What is an enhancer?

Abstract

Tight control of the transcription process is essential for the correct spatial and temporal gene expression pattern during development and in homeostasis. Enhancers are at the core of correct transcriptional activation. The original definition of an enhancer is straightforward: a DNA sequence that activates transcription independent of orientation and direction. Dissection of numerous enhancer loci has shown that many enhancer-like elements might not conform to the original definition, suggesting that enhancers and enhancer-like elements might use multiple different mechanisms to contribute to transcriptional activation. Here, we review methodologies to identify enhancers and enhancer-like elements and discuss pitfalls and consequences for our understanding of transcriptional regulation.

Keywords: enhancer; enhancer-like element; facilitator element; polymerase II; transcriptional regulation.

FIGURE 1

Epigenomic enhancer signature. At a typical enhancer, transcription factors (TFs) bind to their binding motifs and recruit co‐activators including chromatin remodelers or modifiers such as the acetyltransferase p300. Thereby, the nucleosomes are displaced from the core of the enhancer and lead to higher accessibility that can be measured by ATAC‐seq for example. The surrounding nucleosomes are often marked by typical modifications found at enhancers such as H3K27ac or H3K4me1. In addition, enhancers themselves are transcribed, so RNA polymerase II (Pol II) and short RNAs originating from enhancers called eRNAs can used for enhancer identification. Using either ChIP‐seq or ATAC‐seq, a typical epigenomic enhancer signature (bottom) can be used for identification for putative enhancer elements.

FIGURE 2

STARR‐seq as an example for a massively parallel reporter assays (MPRA). (A) When integrated into the transcription unit, elements with intrinsic enhancer activity can activate their own transcription and subsequently be identified through RNA‐Seq. In this example, the enhancer is cloned into the 3′UTR, between a reporter gene and the polyadenylation signal. (B) In a typical MPRA experiment, a library of sequences is incorporated into the MPRA vector and transferred into a population of cells. After RNA‐extraction and next generation sequencing, the elements with intrinsic enhancer activity can be identified and their activity quantified.

FIGURE 3

Canonical and facilitator elements cooperate to activate ɑ‐globin expression. The super‐enhancer at the ɑ‐globin locus consists of two canonical enhancer elements, that show strong intrinsic enhancer activity, and three facilitator elements, that have low or no intrinsic enhancer activity, but are needed within the context of the enhancer cluster to ensure proper activation of the duplicated ɑ‐globin gene promoters. The facilitator elements are redundant, so only deletion of all three elements leads to a strong reduction of the target gene expression. The mechanisms underlying facilitator action on canonical enhancers and target gene promoters is not understood yet.