Figure 4

53BP1 depletion rescues RAD51 foci in Brca1 null cells. (a) RAD51 foci formation in R26^CreERT2;Brca1^SCo/Δ ES cells untreated or treated with 4OHT and transduced with lentiviruses expressing non-targeting shRNAs (NTsh) or shRNAs targeting 53bp1(53BP1sh). Cells were irradiated with 10 Gy, fixed after 6 hours, and stained with anti-RAD51 antibody (red). Nuclei were visualized with DAPI (blue). (b) RAD51 foci formation in Brca1^SCo/Δ MEFs infected with retroviruses expressing self-deleting Cre recombinase (+Cre) or empty vector (−Cre), together with retroviruses expressing 53BP1 or GFP control shRNAs. Cells were irradiated with 10 Gy, fixed after 2 hours, and stained with anti-γH2AX (red) and anti-RAD51 antibodies (green). Nuclei were visualized with DAPI (blue). (c) Quantification of RAD51 foci in R26^CreERT2;Brca1^SCo/Δ ES cells treated as in (a). The percentage of cells with ≥10 Rad51 foci per cell are plotted for each condition. At least 20 nuclei were analyzed for each treatment. (d) Quantification of RAD51 foci in Brca1^SCo/Δ MEFs treated as in (b). The percentage of cells with ≥10 RAD51 foci per cell are plotted for each condition. At least 50 nuclei were analyzed for each treatment.