Members of the SAGA complex regulate susceptibility of AML cells to DNT-mediated cytotoxicity. (A) Volcano plot of all genes enriched in dead (Annexin V⁺) AML3 cells after DNT treatment. X-axis represents the log2-transformed fold changes in normalized sgRNA tag counts in dead vs live cells. Red circles denote genes that are significantly enriched (FDR ≤0.10) in dead cells; genes associated with the SAGA complex are labeled. (B) The top 6 GO gene sets represented by the 17 enriched genes in AML3 dead cells (left panel). Enriched genes in these gene sets are shown (right panel). (C) Western blot analysis of AML2 Cas9-expressing cells transduced with pairs of sgRNAs against ATXN7L3, ENY2, USP22, or AAVS (a targeting genome-wide control). (D-E) Percentage of specific killing of SAGA-deficient AML3 (D) and AML2 (E) Cas9-expressing cells following coculture with DNTs at various E:T ratios for 2 hours. These experiments were conducted independently 3 times, with similar results. (F) Immunoblot analysis of H2B ubiquitination (H2Bubq) levels in ATXN7L3, ENY2, and USP22 in AML2 and AML3-Cas9 cells. Data are the mean ± standard deviation of replicates. *P < .05, **P < .01, ***P < .001, ****P < .0001. n.s., nonsignificant.