoding vector alone, PKR, K296H, K64E/K296H, A67E/K296H, or K64E/A67E/K296H were transfected into RD cells, which were then selected by the addition of 3 μg/mL puromycin. Stable expressing cells were infected with EV-A71 at an m.o.i of 10 and the cellular extracts were harvested after 8 h. Immunoblot analysis was performed to detect PKR, viral 3A protein, and GAPDH. The relative amount of 3A protein was quantified by densitometry and was first normalized to GAPDH and then to that of the vector control. A representative result based on three independent experiments is shown.