Combined inactivation of DEK and PARP1/2 counteracts DNA damage induced by low doses of CPT.

U2-OS control and shDEK cells were pulse-labelled with EdU, then either left untreated or treated with replication stress inducers (25 nM CPT or 10 μm HU) for one hour, in the presence or absence of 1 μM ABT-888. 53BP1 (red) and γH2AX (green) foci formation was visualized via indirect immunofluorescence analysis, EdU (magenta) using click chemistry. DNA was counterstained with Hoechst 33342 (cyan). (A) Representative confocal images. Scale bar: 5 μm. (B) Quantification of 53BP1/γH2AX colocalization in S-phase cells. Foci were counted and colocalization determined using the foci counter of the BIC macro tool box. At least 118 cells per experimental condition were evaluated. The experiment was performed in triplicates. The bands inside the boxes display the median, whiskers indicate the 5th to 95th percentile and outliers are omitted for clarity. t-test: *** p≤0.001.