FIG 8.

Genetic deletion of PERK enhances the antiviral effect of 6-TG. (A) Western blotting of PERK KO cells (clone B3) and nontargeting control gRNA lentivirus-transduced cells (NT) treated with 1 μM thapsigargin (Tg) or 500 μM sodium arsenite (As). Lysates were collected at 1 h post-treatment and analyzed for PERK expression and activation and total and phosphorylated eIF2α. (B) A549 PERK KO cells (clones A2, B3, and C3) and the nontargeting control cell line were infected with PR8 at an MOI of 0.1 and treated with tunicamycin (TM) (5 μg/ml), 6-thioguanine (6-TG) (10 μM), and 6-mercaptopurine (6-MP) (10 μM) for 23 h. The supernatant was collected at 24 hpi, and viral progeny were quantified by plaque assay. Statistical significance was calculated via two-way ANOVA followed by a Dunnett multiple-comparison test (only a subset of statistically significant differences is highlighted with asterisks for clarity). (C) A549 PERK KO clone B3 and nontargeting control cells were treated with escalating doses of 6-TG, 6-TGo, or the vehicle control for 23 h, and cell viability was measured using an alamarBlue assay. Relative fluorescence units were normalized to the vehicle control. Error bars represent standard deviations (n = 3).