DSBs lead to DMSR. (A) The experiment procedure for DMSR analysis in G1 cells is shown. The phase of U2OS cell cycle in indicated time points was determined by flow cytometry. (B) Grayscale inverted images of Fig. 1 A show the increased microtubule number originated from the centrosomes in IR-treated cells. (C) RPE-1 cells were synchronized at G1/S phase, treated with 5 Gy IR, released, and fixed at indicated time points. DMSR was determined by microtubule regrowth assay. Upper: Microtubules (green) were stained with anti–β-tubulin antibody. Scale bar, 20 µm. Lower: Grayscale inverted images show the increased microtubule number originated from the centrosome in IR-treated cells compared with untreated cells. (D) DMSR is confirmed by live-imaging time-lapse experiments. Overlaid GFP-EB3 signal indicated the newly synthesized microtubule tracks within indicated time periods (overlay). The single frames of GFP-EB3 comet associated with the growing microtubule plus ends in steady status (single frame) are also displayed. Images of projections at indicated time points and spanning 0–1 s, 0–15 s, and 0–30 s are shown. See also Videos 1, 2, and 3. Scale bar, 20 µm. UT, untreated; sec, seconds; PI, propidium iodide.