Figure 6

HERC2 dysfunction alters the sensitivity of cells to CX-5461. (a–c) HeLa-shHERC2 (a) and HCT116-shHERC2 (b) cells, with or without Dox-mediated induction, or WT or HERC2^ΔE3/ΔE3 HCT116 cells (c) were exposed to indicated doses of the CX5461 for 24 h and analyzed for clonogenic survival after two weeks. The data are shown with the nonlinear regression fit curves of one phase decay (GraphPad Prism). (d–h) Cells as in (a–c) were transfected with control (–) or BRCA1-specific siRNA, with or without Dox-mediated induction, and either subjected to immunoblotting with the indicated antibodies (d,e) or clonogenic survival assays (f–h, and supplementary Figure S11) as in (a–c). Average ± SD values, normalized to cells without agents were derived from three independent experiments. P-values of interactions were calculated using two-way ANOVA. The concentration that inhibited 50% of the colonies (IC50 value) was as follows: (a) Dox (−): 17.3 nmol/l, Dox ( +): 22.3 nmol/l, (b) Dox (−): 22.4 nmol/l, Dox ( +): 21.6 nmol/l, (c) WT/WT: 16.1 nmol/l, ΔE3/ΔE3: 7.3 nmol/l, (f) Dox (−): 10.0 nmol/l, Dox ( +): 18.3 nmol/l, (g) Dox (−): 6.1 nmol/l, Dox ( +): 18.1 nmol/l, (h) WT/WT: 17.8 nmol/l, ΔE3/ΔE3: 39.3 nmol/l. Full-length blots/gels are presented in Supplementary Figure S12.