FIGURE 4.

T2AA induces DSB response. A, DSB quantification by neutral comet assay. U2OS cells were exposed to cisplatin (30 μm) for 1 h followed by DMSO or T2AA (15 μm) and cultured for 72 h. Cell electrophoresis and comet tail scoring were performed. Representative comet tail images and the comet tail score for each treatment are shown. B, phospho-Ser-1524 BRCA up-regulation in U2OS cells after being treated as indicated for 3 days. Actin served as a loading control. C, chromatin co-foci of phospho-Ser-1981 ATM and 53BP1 in HeLa cells were examined by treating cells with or without cisplatin (15 μm) for 1 h followed by DMSO or T2AA (15 μm) and culturing for 72 h. Cells were immunostained with anti-phospho-Ser-1981-ATM (green) and anti-53BP1 (red) antibodies. Frequency of positive colocalization (percentages of cells exhibiting more than 10 yellow foci in the nucleus (arrows) on confocal images) was 42% for cisplatin/T2AA cotreatment, 0% for other treatments. D, recombination activity assay as described by Jasin and co-workers (16) after cells were cultured with DMSO or T2AA (10 μm) for 72 h.