Fig 5.

BAL1 limits the cellular response to DNA-damaging agents. (A) BAL1 depletion augments the cellular response to DNA-damaging agents. HeLa cells were transfected with control or BAL1 siRNAs (siRNAs 1 and 2), treated with Dox at 50 ng/ml or 200 ng/ml, or left untreated for 1 to 96 h and subsequently evaluated by an MTS assay. The consequences of BAL1 depletion were most striking in cells treated with low-dose Dox (50 ng/ml) (P < 0.001, two-way ANOVA). After 72 h of treatment with low-dose Dox (50 ng/ml), cellular proliferation (as assessed by an MTS assay) was ≈70 to 80% lower in BAL1-depleted cells than in control RNAi or parental cells. OD, optical density. (B) Cellular apoptosis following BAL1 depletion and Dox treatment. Parental, control, and BAL1 siRNA-transfected HeLa cells were untreated or treated with Dox at 50 ng/ml and 200 ng/ml for 24 h and analyzed for apoptosis with anti-annexin V and PI staining. Error bars in panels A and B represent the standard deviations (SD) of the mean for three replicates in a representative experiment. (C) Recruitment of GFP-control, GFP-BAL1, or GFP-BAL1-DM to laser-induced breaks in (5′ UTR-specific) BAL1 siRNA knockdown cells. (a to c) GFP-control, GFP-BAL1, and GFP-BAL1-DM; (d to f) PARP-1 in these cells; (g to i) merged images. (D) Apoptosis of BAL1-depleted HeLa cells repleted with GFP-control, GFP-BAL1, or GFP-BAL1-DM and subsequently treated with doxorubicin (50 ng/ml) or left untreated. Apoptosis was assessed with annexin V and PI staining. Error bars represent the SD of the mean for 3 replicates in a representative experiment.