Figure 4.

ASCL1, ASCL2, and MYOD1 bind open and closed chromatin and satisfy multiple criteria for pioneering transcription factors. (A) Heatmap of ChIP-seq and ATAC-seq from ESCs. Each plot compares the total set of binding sites identified for the factor indicated and shows the ChIP-seq (left) and ATAC-seq (right) signal for each peak region in a 6-kb interval centered on the peak apex. Rows are ordered based on the ATAC-seq signal. Dashed red line approximates the upper and lower quartiles used to define “open” versus “closed” binding sites. (B) Histogram of mean bHLH binding and chromatin accessibility (left) and H3K27ac enrichment (right) at binding sites identified by ChIP-seq for each factor. For each, the bHLH ChIP-seq signal (blue, left axis) is enriched at the peak center. At these sites, there is an increase in ATAC-seq and H3K27ac signal post bHLH induction (green versus red, right axis). (C) Heatmap of ASCL1 ChIP-seq and the ATAC-seq signal for regions showing a significant increase (left) or decrease (right) in chromatin accessibility upon ASCL1 induction. Each row represents a single peak interval with ASCL1 ChIP-seq (left), ATAC-seq from uninduced control ESCs (The ENCODE Project Consortium et al. 2007), or ESCs post-induction (right). (D) Frequency and distribution of Ebox motif distribution at bHLH binding sites identified in nucleosome-occupied (left) and nucleosome-depleted (right) chromatin. Each plot shows the frequency and distribution of a single Ebox permutation across the intervals surrounding the peak apex for the transcription factor, as shown. Motif shown represents the most significantly enriched de novo motif as compared to sequence-normalized background regions.