Figure 5

DNA damage upregulates ERIC RNA levels, and ERIC restricts DNA damage-induced apoptosis. A) U2OS cells were treated with etoposide (50 μg/ml) for times indicated. RNA was extracted from cells and ERIC RNA levels were determined by real-time RT-PCR and normalized to GAPDH. One representative experiment out of 4 is shown. Inner bar-graph indicates percent of cells exhibiting subG1 DNA content at each time point, as analyzed by FACS.

U2OS cells were transfected with either a nonspecific siRNA (NS) or an siRNA directed against ERIC (si#1 or si#2). Then, cells were left untreated or incubated with etoposide (Etop) (50 μg/ml) for 24 hours. B) Upper panel- RNA was extracted and ERIC RNA levels were determined by real-time RT-PCR and normalized to GAPDH levels. Lower panel- Proteins were extracted from cells and western blot analysis performed using antibodies directed against E2F1 and GAPDH. C) Cells were analyzed by FACS following propidium-iodide (PI) staining. One representative experiment out of 3 is shown. Numbers represent percent of cells with a subG1 DNA content. D) Summarized results of three independent FACS experiments, *P value = 0.01 (Two-tailed Student’s T-test).