FIG. 6.

Induction of cytokine expression and PKR phosphorylation by RNA associated with vaccinia virus infection. (A) Confluent HeLa cells were infected with vvΔE3L-Rev or vvΔE3L at an MOI of 5 and collected at early (2 hpi) or late (8 hpi) times postinfection. Cells were also pretreated with 50 μg/ml of araC 1 h prior to infection, infected with vvΔE3L-Rev or vvΔE3L, and collected at 8 hpi. Total RNA samples were extracted with TRIzol and digested with DNase. Five micrograms of each RNA sample or 1 μg pI or pIC was then transfected into HeLa cells. RNA samples were also pretreated prior to transfection with RNase III. Cells were collected at 6 h posttransfection, and cytokine expression was examined by RT-PCR. (B) HeLa cells were transfected with the RNA samples described for panel A and collected at 6 h posttransfection. The levels of β-actin, PKR, and phosphorylated PKR were examined by Western blotting.