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. 2020 Mar 30;48(9):4915–4927. doi: 10.1093/nar/gkaa196

Figure 1.

Figure 1.

PHF2 knockdown alters 53BP1 and BRCA1 focus dynamics in response to IR. (A) U2OS cells were depleted for Luciferase (Luc), CtIP or PHF2 by siRNA. After 48 h, cells were treated with IR (10 Gy) and fixed after 1, 4 or 7 h. 53BP1 focus formation was analysed by immunofluorescence. Left panel: representative images. Right panel: quantification of three independent experiments with each at least 100 cells. (B) U2OS Rap80 knockout cells were depleted for Luc or PHF2. Cells were treated with IR (10 Gy), fixed at the indicated time points and γH2AX (positive control for DNA damage induction) and BRCA1 focus formation was analysed as in (A). Left panel: representative images (IR, 4 h). (C) U2OS cells depleted for Luc or PHF2 were subjected to different DNA damaging agents and lysed after 1 h. Extracts were analysed by western blot using the indicated antibodies. (D) U2OS cells were transfected as in (C), treated with CPT or ETP and analysed by western blot with the indicated antibodies. (E) U2OS cells were depleted for CtIP or PHF2 by siRNA. Cells were treated with IR (3 Gy), fixed at the indicated time points and RPA2 focus formation was analysed as in (A). Left panel: representative images (IR, 1 h).