Figure 6.

A-T post-mitotic neurons are defective in DDR. In a, D30 post-mitotic neurons (D30) were tested by western blot with antibodies specific for the various ATM phosphosubstrates and cleaved PARP at various times after treatment with 5 Gy IR. β-Actin was used as a loading control. DNA SSBs (b) and DSBs (c) were analyzed in neurons (D30) by alkaline and neutral comet assay, respectively, at two different times after treatment. 20 min after H₂O₂ or IR treatment was considered the time point with the maximum DNA damage. In b and c, representative photos of comets from untreated cells and at different time points following treatment. The ratio between treated and untreated tail moments of at least 50–70 cells per experimental point is shown in the graphs (values in %). One representative experiment out of three is shown. In d and e, formation and resolution of IR-induced nuclear foci. Neurons (D30) were irradiated with 0.5 Gy, collected at the indicated times and labeled for γ-H2AX (d) and 53BP1 (e). For each treatment, the number of foci was scored from 100 cell nuclei per duplicate preparations and from three independent experiments (mean±S.D.) (d and e, bottom) For each time point, the difference between Ctrl and A-T was statistically significant (^#P<0.01) (analysis performed by the Student's t-test)