(A) Wound-healing assays of MDA-MB-231 cells untreated (Unt) or treated with doxorubicin (Dox, 100 nM), phleomycin (Phleo, 90 μg/ml), etoposide (Et, 40 μM) or ionizing radiation (IR, 20 Gy). Drug treatments were for 48 hr. IR treatment was performed at time 0. All samples were analyzed post-48 hr from wound induction. Images were acquired at 0 and 48 hr. Representative images from three independent experiments are shown. (B) Verification of DNA damage induction and quantification of wound healing from (A). Top: Western blot analysis of samples from (A) with the DNA damage marker γH2AX. H2AX is a loading control. Bottom: Quantification of wound healing experiments from (A). (C) Cell cycle analysis of cells treated in panel A by flow cytometry. Cells were treated as in (A) and analyzed by FACS 48 hr post-treatment. (D) Proliferation of cells treated as in (A). After 48 hr, cells were trypsinized, counted and normalized to untreated cells at 0 hr. (E) ATM promotes cell migration in the absence of induced double-strand breaks (DSBs). Wound-healing assays were performed in siRNA-treated MDA-MB-231 human breast cancercells with siNon-coding (siNC) or siATM siRNAs. (F) Verification of DNA damage induction and quantification of wound healing from (E). Top: Western blot analysis of samples from (E) with the ATM markers KAP1-pS824 and mut-p53-pS15. Unmodified proteins are loading controls and ATM controls siRNA depletion. Bottom: Quantification of wound healing experiments from (E). (G) ATM depletion impairs cell migration and invasion, but not proliferation. Left panel: siNC or siATM cells were analyzed with xCELLigence Real-time cell analyzer (RTCA) to measure proliferation, migration and invasion in parallel and real-time. Experiments performed as detailed in ‘Materials and methods’. Right panel: ATM depletion 96 hr post-transfection. (mean ± s.e.m., n = 3).