Fig. 1: DDR-focused RNAi screen identifies HORMAD1-driven genetic dependencies.

A Schematic diagram describing workflow for parallel siRNA screens in parental SUM159, and clonally-derived HORMAD1-inducible SUM159. Cells were reverse-transfected into siRNA-containing 384-well plates, and doxycycline added 24 h post-transfection. Cell viability was measured 5 days post-transfection using CellTiter-Glo. CellTiter-Glo readings were converted into Z scores, and doxycycline-inducible effects were identified using drug effect (DE) Z-scores. Candidate genetic dependencies were selected using the following criteria: 1) DE Z-score < -3 in HORMAD1-inducible clone 1, 2) DE Z-score > -2 in SUM159 parental clone and 3) Z-score > -3 in DMSO-treated arms. B Scatter plot displaying the distribution of DE-Z scores in HORMAD1-inducible SUM159 clone 1. Negative DE Z-scores are indicative of HORMAD1-driven dependencies. A numerical threshold of DE Z-score < -3 was used for candidate selection. Fourteen candidate DDR genetic dependencies were interrogated in secondary deconvolution experiments, of which 5 were validated as HORMAD1-induced genetic dependencies (marked in red). C–G Bar plots displaying increased normalised percentage inhibition (NPI) of clonally-derived HORMAD1-inducible SUM159 cells (+DOX/ + HORMAD1 vs. -DOX/-HORMAD1) transfected with an siRNA pool or four individual siRNAs targeting ATR, BRIP1, POLH, TDP1 and XRCC1 and exposed to HORMAD1 expression for 4 days. Non-targeting (siALLSTAR) and targeting (siPLK1) siRNAs were used as normalisation controls. Error bars indicate SD from mean effects (n = 3), p values represent multiple Student t-tests (***p = < 0.0001, **p = < 0.001, *p = < 0.05).