MHV-68 de novo infection increases COX-2 protein levels and requires viral transcription. (A) COX-2 protein expression was induced during MHV-68 replication. NIH 3T3 cells in low-serum conditions were uninfected (lane 6) or infected with EGFP-MHV-68 (lanes 1 to 4). As a positive control, uninfected NIH 3T3 cells were treated with a serum shock (lane 5). Cell extracts were harvested at 1, 5, 9, or 24 h postinfection (PI) and analyzed by Western blotting using a COX-2 polyclonal antibody (top panel). Each lane contained 15 μg of total protein as determined by Bradford assays. The membrane was reprobed with antiactin providing a loading control (bottom panel). (B) COX-2 protein expression was not induced by UV-irradiated MHV-68 compared to wt MHV-68. NIH 3T3 cells in low-serum conditions were infected with wt MHV-68 (lanes 2 and 5), infected with an equal volume of UV-irradiated wt MHV-68 (lanes 1 and 4), or uninfected (lanes 3 and 6). Cell extracts were harvested 24 and 42 h postinfection for Western blot analysis using a COX-2 polyclonal antibody. Each lane contains 15 μg of total protein as determined by Bradford assays. The membrane was reprobed with antiactin, providing a loading control (bottom panel).