Where is the enhancer located

Thereof, where are enhancers located? Enhancers can be located upstream of a gene, within the coding region of the gene, downstream of a gene, or thousands of nucleotides away. When a DNA -bending protein binds to the enhancer , the shape of the DNA changes, which allows interactions between the activators and transcription factors to occur.

Secondly, what is the difference between a promoter and an enhancer? An enhancer is a sequence of DNA that functions to enhance transcription. A promoter is a sequence of DNA that initiates the process of transcription. A promoter has to be close to the gene that is being transcribed while an enhancer does not need to be close to the gene of interest. Silencers are antagonists of enhancers that, when bound to its proper transcription factors called repressors, repress the transcription of the gene.

Enhancers do not act on the promoter region itself, but are bound by activator proteins. Enhancers are DNA-regulatory elements that activate transcription of a gene or genes to higher levels than would be the case in their absence. These elements function at a distance by forming chromatin loops to bring the enhancer and target gene into proximity Enhancers can be found in many of the same areas that silencers are found , such as upstream of the promoter by many kilobase pairs, or even downstream within the intron of the gene.

How do I identify an enhancer? Enhancer elements require protein binding to exert their regulatory functions, and therefore tend to be in nucleosome-free chromatin regions.

Why are introns called introns? The parts of the gene sequence that are expressed in the protein are called exons, because they are expressed, while the parts of the gene sequence that are not expressed in the protein are called introns, because they come in between the exons. Are promoters transcribed? A promoter is a sequence of DNA needed to turn a gene on or off. Evidence for the formation of loops between long-range enhancers and promoters comes mainly from two strands of evidence.

The first is the cross-linking, by formaldehyde, and subsequent ligation together of enhancer and promoter DNA sequences as detected in chromosome conformation capture 3C -type methods. The second is the visualization, by fluorescence in situ hybridization FISH , of the spatial proximity of enhancer and promoter regions in the cell nucleus.

However, in some other cases in which bona fide regulatory elements can be captured by cross-linking to the appropriate gene promoter, visual assays do not detect a significant frequency of spatial co-localization between the enhancer and the promoter This may be because the chromatin loops are too transient to detect by FISH or because the 3C ligation products are established through indirect cross-linking of enhancers and promoters to relatively large — nm nuclear substructures or supramolecular complexes.

In the latter case, I would say that there is not a DNA loop as such being formed between the enhancer and the promoter. In cases where looping does occur, how do the loops form? Generally, the assumption is that enhancer—promoter looping serves to deliver factors for example, RNA polymerase, transactivators and transcription factors to the promoter in the right tissue and at the right time.

Since chromatin is a very large flexible polymer, the default conformation of which is not a series of structured loops, there must be specific mechanisms for stable loop formation. The directed formation of large loops by active chromatin bending would require considerable energy input, and we do not know of active mechanisms operating in interphase that can do this over such large distances.

However, chromatin continuously undergoes movement by constrained diffusion The radius of this constraint is sufficiently large that any two sequences within approximately 1 Mb of each other can randomly encounter each other in the nucleus. If there are protein complexes bound at the promoter and enhancer that have affinity for one another, a chromatin loop may then be stabilized through this passive mechanism.

Proteins that might be able to do this include those with dimerization or oligomerization domains and that are present at both the promoter and the enhancer. The formation of a loop, which juxtaposes sequences associated with multiple transcription, chromatin-modifying and chromatin-remodelling factors, will increase the local concentration of these factors and so promote the formation of further protein— protein and protein—DNA complexes.

Indeed, increased local protein concentration has been shown to be a key mechanism through which looping affects repression by the lac repressor So what about situations where DNA loops between enhancers and gene targets cannot be directly visualized in the nucleus? In some of these cases, the intervening chromatin seems to be in a compact state so that enhancer and promoter are still relatively close to each other — nm The high concentrations of transcription factors and protein complexes nucleated by enhancer binding could then simply diffuse through this restricted nuclear volume to find and activate transcription from the target promoter.

