Figure 2

Translational arrest is inhibited by ablation of PKR expression. (A) MOSEC cells are efficiently transfected with GFP control vector. At 48 hours post transfection MOSEC cell were harvested for FACS analysis monitoring FL-1 for GFP expression. (B) Cells transfected with GADD34, but not the GADD34 PP1c deletion mutant, can dephosphorylate eIF2α. MOSEC cells transfected with GADD34, PP1c mutant or GFP control vector were infected at 48 hours. 8 h.p.i. lysates were collected and subjected to western blotting for phospho-eIF2α. In panel B immunoblot was stripped and reprobed with β actin for loading control. (C) Transfection of GADD34 but not the PP1c mutant alleviates Sindbis vector-induced translational arrest. MOSEC cells transfected with GADD34, PP1c mutant or GFP control vector were infected at 48 hours. 24 h.p.i. cells were subjected to ³⁵S labeling. (D) Dephosphorylation of eIF2α by GADD34 but not the GADD34 mutant lacking the PP1c interacting domain, increases the cell viability at 24 h.p.i. MOSEC cells were transfected with either GADD34, GADD34 PP1c mutant or GFP control vector. 48 hours after transfection cells were infected with SV. 24 h.p.i. cell viability was assessed. Data in D represents the SEM (error bars) of three experiments. Cell viability for each sample was compared to the infected control at the same time point and was corrected for the percentage of infected cells. Statistical significance was calculated by a two-tailed student t-test (** P < 0.005).