Figure 5: Changes in gene expression and DNA modification (5mC+5hmC) in Tet2/3 DKO cells.
From: Acute loss of TET function results in aggressive myeloid cancer in mice
(a) Differential gene expression in WT versus Tet2/3 DKO LSK cells. Red dots—differentially expressed genes (P value ≤0.05, fold change >1.5 or <0.67; thresholds indicated by blue lines). Grey dots—all other genes. (b) Gene set enrichment analysis (GSEA) of RNA-Seq data. Myeloid gene signatures are significantly enriched for genes upregulated in Tet2/3 DKO versus WT LSK cells. (c) Genome browser visualization showing increased and decreased expression of myeloid (Mpo, Csf1r) and lymphoid (Dntt, Cd72) genes in Tet2/3 DKO versus control LSK cells. (d) Slight but consistent increase in average DNA modification at gene regions in Tet2/3 DKO (red lines) versus WT LSK cells (blue lines), regardless of whether gene expression is upregulated (top) or downregulated (bottom). Similar results were obtained for all genes (Supplementary Fig. 17c). Shaded areas=±1 s.d. TSS, transcription start site; TTS, transcription termination site. (e) Changes in gene expression versus changes in average DNA methylation for all genes differentially expressed in Tet2/3 DKO LSK versus WT LSK cells. Top, gene bodies; bottom, promoter regions (TSS±2 kb). Each gene body/TSS region is represented by a dot. Most regions show increased methylation, regardless of whether they are up- or downregulated in DKO LSK cells relative to WT. (f) Active enhancer regions in LT-HSC43 show increased DNA modification (5mC+5hmC) in Tet2/3 DKO versus WT LSK cells. Each bin contains all the CpGs falling within the ∼7,000 active enhancer regions. Colour scale, density of CpGs with the indicated methylation levels. Enhancer centres in WT LSK cells show an increased fraction of unmethylated CpGs relative to Tet2/3 DKO LSK cells (green colour at the bottom), whereas enhancer edges in Tet2/3 DKO LSK cells show an increased fraction of methylated CpGs (5mC+5hmC) compared with WT (red colour at the top). For details, see Supplementary Methods. (g) Graphical representation of increased DNA modification in Tet2/3 DKO versus WT LSK cells. Data are from central regions of the enhancers (blue and red rectangles in f).
