Fig. 7: Atoh8 restrains cellular plasticity by binding specific enhancers to limit WNT signalling activity.
a, Heatmap displaying Atoh8 read counts ± 1 kilobase around merged peak summits. b, Genomic distribution of Atoh8-specific peaks. c, The most enriched DNA-binding motifs associated with Atoh8, derived from a de novo motif analysis (MEME). d, Mean read count enrichment density associated with H3K27Ac and H3K4Me1. e, Venn diagram showing the numbers of Bcl11b versus Atoh8 peaks. f, Fraction of Atoh8 sites retrieved within open chromatin regions (ATAC-Seq) during reprogramming and transformation. g, Examples of open chromatin sites. Chr, chromosome. h, Atoh8-centred chromatin state analysis. ES, Embryonic stem cells. i, Volcano plot showing differentially expressed genes in Atoh8 KD versus control MEFs. Samples were collected 5 d after RNA interference induction. Each dot corresponds to a transcript. Benjamini–Hochberg-adjusted P values of the comparisons were computed using the limma-voom workflow modified two-sided t-test. j, Top, schematic depicting the c-Myc binding site (BS) on the Atoh8 promoter. Bottom, real-time qPCR showing the levels of DNA immunoprecipitated. The data are represented as a percentage of Atoh8 DNA levels in ChIP input and are from two independent experiments. k, Western blot for c-Myc, Atoh8 and Gapdh in MEFs in response to c-Myc exogenous expression. l, Graph depicting the fold-enrichment as determined by statistical over-representation analysis. RTK, Receptor tyrosine kinase. m, Top, pictures representing AP+ colonies. Bottom, pictures representing cresyl violet foci. n, Quantification (n = 3 independent experiments). o, Quantification of AP+ colonies and foci (n = 3 independent experiments). p, Top, pictures representing AP+ colonies. Bottom, pictures representing cresyl violet foci. q, Quantification (n = 6 independent experiments for the left panel and n = 3 for the right panel). r, Western blot of Atoh8 expression in MEFs treated with recombinant Wnt3a for 48 h. s, Schematic recapitulating the main findings of the study. In n, o and q, the data represent means ± s.d. and statistical significance was determined by two-tailed Student’s t-test.
