Fig. 1: PARP1/PARP2 inhibitor sensitivity is determined by serine ADPr levels, which are controlled by HPF1 and ARH3 activity.

a Reduced survival of HPF1 KO cells after treatment with the indicated PARP1/PARP2 inhibitors. Representative images (top) and quantification of colony formation assay (bottom). b HPF1 loss results in further sensitisation of BRCA1- or BRCA2-deficient cells to Olaparib. See Supplementary Fig. 2a for BRCA1, BRCA2, HPF1 and tubulin control immunoblots. c Effects of 6-day Olaparib treatment on γH2AX formation and H3S10P reduction depend on cellular HPF1 and ARH3 protein levels. The experiment was repeated independently 3 times with similar results. See Supplementary Fig. 2b for Pan ADPr immunoblot. A repeat of the experiment with an additional concentration of Olaparib is provided in Supplementary Fig. 2c. d Flow cytometry analysis of cell cycle profiles following 4-day exposure to Olaparib. Asterisks in different colours indicate significant difference in corresponding cell populations between WT and HPF1 KO cells. e HPF1 and ARH3 status determines the effects of 4-day Olaparib treatment on γH2AX levels (top) and percentage of cells with >4 N DNA (bottom) as determined by flow cytometry. f ARH3 overexpression (OE) renders cells more sensitive to PARP1/PARP2 inhibition. g Loss of ARH3 confers resistance to Olaparib. h Schematic representation of ADPr synthesis and removal (top), and summary of the impact of HPF1 and ARH3 status on cell serine ADPr levels and PARP1/PARP2 inhibitor (PARPi) sensitivity (bottom). a, f, g Data are shown as mean ± SD of three independent experiments. b, d, e Data are shown as mean ± SEM of three (b) or five (d, e) independent experiments. *p < 0.05; **p < 0.01; ***p < 0.001 (two-tailed Student’s t-test). Flow cytometry gating strategy for the analyses shown in (d) and (e) is shown in Supplementary Fig. 2d.