Diffusion might also be facilitated by nonspecific binding to the intervening chromatin, and indeed this type of scanning has been observed for the lac repressor in vivo Examples of proteins scanning the chromatin between enhancers and promoters have also been reported in eukaryotes Akin to the scanning model, the linking model proposes that chromatin complexes assembled at enhancers actively reorganize the chromatin between enhancers and promoters and is supported by evidence for propagation of histone modifications across the intervening chromatin 21 and by the activity of enhancer-blocking sequences It is harder to imagine the scanning and linking mechanisms operating at enhancers located hundreds of thousands of base pairs away from their target promoter, often with intervening genes that do not respond to the enhancer.

Nor is there any reason to think that all enhancers function through the same mechanism. Indeed, components of both one-dimensional and three-dimensional diffusion have been observed for the lac repressor in living E.

Ann Dean. Enhancers are DNA-regulatory elements that activate transcription of a gene or genes to higher levels than would be the case in their absence. These elements function at a distance by forming chromatin loops to bring the enhancer and target gene into proximity Recent data also suggest that insulator-binding proteins CTCF and cohesin may facilitate enhancer—promoter interactions.

How do enhancers affect transcription? Thus, it appears that enhancers serve as centres for the assembly of the pre-initiation complex PIC. Enhancers might be important for nuclear relocation of the enhancer—promoter pair to a neighbourhood that is favourable for transcription. There is evidence for each of these models, but important questions remain about the mechanistic details and how the models might relate to each other. Can Mediator bridge to activators bound at enhancers over long distances?

Indeed, in embryonic stem cells ESCs , Mediator subunits MED1 and MED12 co-localize with cohesin at enhancers and promoters, and cohesin is necessary for loop formation between them Thus, Mediator may coordinate enhancer signalling to the transcription machinery by interacting with enhancer-bound transcription factors and Pol II and serve as a hub for transcriptional regulation by distant enhancers. Although it is not clear whether these sites are bona fide enhancers, in at least one example, the distal site loops to a promoter in a TAF3-dependent fashion, and knockdown of TAF3 or CTCF reduces expression of the gene, suggesting that the loop is functional.

Enhancer looping also appears to have a role in Pol II elongation. The association of ELL3 with the enhancers was required for proper Pol II occupancy at developmentally regulated genes.

Together, these studies show that enhancers can influence both Pol II initiation and elongation through direct participation of transcription machinery components in looping. Another means by which enhancers could influence transcription of their target genes is through their own transcription. It has been known for many years that sense and antisense transcripts arise from certain enhancers, although the function of the transcripts was unclear.

Is the RNA or the transcription per se important, or is the transcription simply incidental to transcription of a looped gene? Now, genome-wide studies have revealed that enhancers are frequently transcribed into non-coding RNAs of various length, polyadenylation status and strand specificity 33 , 24 , Furthermore, enhancer RNAs eRNAs have been used to identify active enhancers, suggesting that enhancer transcription is a part of the enhancer activation process Transcription of the eRNAs correlated with mRNA synthesis at nearby genes, suggesting an involvement in transcription regulation 33 , It seems unlikely that the transcription is a by-product of activation of the target gene, since knockdown of a subset of the eRNAs resulted in decreased gene transcription An intriguing possibility is that eRNAs may have a structural role in establishing or stabilizing enhancer—promoter loops.

In fact, new data provide support for this idea Nevertheless, at this point, the function of enhancer ncRNAs requires considerable further study and validation. Recent studies document looping interactions between enhancers and promoters on a genome-wide scale. Comprehensive mapping of RNA Pol II-associated long-range interactions in different cell types suggested a structural framework of multi-gene complexes involving close enhancer—promoter interactions to accomplish cell-specific functions These studies of genome-wide enhancer looping have been tied together with eRNAs in a report indicating a significant correlation among gene expression, promoter—enhancer looping and transcription of the enhancers The picture that emerges is of an ensemble of enhancer—gene interactions that determine a specific cellular transcriptome.

