Figure 3

DNA binding and expression of the transcription factors AP-1, E2F1 and STAT-1 in PARP-1^+/+ and PARP-1^-/- cells. (A) EMSA analysis. Crude nuclear proteins (5 μg) from both PARP-1^+/+ and PARP-1^-/- cells were incubated with a 5' end-labeled probe bearing the high affinity binding site for AP-1, E2F1 and STAT-1. Formation of DNA/protein complexes was then monitored by EMSA on an 8% gel as detailed in Figure 2. The position of the AP-1, E2F1 and STAT-1 DNA-protein complexes is shown, as well as that of the free probe (U). P: labeled probe alone. (B) Coomassie blue staining of the protein samples used for conducting both the EMSA (panel A) and the Western blot experiment (panel C). One protein band present in both extract was randomly selected and its intensity determined by densitometric analysis in order to precisely calibrate the protein concentration used for the assays. (C) Nuclear extracts (10 μg) from both PARP-1^+/+ and PARP-1^-/- cells were examined in Western blot as in Figure 1 using antibodies directed against E2F1, STAT-1 and the AP-1 subunit c-jun. The position of the nearest molecular mass markers is indicated (60 kDa and 85 kDa).