Fig. 2.

Caspase-2 activation by AD169 is cell cycle-dependent. (A) MRC-5 fibroblasts were pretreated with a mitotic entry inhibitor (RO-3306, 10 µM) or a mitotic exit inhibitor (nocodazole, 0.66 µM) for 1 h. Cells were infected with HCMV AD169 (MOI 5) and collected at 24 hpi. Caspase-2 activation was analyzed by immunoblot. IE1 and β-actin were used as infection and loading controls, respectively. (B) MRC-5 fibroblasts were seeded and synchronized according to a previously described protocol (77) with a few modifications. Asynchronous (asyn) cells were infected before serum starvation. Synchronized cells were infected in G0, early G1 (E-G1), late G1(L-G1), or G2/M phase. (C) Synchronized cells were collected, fixed with ethanol, stained with propidium iodide (PI), and analyzed by flow cytometry. (D) Synchronized cells infected with HCMV AD169 for 24 h were analyzed for caspase-2 activation by immunoblot. The data shown in this figure are representative of 2 independent experiments.