Fig. 4: HNF4A activity maintains colonic epithelial identity.

a, DiffTF analysis of combined ATAC–seq and RNA-seq data with selected TFs highlighted. Shaded areas indicate gain (red) or loss (blue) of TF activity in AKP Atrx^KO organoids compared with AKP controls. TFs highlighted in green are associated with transcriptional activation and in red with transcriptional repression. b, Volcano plot of H3K27ac CUT&RUN data in AKP control and AKP Atrx^KO organoids. Each data point represents a H3K27ac binding peak. Significantly altered sites (false discovery rate (FDR) ≤ 0.05) are highlighted in pink. c, Table outlining the overlap between ATAC–seq accessibility changes and altered H3K27ac peaks in AKP control and AKP Atrx^KO organoids. H3K27ac losses are mostly associated with reduced chromatin accessibility. d, Representative Integrative Genomics Viewer (IGV) browser tracks of AKP control and AKP Atrx^KO ATAC–seq accessibility and H3K27ac CUT&RUN data. Cdx1 and Hnf4a gene loci are shown. Regions shaded grey have significant loss of chromatin accessibility and corresponding depletion of H3K27ac. e, Table outlining the overlap between RNA-seq gene expression changes in AKP Atrx^KO and AKP Hnf4a^KO organoids. f, Fluorescence microscopy of phalloidin-stained AKP control or AKP Hnf4a^KO organoids after treatment with TGFβ (5 ng ml^–1). Scale bars, 400 μm. g, Quantification of the percentage of AKP control and AKP Hnf4a^KO organoids adopting a spindle-like morphology after TGFβ treatment (n = 3 independent experiments each). h, Representative IHC images of β-catenin-stained subcutaneous tumours from mice injected with AKP control or AKP Hnf4a^KO organoid cells (n = 5 mice each). β-catenin staining is used to identify tumour cells. Scale bars, 100 µm (overview) and 50 μm (zoom). For a, two-sided P values for each transcription factor was calculated with Welch two-sample t-tests using the bootstrap approach. Adjusted P values were calculated using the Benjamini–Hochberg method for multiple-testing correction. For g, data are mean ± s.d., and P values were calculated using two-tailed Student’s t-tests.