FIG 7.
MHV68 infection prevents p21 induction following p53 stimulation. (A) 3T3 fibroblasts were mock infected or infected with MHV68 at an MOI of 5 PFU/cell. Twelve hours postinfection, cells were treated with DMSO, doxorubicin (Doxo; 5 μM), or nutlin-3a (10 μM) for 4 h prior to harvest. Cells were harvested in RIPA buffer 16 h postinfection, and proteins were resolved by SDS-PAGE. Immunoblot analyses were performed using antibodies to detect the indicated proteins. Detection of β-actin serves as a loading control. (B) 3T3 fibroblasts were infected with MHV68 at an MOI of 5 PFU/cell and either mock treated or treated with 5 μM nutlin-3a 14 h postinfection. RNA was isolated from cells 18 h postinfection, and quantitative RT-PCR analyses were performed using RT2 profiler p53 signaling pathway PCR arrays. Data represent the change (n-fold) in transcript levels normalized to the levels for β-actin, as determined using the ΔΔCT method for vehicle-treated versus drug-treated samples. For comparison purposes, data for uninfected cells are reproduced from Fig. 5. Data represent mean values from two independent experiments. Error bars represent the range of data.