Validation and characterization of candidate MEIS1 regulators. (A) Schematic of competition assay with cells transduced with sgRNAs expressed in a BFP-positive backbone. The proportion of BFP-positive, sgRNA-transduced cells (% BFP) is assessed over time using flow cytometry and compared to BFP negative, sgRNA nontransduced cells. (B) Bar graphs indicating flow cytometry measurements of the percentage of UB3 BFP-positive cells over time (y-axis), normalized to the baseline (time 0 or T “0”) measurement (n = 3); the x-axis indicates the sgRNA number for NTCs (gray bars) or each gene (n = 2-3 sgRNAs per gene). Each bar represents a time point measurement: baseline (T0), day 8, day 11, and day 14. (C) Uniform manifold approximation (UMAP) plots of synthetic gene expression profiles from CROP-seq perturbations in U937 GFP-MEIS1 cells. (D) Dot plot comparing the expression of self-renewal–associated and differentiation-associated genes in cells with each perturbation. Dot sizes scale with the percentage of cells per gene knockout. (E) A relative score of DepMap dependencies for leukemia (n = 41) compared with nonleukemia cell lines (labeled “other”; n = 979) is shown for candidate hits identified in our screen. The y-axis is the rank of AML selectivity (with a rank of 1 being the most selective), and the x-axis shows the negative log10 P values. The dotted line separates the significant (false discovery rate [FDR] P value <.05) from the nonsignificant (FDR P value >.05). The P value was calculated using the Wilcox test of Chronos scores in leukemia compared with other cell lines. (F) A sigmoid plot of DepMap data showing the dependency score (Chronos, x-axis), compared with the normalized dependency rank (y-axis) for SGF29 in 41 leukemia cell lines in blue compared to 979 nonleukemia cell lines labeled “other” (gray).