Extended Data Fig. 3: Combo treatment triggers transcriptional changes associated with myeloid differentiation and interferon response in AML cells.
From: Perturbing LSD1 and WNT rewires transcription to synergistically induce AML differentiation
a, GSEA of 47-gene leukemia stem cell (LSC) signature in ER-HoxA9 cells treated with drug combo vs. vehicle. b, PCA of RNA-seq of THP-1 cells treated with vehicle, GSK-LSD1, LY2090314 or combination of both inhibitors. c, GSEA of myeloid maturation signatures in drug-treated THP-1 cells. d, GSEA of additional myeloid maturation signatures in THP-1 cells treated with combo vs. vehicle. e, GSEA of Somervaille leukemia stem cell (LSC) signatures in THP-1 cells treated with combo vs. vehicle. f, Western immunoblots for β-catenin, phospho-GSK3α/β (Y279/Y216), IRF7 and vinculin after treatment with the indicated inhibitors for 3 days in THP-1 cells. Vinculin was used as a loading control. The experiment was repeated three times with similar results. g, GSEA of Wnt and β-catenin signaling genes in ER-HoxA9 cells treated with drug combo vs. vehicle. h, GSEA of Wnt and β-catenin signaling genes in THP-1 cells treated with drug combo vs. vehicle. i, Expression of TCF7 (TCF1) in drug-treated ER-HoxA9 cells. Data are presented as mean values ± SD from three independent biological replicates. P-values indicate the significance of unpaired, two-tailed Student t-tests. “ns” indicates not significant. j, Heatmap of expression of genes synergistically upregulated or downregulated upon combo treatment (left), and type I interferon signature genes (right). k, Expression of genes relating to the type I interferon response in drug-treated THP-1 cells. Data are presented as mean values ± SD from three independent biological replicates. P-values indicate the significance of unpaired, two-tailed Student t-tests. See source data for individual P-values. Asterisks indicate significance at these levels: *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, “ns” indicates not significant. l, EnrichR database pathways enriched in upregulated combo synergy signature genes. Significance of enrichment z-scores is shown as q-values (corresponding to p-values adjusted for significance by Benjamini-Hochberg method). m, Dot blot for dsRNA using total RNA from THP-1 cells treated with indicated inhibitors for 3 days. Total RNA extract treated with mock, RNase T1, RNase III, or RNase A (350 mM NaCl) was dotted on Hybond N+ membranes, visualized by methylene blue staining and immunoblotted with J2 antibody. n, Table shows rank and P-value of enrichment of top transcription factor (TF) motifs in accessible chromatin of ER-HoxA9 cells treated with vehicle, GSK-LSD1, LY2090314 or combination of both inhibitors. Motifs shown in all treatments are the top 10 motifs from combo-treated cells. Enrichment rank and P-values of TCF7 (TCF1) and LEF motifs are also shown. Motif enrichment significance was determined via hypergeometric tests.
