Figure 1. Wnt Pathway Activation Sensitizes Leukemia Cells to Asparaginase.
(A) CCRF-CEM cells were transduced with the GeCKO genome-wide guide RNA library in biologic duplicates, split into treatment with vehicle or asparaginase (10 U/L), and guide RNA representation was assessed after 5 days of treatment.
(B) Significance of gene depletion in asparaginase-treated conditions, as assessed by robust ranking aggregation (RRA) score calculated using MAGeCK analysis. Note that microRNA genes are not shown.
(C) CCRF-CEM cells were transduced with the indicated shRNAs, and knockdown efficiency was assessed by RT-PCR analysis. CT values greater than 36 were defined as not detected (N.D.).
(D) CCRF-CEM cells were transduced with the indicated shRNAs and subjected to Western blot analysis for active (nonphosphorylated) β-catenin or GAPDH.
(E) CCRF-CEM cells transduced with a lentiviral TOPFlash-EGFP (7xTcf-EGFP) reporter of canonical Wnt/β-catenin driven transcription were transduced with the indicated shRNAs, and reporter-driven EGFP fluorescence was assessed.
(F) The indicated cell lines were transduced with the indicated shRNAs and treated with the indicated doses of asparaginase. Relative viability was assessed after 8 days of treatment by counting viable cells. All cell counts were normalized to those in shLuc-transduced, no-asparaginase controls.
(G) CCRF-CEM cells were transduced with the indicated shRNAs, treated with asparaginase (10 U/L) for 48 hr and caspase 3/7 activity assay was assessed.
(H) CCRF-CEM cells were treated as indicated and the number of viable cells after 8 days of treatment was assessed. All cell counts were normalized to those in no-Wnt3a, no-asparaginase controls.
All error bars represent SEM.
See also Figures S1 and S2 and Tables S1 and S2.