Figure 5: Deletion of the 20q-TERRA locus decreases telomere protection in U2OS cells.

(a) Quantification of the total γH2AX signal per nucleus (mean values±s.e.m., n=number of cells) is shown. The total number of cells analyzed is indicated. (b) Quantification of the total 53BP1 spot signal per nucleus (mean values±s.e.m., n=number of cells is shown). The total number of cells analyzed is indicated. (c) Graphs showing the quantification of the co-localization (TIF) between TRF2 and either γH2AX or 53BP1 in WT cells and in all 20q-KO clones (mean values±s.e.m., n=3 independent experiments for γH2AX and n=number of cells for 53BP1) per cell is shown. The total number of nuclei analyzed is indicated. (d) Representative images of the average number of TIFs found on double inmunostain to detect the telomere protein TRF2 (green) and either the DNA damage markers phospho-Histone γH2AX or 53BP1 (red) in the U2OS cells WT or deleted for the 20q locus. Arrowheads indicate co-localization events. Scale bar, 10 μm. (e) Quantification of chromosomal end-to-end fusions in WT and in the 20q-KO cells from three independent experiments (mean values±s.e.m., n=metaphases). Examples of end-to-end fusions are shown as well. Scale bar, 1 μm. (f) Array-CGH analysis was performed on hybridization on the same membrane of DNA differentially labelled from WT and 20q-KO cells. The chromosomal gains and losses in 20q-KO cells normalized by WT cells are represented. The chromosomal gains are shown in green and in red the chromosomal losses. One-way Anova with Dunnett’s post test was used for all statistical analysis except for the quantification of chromosomal fusions in which the Student’s t-test was used (*P<0.05, **P<0.01 and ***P<0.001).