Fig. 3: Cas13d enables the highly reproducible identification of GIs in oncogenic signaling pathways.

A Genetic interaction maps showing GI scores from single gRNA (left panel) and single-gene array screens (right panel), without and with imatinib treatment. Tau values from single gene perturbations in both possible orientations (Gene1-gNTC and gNTC-Gene1) are displayed along the edges of each GI map. GI scores derived from gRNA/array orientation 1 are shown in the bottom left of each GI map, while GI scores derived from gRNA/array orientation 2 are shown in the top right. Positive GI scores indicate buffering interactions while negative GI scores indicate synergistic interactions. B Volcano plots showing GI scores and associated significance (−log₁₀ (p-value)) for all possible gene-gene combinations from single gRNA (top) and single-gene array screens (bottom), with and without imatinib treatment. Blue and yellow data points indicate interactions that meet the threshold of GI  > < ±0.2 and FDR  <  0.5. Data points with black labels indicate GIs that were identified in both orientations (gRNA-gNTC and gNTC-gRNA). Data points labeled in gray indicate “same gene GIs” that passed the threshold, where gRNAs in both positions targeted the same gene. Red data points indicate ‘single gene controls’ where GI scores were calculated between genes, with one gene replaced by non-target control gRNAs (see methods for details). Significance was calculated using limma moderated t test followed by Benjamini–Hochberg multiple testing correction. C Pearson correlation (r) between GI scores determined from imatinib treated and untreated cells via the single gRNA and single-gene array approach, in both orientations. D Genetic interaction network of reproducible GIs in both gene orientations (Gene1-Gene2 and Gene2-Gene1) that were identified between all six investigated genes derived from the single-gene array strategy. Only interactions that were identified by GIs of ><±0.2 and FDR < 0.5 in both orientations (gRNA-gNTC and gNTC-gRNA) are shown. Ras-GEF = Ras guanine nucleotide exchange factor. Ras-GAP = Ras-GTPase activating protein. Edges between genes are colored based on the average GI score of screen replicates and gRNA-gNTC orientations. Created in BioRender. Böttcher, M. (2025) https://BioRender.com/e57j701. Source data are provided as a Source Data file.