Fig. 1.

Differential H3K27ac analysis across tissues is an effective method to identify tissue-specific enhancers. a Scatterplots of H3K27ac ChIP-seq enrichment at each known DV enhancer (1 kb centered on midpoint) against the transcript levels of the known target gene show a good but modest correlation in dorsal ectoderm precursor embryos gd ⁷ (left) and in mesoderm precursor embryos Tl ^10b (right). ChIP-seq enrichment values represent fold change over the corresponding input control after normalizing for differences in read count and fragment size. MEs mesoderm enhancers, DEEs dorsal ectoderm enhancers. b The correlation between H3K27ac and transcript levels becomes stronger when the fold changes in H3K27ac levels between the two mutant embryos at each enhancer are plotted against the fold changes in transcript levels of the corresponding target genes. See Additional file 3: Figure S1 for gene names. c De novo identification of DV enhancers based on CBP candidate regions and differential H3K27ac analysis by the package DESeq2. Numerous candidate enhancers were located with CBP ChIP-seq data in wild-type embryos. DESeq2 was then used to identify significant differences in H3K27ac between gd ⁷ and Tl ^10b embryos within 1-kb windows centered on the CBP peaks. The average DESeq2-normalized ChIP signal for all replicates is shown as a scatterplot on the right. CBP regions significantly different for H3K27ac are shown in blue (MEs) and yellow (DEEs), while non-differential regions are shown in gray. d Average enrichment of H3K27ac in gd ⁷ and Tl ^10b embryos, as well as CBP in wild-type embryos, is shown for MEs and DEEs that were located distally, at least 1 kb from any transcription start site (TSS). The gray bar represents the 1-kb window used to calculate H3K27ac enrichments, and the red box represents the 201-bp enhancer region. e Examples of newly identified distal enhancers zfh1-ME2 and C15-DEE2. H3K27ac is shown as normalized ChIP enrichments over input, while CBP is shown as normalized ChIP reads (reads per million). The gray bar represents the 1-kb H3K27ac window, and the blue and yellow boxes represent the 201-bp ME or DEE enhancer region, respectively