Analysis of TLX3-dependent transcriptional programs. (A) Bar plot representation of BFP+ cells relative to starting cell culture (day 0) in DND-41 cells expressing shRNA against TLX3 (shTLX3) or Renilla luciferase (shRen). Results are mean ± SD and were analyzed by two-way analysis of variance (ANOVA). (B) Genome-wide distribution of TLX3 ChIP-seq peaks in DND-41 cells. (C) Top enriched motifs in TLX3 ChIP-seq binding sites by HOMER. (D) Heatmaps of TLX3 (blue) and MYB (green) ChIP-seq signals centered on TLX3-binding sites in DND-41 cells. Each row represents a single TLX3-binding region. (E) Venn diagram of the overlap between TLX3- and MYB-binding sites in DND-41 cells. P value was calculated by the Fisher exact test. (F) Top enriched gene ontology (GO) biological process, human phenotype, and mouse phenotype terms associated with the nearest-neighbor genes within 100 kb of TLX3 and MYB co-occupied genomic regions. The –log10(P value) was calculated using binomial test with the GREAT software. (G) Genome browser tracks for ATAC-seq and H3K27ac, TLX3, and MYB ChIP-seq in DND-41 cells for CDK6 and BCL2L1. (H) Gene set enrichment analysis of gene sets enriched in TLX3+ leukemia samples from 3 independent studies17,43,44 comparing differentially expressed genes upon TLX3-depletion in DND-41 cells (N = 2 replicate RNA-seq) or TLX3 overexpression (TLX3-OE) in Jurkat cells (N = 3 replicate RNA-seq).
