FIG 3.

Triggering of apoptosis by IFN in IBDV-infected HeLa cells is dependent on PKR expression. HeLa cells mock infected or infected with IBDV (MOI of 2) were treated with 1,000 IU/ml of hIFN-α at 3, 6, 9, or 12 h p.i. (samples named M+3, M+6, M+9, and M+12 and I+3, I+6, I+9, and I+12, respectively, throughout the text), as indicated, or remained untreated (−) (named M and I, respectively, throughout the text), and cells were analyzed at 24 h p.i. (A) Western blot analysis of total cell extracts with different antibodies: anti-PARP, anti-t-PKR, anti-phosphorylated (Thr446) PKR, anti-t-eIF2α, anti-phosphorylated (Ser52) eIF2α, anti-Mx, and anti-VP3. Antibodies to β-actin were used for a protein loading control. The PARP cleavage product is denoted c-PARP. (B) Apoptosis was measured for duplicate samples by using the Caspase-Glo 3/7 assay kit, and each determination was carried out in duplicate. Caspase values from infected cell samples were normalized to those from mock-infected cells. Bars indicate means ± standard deviations based on duplicate samples from two independent experiments. Striped bars, infected cell samples not treated with IFN; solid bars, infected cell samples treated with IFN. * and ** indicate P values of <0.05 and <0.01, respectively, as determined by unpaired Student's t test.