FIG 7.

Characterization of C9 protein and IFN resistance in C9-expressing cells. (A) Predicted domain structure of C9 protein showing ankyrin (ANK) repeats and the C-terminal F-box domain. (B) Expression of HA-tagged C9 protein. A549 cells were mock infected or infected with vC9rev or vHA-C9 in the absence or presence of AraC. At the indicated hours postinfection (h.p.i.), the cells were harvested, and lysates were analyzed by Western blotting with antibody to the HA epitope, VACV, or actin. (C) Ectopic expression of 2×Myc-C9 overcomes the IFN-β-sensitive phenotype of vΔC9. A549 control cells and A549 cells stably expressing 2×Myc-C9 were untreated (lanes −) or treated with 2,000 IU/ml of IFN-β for 24 h (lanes +) and then were mock infected or infected with 0.01 PFU/cell of purified WR or vΔC9. At 48 h after infection, the lysates were analyzed by Western blotting using antibody to VACV, A3, L1, the Myc epitope, and actin. The positions of protein markers (in kilodaltons) are shown on the right of panels B and C.