Fig. 5
From: Immunoproteasome function maintains oncogenic gene expression in KMT2A-complex driven leukemia
Enrichment of BASP1 by PSMB8 inhibition inhibits KMT2A target gene expression. A Heatmap of differentially expressed genes in MOLM-13 cells: 100 nM PR-957 vs. DMSO (72 h). Upregulated (red; fold-change (FC) > 2, p < 0.05) and downregulated (blue; FC < -2, p < 0.05) genes. B Gene Set Enrichment Analysis (GSEA) of PR-957 treated MOLM-13 cells compared to EPZ5676- or VTP-50469-treatment; NES (normalized enrichment score). C Hockey-stick-plot of ranked genes found to be differentially enriched/depleted in genome wide CRISPR/Cas9 screening in MOLM-13 cells. D Heatmap depicting unsupervised hierarchical clustering of differentially abundant proteins detected by mass-spectrometry. 100 nM PR-957 vs. DMSO, 72 h, MOLM-13. Upregulated (red; FC > 2, p < 0.05) and downregulated (blue; FC < -2, p < 0.05) proteins. E Correlation between the magnitude of differential protein abundance (-log10 Padj) as determined by proteome analysis and the functional dependencies obtained by CRISPR/Cas9 screening (Δ beta-scores PR-957 vs. DMSO) in MOLM-13 cells. F Western Blotting showing expression of BASP1 in nuclear (N) and cytoplasmic (C) fractions of MOLM-13, KOPN-8, ML-2 and MV4;11 cells. PR-957 (100 nM, 200 nM) vs. DMSO, 72 h. G Heatmaps displaying IgG and BASP1 Cut&Run signal mapping to a 2-kb window around the TSS (Transcription Start Site). 100 nM PR-957 vs. DMSO, 48 h, MOLM-13 cells; 1 (out of n = 3) representative replicate. H Stacked bar plot depicting genomic distribution of BASP1 Cut&Run peaks. MOLM-13 cells; n = 3 independent replicates. I Dot-plot of ranked BASP1-bound TSS according to their abundance; 1 representative replicate. J Growth curves of KMT2A-r cells (MOLM-13, MV-4;11, ML-2, KOPN-8) transduced with pLEX-BASP1-HA-Tag (BASP1-HA) or pLEX-HA-Tag (EV-HA). n = 4 independent experiments, in triplicate; mean with SEM; 2-way ANOVA. K Survival curves of NXG recipient mice transplanted with 1 × 105 MOLM-13 or MV-4;11 cells expressing pLEX-BASP1 (BASP1) or pLEX-EV (EV) (n = 10 per construct and cell line). Two independent cohorts; Mantel-Cox test