FIG 2.
Lytic replication is not altered by the expression of IKKα-SA in murine fibroblast cells or the lungs of infected mice. (A) MEFs were generated from CreERT2/IKKαfl/fl mice and treated with tamoxifen (Tam) daily for 8 days to drive IKKα deletion. Cells were lysed and probed for IKKα protein to verify the loss. GAPDH was used as a loading control. (B) IKKα−/− and WT MEFs generated as described above for panel A were infected with WT MHV68 at an MOI of 5. Virus replication was measured by plaque assays at 24 and 48 hpi. Time points were measured in triplicate. Data were analyzed by an unpaired t test. n.s., not significant (P > 0.05). (C) Murine embryonic fibroblasts were infected at an MOI of 0.05, and cell lysates were harvested at the indicated time points. Viral titers were determined by plaque assays and measured in triplicate. Results were analyzed using two-way ANOVA with Tukey’s multiple comparisons. WT MHV68 differs from other viruses at multiple time points (P < 0.05) due to input differences. The WT differs from IKKα-SA.STOP2 at 12 hpi, all viruses at 24 hpi and 80 hpi, IKKα-SA1 and -2 and IKKα-SA.STOP1 at 48 hpi, and IKKα-SA.STOP2 at 144 hpi. IKKα-SA2 differs from IKKα-SA.STOP1 at 104 hpi. (D) C57BL/6 mice were infected with 1,000 PFU of the indicated viruses via the intranasal route. Lungs were removed and homogenized at the indicated times postinfection, and virus titers were determined by plaque assays. Symbols represent PFU per milliliter of lung homogenate for individual mice, bars represent the mean titers ± standard deviations (SD), and the dashed line indicates the limit of detection (50 PFU/mL). “WT” indicates BAC-derived MHV68. No significant differences were found by one-way ANOVA for each time point.