Dexamethasone induces autophagy, and lack of its component decreases B-ALL cell sensitivity to dexamethasone. (A) WT and NPRL2-KO NALM-6 cells were treated for 24 hours with either DMSO or 100 nM dexamethasone. Changes in LC3B-II and p62 protein levels (both indicators of autophagic activity) were evaluated by western blot using LC3B and p62 antibodies. (B) WT and NPRL2-KO NALM-6 cells were treated as in panel A, and autophagic activity was measured by FACS using LysoTracker Red DND-99 dye. (C) Competition assays were performed using NALM-6 or MUTZ-5-Cas9 cells expressing either empty vector or a lentivirus vector encoding indicated sgRNAs targeting either ATG14 or ATG10, in the presence or absence of dexamethasone (10 nM for NALM-6, and 1 nM for MUTZ-5), as described in Figure 2B. Each condition was analyzed in triplicate, and data are represented as means ± SD. (D) WT and NPRL2-KO NALM-6–Cas9 cells were transduced with a lentivirus vector encoding an sgRNA targeting ATG14, and competition assays were performed with or without dexamethasone (10 nM), as described in Figure 2B. Each condition was assessed in triplicate, and data are represented as means ± SD. The data presented are representative of multiple independent experiments.