Nucleic Acids Research, 2011, Vol. 39, No. 8 3173 Figure 6. A substantial amount of damages are still being repaired 2 h after exposure to DMS. Level of SSBs in EM9-XH cells treated with (A) DMS (2 mM) or (B) mock treated and left to repair. PARP inhibitor was added (open symbols, time point indicated by arrow) to slow down the ligation step, 2 h after treatment was terminated. The means and standard errors of three experiments are shown. The ongoing SSB formation 2 h after terminated treatment, represents MPG initiated BER events. Level of SSBs in A549 cells with or without siRNA depleted MPG and treated with (A) DMS (0.5 mM) or (B) mock treated and left to repair. PARP inhibitor was added (open symbols, time point indicated by arrow) to slow down the ligation step, 2 h after treatment was terminated. The means and standard errors of three experiments are shown. PARP inhibitor) occur in the cells 2 h after DMS was removed from the medium (Figure 6), which suggests that there is still ongoing repair at this time. According to our model, the repair events we detect through PARP inhibition represents only a fraction of uncoupled SSB intermediates, indicating that an even higher amount of total BER events still occur in the cell at this time. We speculate that this ongoing repair could be the result of slow damage detection by the glycosylase, possibly because of inaccessibility to the location of the lesion or the complexity of the lesion. We also see a difference in cells with siRNA depleted MPG compared to wild-type cells, indicating that we are indeed studying actual BER events (Figure 6D). However, it should be noted that there is no certain way to ensure that the damages repaired after 2 h are in fact original damages that were produced at the time of treatment. There is a possibility that cellular DMS is not removed in the washing step and that this could sit in the cell and make damage 2.25 h after initiation of the treatment. Even in this scenario DMS hydrolyzes in