Dynamic assembly of PARP1 at DSB sites. a, scheme of the laser microirradiation experiment. b, laser microirradiation induces rapid redistribution of PARP1-GFP at DSB sites. The top panels show the photobleaching of Hoechst 33258 dye following microirradiation. PARP1-deficient cells transiently expressing physiological levels of PARP1-GFP (bottom) were microirradiated with a 780-nm laser line (10% of maximum laser output) to induce DSBs. The numbers at the bottom right of each image indicate elapsed time (s). Bar, 5 μm. c, the graph shows a plot of intensity versus time illustrating the increase in the concentration of both PARP1-GFP and GFP-PARP1 immediately following DSBs induction. Data are presented as mean ± S.E. from at least 10 cells for each construct. d, differential interference contrast (DIC) images were collected to monitor nuclear membrane integrity and cellular viability 2 min after laser microirradiation. Immunofluorescence shows localization of both PARP1-GFP and PAR polymer after laser microirradiation. The arrow indicates the region where DSBs were introduced (1-μm-wide region).
