Figure 4. Topo I cleavage complex formation and induction of DNA double-strand breaks.

(A) Cleavage complex formation in untreated OVCAR-3 cells (bars 1,5), or 2 days after treatment with 20 moi Adp14 (bars 2,6), 10 nM of the CK2 activator 1-ethyl, 4,5 dicarbamoyl imidazole (bars 3,7), or both Adp14 and the CK2 activator (bars 4,8). Samples 5–8 were also treated with 10 µM of the ROS inducer pyocyanin. Cells were pulsed with [³H]-thymidine to label DNA, cleavage complexes were captured by K⁺SDS precipitation, and DNA was quantified by scintillation counting. (B) Western analysis of γ-H2A.X, an indicator of DNA double-strand break formation, in lysates of H358 cells subjected to the same treatments 1–8 as in part (A). Total H2A.X levels are shown as a control.