How are these multi-gene complexes organized? The close approximation of active enhancer—gene pairs and similarly regulated genes fits well with the concept of transcription factories that are focal concentrations of RNA Pol II. This now seems likely to be a generalizable phenomenon. Can a connection be drawn between transcription factory residence and loops between enhancers and promoters? Enhancer loops might serve to deliver the activated gene to a transcription factory.

However, other scenarios can be envisioned, and this question remains to be rigorously addressed. Future work may broaden the perspective; however, thus far, mechanistic insights into how enhancers bring about gene expression all invoke looping. In some cases, looping can directly involve components of the transcriptional machinery. Moreover, looping may be influenced by enhancer transcription. Finally, enhancer looping on a genome-wide scale may organize active regions of the genome and may determine the destiny of certain genes for transcription factories.

Is enhancer looping sufficient for gene activation? How looping relates to transcription factory residence is an intriguing question. A single-cell technology for determining interaction frequency between sites in chromatin, comparable to the resolution obtained using FISH, would be a substantial advance.

Other pressing needs for the future are to determine in an unbiased way the proteins underlying nuclear enhancer—promoter loop organization and to uncover how movement in the nucleus orders the landscape for gene expression. Q How do mutations and variants in enhancers influence human disease?

Marcelo A. It is not surprising, thus, that genetic variation within these regulatory sequences has the potential of resulting in phenotypic variation and underlies the aetiology of human diseases. In the absence of mutations in the globin genes, the disease emerged as a consequence of the disruption in the linear relationship between the globin genes and their distant cis -regulatory elements Over the past decade, genomic sequencing efforts confirmed these predictions and afforded a better understanding of the pervasiveness of mutations in distant cis -regulatory elements — the vast majority of which are enhancers — underlying human diseases The picture that has gradually emerged from these studies is that regulatory mutations result in both Mendelian and complex disease traits, that their frequency spectra range from rare to common and that their phenotypic effects range from small to large.

However, the functional characterization of putative disease-causing regulatory mutations remains an important challenge, and most mechanistic demonstrations resort to experimental strategies that involve large amounts of labour, cost and time. Genetic variation in distant enhancers has been linked to several human Mendelian disorders. In an early demonstration of this, mutations in an enhancer controlling the expression of sonic hedgehog SHH from a megabase away was shown to result in pre-axial polydactyly in families.

This phenotype is shared with patients carrying a chromosomal translocation that removes this enhancer from the general vicinity of SHH However, the phenotypic impact of mutations in enhancers may vary substantially from that of protein-coding mutations, even if both are connected to the same gene.

Mutations in enhancers are largely limited to cis effects on transcription, whereas those in protein-coding sequences may alter broader aspects of gene expression, such as RNA processing and stability, protein folding, and so on Another central distinction between the impact of coding and non-coding mutations relates to the modularity of distant enhancers: each enhancer of a gene is responsible for a subset of the quantitative, temporal and spatial expression of that corresponding gene.

As an example, coding mutations in TBX5 — a gene involved in heart and forelimb development — results in Holt—Oram syndrome, which is characterized by cardiac and forelimb malformations. Smemo et al. While most regulatory mutations leading to disease that have thus far been characterized disrupt pre-existing enhancers, gain-of-function mutations are also likely to participate in disease processes. De Gobbi et al.

Thus, the mutational space of non-coding sequences, already estimated to be much larger than that of coding sequences, is likely to be an underestimation of the true figure. The modularity of enhancers and their functional compartmentalization imply that regulatory mutations will often have a lower burden on fitness than will coding mutations and may reach high frequency in populations.

As a prelude to understanding how common variation in distal enhancers might underlie the genetic architecture of several complex traits and diseases in humans, Emison et al. These regulatory variants often reach high frequency in populations and are predicted to affect disease risk through small phenotypic effects, contrasting with the large effect Mendelian variants discussed above.

The precise identification of disease-causing regulatory variants within GWAS loci remains an important challenge, especially in terms of the experimental validation of the putative functional effects of these variants. Nevertheless, a number of regulatory variants in enhancers emerging from GWAS hits have been functionally characterized, and several insights have come out of these studies.

