Fig. 5: Aphidicolin induced replication stress increases TRF1/PARP1 interaction, TRF1 PARylation and TIFs induction upon PARP1 interference.

HeLa were treated with 0,2 μM aphidicolin for 20 h Then cells were PI stained and analyzed for cell cycle distribution by flowcytometry (a). Parallel samples were processed for IP with an anti-TRF1 followed by anti-PARP1 to measure TRF1/PARP1 binding, or with an anti-PAR followed by an anti TRF1 antibody to detect TRF1 PARylation (b). Hela cells interfered with siPARP1, or a SCR sequence were exposed to 0.2 μM aphidicolin for 20 h and then processed for IF-FISH to detect γH2AX and telomeric sequences. Representative images at 63X magnification were shown (c). The percentage of TIFs positive cells and the mean number of colocalization per nucleus among the γH2AX positive nuclei were scored and reported in histograms d and e respectively. The histograms report the mean of three independent experiments. Bars are SD. P value was determined by unpaired two tailed t-student test **P ≤ 0.01, ***P ≤ 0.001.