Figure 2: PARP inhibition causes mitotic chromatin bridges.
Throughout the figure blue bars represent BRCA1/2 proficient cells, red bars represent BRCA1/2 deficient cells, and P values were calculated using two-tailed Student’s t-test. (a) HeLa cells were transfected with indicated siRNAs and immunoblotted for BRCA2 and β-Actin levels after 48 h. Lines next to blots indicate positions of molecular weight markers. In parallel, cells were treated with DMSO or olaparib (0.5 μM) for 24 h and stained for α-Tubulin (red) and counterstained with DAPI (white). Representative immunofluorescence images are presented. Scale bars represent 5 μm. (b) HeLa cells were treated as for a. The percentages of cells containing chromatin bridges (n>20 events per condition) and lagging chromosomes (n>40 events per condition) were quantified. (c) BT-549 cells harbouring indicated shRNA vectors were pre-treated with doxycycline for 48 h and treated with olaparib (0.5 μM) or DMSO for 24 h. Percentages of cells containing chromatin bridges in anaphase and telophase (n>20 events per condition) were quantified. (d) HCC1937 cells were treated with olaparib (0.5 μM) or DMSO for 24 h. Percentages of anaphase (ana.) or telophase (telo.) cells containing chromatin bridges (n>20 events per condition) were quantified. (e) KB2P1.21 (Brca2−/−) and KB2P1.21R1 (Brca2iBac) cells were treated with olaparib (0.5 μM) or DMSO for 24 h. Percentages of anaphase or telophase cells containing chromatin bridges (left panel, n>20 events per condition) and cells containing lagging chromosomes (right panel, n>40 events per condition) were quantified. (f,g) HeLa cells were transfected with indicated siRNAs and after 24 h were treated with olaparib (0.5 μM) or DMSO and/or mirin (50 μM) for 24 h. The percentages of anaphase or telophase cells containing chromatin bridges (n>20 events per condition) were quantified. Throughout the figure ‘NS’ indicates not significant and ‘NA’ indicates not analysable. All error bars indicate s.d. of three independent experiments.