First, the same variant may have an impact on the risk for more than one disease 50 , Second, new mechanisms of disease have been uncovered or confirmed, such as an altered response to inflammatory signalling underlying the risk of coronary artery disease Finally, the detailed characterization of a genetic or signalling pathway associated with a disease may reorient the target of biological exploration.

However, recent data posit that non-pancreatic actions of TCF7L2 may in fact underlie the increased disease risk to T2D 54 — Clear challenges remain to be addressed both in the identification of regulatory variants contributing to human diseases and the experimental interrogation of the impact of those variants on biological processes. New technologies that effectively assay functional variants, teasing out their biological impact in high throughput, will be necessary to replace the laborious and low-throughput experimental strategies used thus far.

Extrapolating the notion of mutation burden and aggregate analysis used in exome sequencing to regulatory elements will prove a formidable task, and yet it is likely that the genetic architecture of several common diseases will include various regulatory mutations in multiple enhancers within an individual.

Finally, the almost exclusive focus of regulatory mutations on distal enhancers reflects our inability to assay functionally other types of regulatory elements in the genome.

Yet other classes of regulatory elements, such as insulators, repressors and matrix attachment regions, are abundant in the human genome and almost certainly have their function modified by rare and common genetic variants. The development of new experimental assays to interrogate these elements and their putative allelic variants will contribute to uncovering the genetic mechanisms of several human diseases that have as their molecular base variants in distal cis -regulatory sequences.

Gill Bejerano. Modern technologies driven by next-generation sequencing, such as ChIP—seq, that reveal all genomic DNA in a particular functional state provide breathtaking snapshots of gene regulation in action.

We see large amounts of open chromatin, dynamic domains of histone modifications and many thousands of binding sites for virtually any transcription factor and co-factor under any condition How many of these biochemical events that we observe actually contribute to gene regulation is an open question. How this landscape is exactly divided between enhancers and other gene regulatory regions, such as repressors and insulators, and indeed how many of these elements have multiple roles under different cellular conditions is only starting to unfold.

The evidence we have suggests that a large fraction of gene regulatory regions can act as enhancers 4 , 5. By virtue of occupying so much genome landscape, enhancers provide a large potential target for evolution. Genome evolution is driven by mutation and selection. An adaptive genomic mutation can take hold by improving fitness in at least one context that the locus is used in while not perturbing function too much in any other context of its use.

Most human genes are expressed in multiple cells and tissues at different times. A gene sequence mutation may perturb the organism in all contexts where the transcript is in use, increasing the likelihood of a detrimental effect. The expression domain of a gene, however, is often the sum of inputs from multiple enhancers and other cis -regulatory elements , each active in only a subset of contexts for example, ref.

Thus an enhancer mutation may affect a smaller subset of functional contexts. If it happens to be beneficial in one context, there are fewer other contexts to reconcile with. This modularity makes enhancers even more likely fodder for evolution Framed by our growing interest in them, there are still a number of fascinating fundamental questions to be addressed at a number of levels about the contribution of enhancers to evolution.

For example, we do not know how small or simple enhancers can be at their birth. The handful of enhancers that have been studied in detail are bound by multiple transcription factors over dozens of bases We also see numerous co-regulated groups of genes in multiple contexts, but our understanding of enhancer logic and gene networks is not deep enough to know how sequence-constrained the enhancers driving these gene groups must be.

The more complex and constrained we presume an enhancer must be to contribute to fitness, the less likely it is that functional enhancers are to arise de novo in neutrally evolving DNA. Duplication and divergence of pre-existing enhancers, including in the context of gene duplication, is an appealing model.

An enhancer's orientation may even be reversed without affecting its function. Furthermore, an enhancer may be excised and inserted elsewhere in the chromosome, and still affect gene transcription. That is the reason that intron polymorphisms are checked though they are not transcribed and translated.

Currently, there are two different theories on the information processing that occurs on enhancers: [2]. Category : Gene expression. Read what you need to know about our industry portal chemeurope. My watch list my. My watch list My saved searches My saved topics My newsletter Register free of charge. Keep logged in. Cookies deactivated. To use all functions of this page, please activate cookies in your browser.