Super-enhancers are transcriptionally more active and cell type-specific than stretch enhancers

Abstract

Super-enhancers and stretch enhancers represent classes of transcriptional enhancers that have been shown to control the expression of cell identity genes and carry disease- and trait-associated variants. Specifically, super-enhancers are clusters of enhancers defined based on the binding occupancy of master transcription factors, chromatin regulators, or chromatin marks, while stretch enhancers are large chromatin-defined regulatory regions of at least 3,000 base pairs. Several studies have characterized these regulatory regions in numerous cell types and tissues to decipher their functional importance. However, the differences and similarities between these regulatory regions have not been fully assessed. We integrated genomic, epigenomic, and transcriptomic data from ten human cell types to perform a comparative analysis of super and stretch enhancers with respect to their chromatin profiles, cell type-specificity, and ability to control gene expression. We found that stretch enhancers are more abundant, more distal to transcription start sites, cover twice as much the genome, and are significantly less conserved than super-enhancers. In contrast, super-enhancers are significantly more enriched for active chromatin marks and cohesin complex, and more transcriptionally active than stretch enhancers. Importantly, a vast majority of super-enhancers (85%) overlap with only a small subset of stretch enhancers (13%), which are enriched for cell type-specific biological functions, and control cell identity genes. These results suggest that super-enhancers are transcriptionally more active and cell type-specific than stretch enhancers, and importantly, most of the stretch enhancers that are distinct from super-enhancers do not show an association with cell identity genes, are less active, and more likely to be poised enhancers.

Keywords: Gene regulation; epigenomics; genomics; stretch enhancers; super-enhancers.

Figure 1.

Genomic distribution and conservation of super-enhancers and stretch enhancers in 10 human cell types. (a) Number of super- and stretch enhancers in 10 cell types. (b) Fraction of the human genome covered by super- and stretch enhancers across 10 human cell types. (c) Distribution of distances to TSS for super- and stretch enhancers (average across the 10 cell types) (P value <2.2e-16, Wilcoxon rank sum test). (d) Evolutionary conservation score, phastCons scores obtained from UCSC 100 vertebrate species (phastCons100way) at super- and stretch enhancers with 6 kb flanking regions in H1-ES, K562, NHLF, and Islets cell types.

Figure 2.

Chromatin modifications at super-enhancers and stretch enhancers. (a) Genome-wide average ChIP-seq profiles for H3K27ac, H3K4me, H3K4me3, and H3K27me3 at super- and stretch enhancers in H1-ESC, GM12878, and K562. (b) Genomic browser screenshot showing super- and stretch enhancers with ChIP-seq signals for H3K27ac, H3K4me1, H3K4me3, P300, and CTCF, and open chromatin (DNaseI), RNA-seq, and conservation at the locus of SOX2 gene in H1-ES cells.

Figure 3.

Chromatin organization at super-enhancers and stretch enhancers. (a-b) Spatial distribution of two Cohesin components, RAD21 and SMC3 (a) and CTCF (b) at super- and stretch enhancers from K562 and GM12878 cells.

Figure 4.

Transcriptional activity at super-enhancers and stretch enhancers. (a) Genome-wide profile of RNA Pol II at super- and stretch enhancers in H1-ESC, GM12878, K562, and HUVEC cell-lines. (b) Transcriptional abundance in reads per kilobase of transcript per million mapped reads (RPKM) of genes near (within a 50 kb window) super- and stretch enhancers across 10 cell types (P value <2.2e-16, Wilcoxon rank sum test). (c) GRO-seq profiles at the constituents of super- and stretch enhancers in K562 and GM12878 cell-lines. (d) GRO-cap profiles at the constituents of super- and stretch enhancers in K562 and GM12878 cell-lines.

Figure 5.

Overlap analysis of super-enhancers and stretch enhancers. (a) Fraction of overlap between super- and stretch enhancers across the ten cell types. (b) Pie chart of average overlap of super- and stretch enhancers. (c) The region length in base pairs (bp) of super-stretch and stretch enhancers across 10 cell types (p-value < 2.2e-16, Wilcoxon rank sum test).

Figure 6.

Cell type-specificity analysis of super, super-stretch and stretch enhancers. (a) K-means clustering on the histone modification H3K27ac profile at super, super-stretch and stretch enhancers in five ENCODE cell types (GM12878, K562, H1-ES, HepG2, and HUVEC). (b) Transcriptional abundance in units of RPKM of genes associated with H1-ESC and how these genes are expressed in the other nine cell types tested as shown along the axis. (c) GO analysis of super-, super-stretch and stretch enhancers in H1 ES cell type. (d) Venn diagram shows the overlap of genes associated with super, super-stretch and stretch enhancers, and label the known key cell-identity genes in H1 ES, K562, and Islets cells.