Finding distal regulatory elements in the human genome

Abstract

Transcriptional regulation of human genes depends not only on promoters and nearby cis-regulatory elements, but also on distal regulatory elements such as enhancers, insulators, locus control regions, and silencing elements, which are often located far away from the genes they control. Our knowledge of human distal regulatory elements is very limited, but the last several years have seen rapid progress in the development of strategies to identify these long-range regulatory sequences throughout the human genome. Here, we review these advances, focusing on two important classes of distal regulatory sequences-enhancers and insulators.

Figure 1

Different classes of cis-regulatory elements in a typical human genome. Transcription initiates at promoters (blue DNA), which are further activated by enhancers (green DNA) or repressed by silencers (red DNA). The activity of enhancers and silencers may be confined by insulators (yellow DNA), which also prevent the spreading of repressive condensed chromatin structures (shown at each end of this chromosomal region). This model depicts nucleosomes as DNA (gray helix) wound around histone proteins (various colors), which are less dense at exposed DNA regulatory elements when bound by various transcription factors (blue ovals), activator and repressor proteins (green and red ovals, respectively), and CTCF (yellow oval).

Figure 2

Three mechanisms by which enhancers act to enhance transcription at target promoters. (a) Transcription factors recruit nucleosome-remodeling complexes containing SWI/SNF proteins (green ovals), ATPases that can slide nucleosomes along the DNA in an ATP-dependent fashion. The resulting change of nucleosome positions at the target promoter (blue DNA) facilitates assembly of the transcription machinery (blue ovals) and gene expression (represented by green block arrow). (b) Transcription factors also recruit coactivator complexes (green ovals) with histone acetyltransferase (HAT) activities, which in turn lead to histone acetylation (represented by stars on the nucleosomes). Such chromatin modification may provide binding surfaces for other activator proteins (additional green ovals) to bind, or result in decondensation of chromatin to allow transcription machinery (blue ovals) easier access. (c) Transcription factors may also recruit the mediator complex (purple ovals), which directly interact with RNAPII to activate transcription.

Figure 3

A strategy to map enhancers based on their chromatin signatures. (a) Derivation of the average chromatin modification profiles from known enhancers. Enhancers (green DNA) are flanked by nucleosomes containing mono-methylated histone H3 lysine 4 (H3K4me1, represented by single pentagons). Plotting H3K4me1 enrichment relative to chromosomal position for many enhancers yields an average enhancer H3K4me1 profile or ‘chromatin signature.’ (b) Scanning the chromatin signature along the genome (fading green profile) to discover regions with similar chromatin profiles. A new enhancer is identified based on correlation of its H3K4me1 profile to the enhancer chromatin signature (see thick green profile), whereas other regulatory elements lacking this signature (insulators, yellow; promoters, blue) are passed over by the scanning algorithm. Promoters are characterized by tri-methylated and di-methylated H3K4 (represented by triple and double pentagons), and acetylation (stars) may be found at either kind of regulatory element.

Figure 4

Enhancer activity in the absence of CTCF. The effect of CTCF knockdown in human cells is depicted in this model, illustrating that activation of transcription (green block arrows) occurs due to loss of the enhancer-blocking insulator function, as described in Ref. [22^•]. When CTCF (yellow ovals) is knocked down by siRNA (red no symbol), enhancers (green) are free to upregulate promoters (blue) that were previously separated by insulators (yellow DNA) bound by CTCF.

Figure 5

Looping model of enhancer function. During transcriptional activation, an enhancer (green) is positioned in close proximity to the target promoter (blue) via DNA looping.