Figure 6
From: Mir142 loss unlocks IDH2R140-dependent leukemogenesis through antagonistic regulation of HOX genes
Mir142 loss of function upregulates a leukemic HOX/Meis1/Pbx3 signature and antagonizes IDH2R140Q-dependent silencing of Hoxa cluster genes. (a) Volcano plot showing differentially expressed genes (DEGs) in Mir142–/– versus WT GMPs (padj < 0.05 colored dots, WT n = 4, Mir142–/– n = 3). miR-142-3p targets predicted by Targetscan highlighted in blue. Light blue: miR–142–3p.1 specific targets. Dark blue: miR-142-3p.2 specific targets. Turquoise: targets of both miR-142-3p.1 and miR-142-3p.2 isomirs. Top ten most significantly differentially expressed genes are labelled. (b) Gene set enrichment analysis (GSEA) reveals strong enrichment of predicted miR-142-3p targets (union of miR-142-3p.1 and miR-142-3p.2 targets) in genes upregulated in Mir142–/– GMPs compared to WT. (c) Venn diagram showing overlap of miR-142-3p.1 and miR-142-3p.2 targets with DEGs from Mir142–/– versus WT GMPs and Mir142–/– + IDH2R140Q versus WT + IDH2R140Q GMPs (all DEGs with padj < 0.05 were used for the analysis). Highlighted targets Ash1l and Ctnnb1 are differentially expressed in both Mir142–/– and Mir142–/– + IDH2R140Q GMPs. (d) GSEA reveals strong enrichment of a mutant NPM1–associated leukemic signature in genes upregulated in Mir142–/– + IDH2R140Q GMPs compared to WT + IDH2R140Q. (e) Heatmap showing expression of the significantly differential leading-edge genes contributing to enrichment of the mutant NPM1–associated leukemic signature in Mir142–/– + IDH2R140Q verus WT + IDH2R140Q GMPs (padj < 0.05). Signature includes multiple homeobox family genes (Hoxa5/7/9/10, Meis1 and Pbx3) highlighted with arrows. Expression in Mir142–/–GMPs is shown for comparison. Columns represent normalized expression values from each sample isolated from individual mice (WT + IDH2R140Q (n = 4), Mir142–/– + IDH2R140Q (n = 6), and Mir142–/– (n = 3)). (f) Heatmap of most significantly differentially expressed genes in CD34+Mac1+ myeloblasts and Mac1+(CD34–) myeloid cells isolated from Mir142–/– mice and Mir142–/– + IDH2R140Q recipients (GFP + cells). (g) Expression of differentially expressed Hoxa cluster genes and Meis1 in CD34+Mac1+ myeloblasts and Mac1+(CD34–) myeloid cells in bone marrow of Mir142–/– mice (padj < 0.05, n = 2). (h) Expression of differentially expressed Hoxa cluster genes and Meis1 in Mir142–/– + IDH2R140Q GMPs, CD34+Mac1+ myeloblasts and mature Mac1+(CD34–) cells (padj < 0.05, n = 2). (i) Expression of homeobox genes including Hoxa cluster genes (Hoxa5/7/9/10), Meis1 and Pbx3, and the HOX regulator Ash1l in WT + CTL (n = 3), WT + IDH2R140Q (n = 4), Mir142–/– + IDH2R140Q (n = 6) and Mir142–/– (n =3) GMPs. (j) CFC assay with WT and Mir142–/– HSPCs co-transduced with IDH2R140Q and either an shRNA targeting Ash1l (shAsh1l) or non-targeting control (shNeg) (n = 2). Statistical significance assessed by unpaired two-tailed t test.
