Epigenetics is the study of mechanisms which affect gene expression without changing the sequence of DNA. Many of these mechanisms are heritable across generations. Picturing Waddington’s Epigenetic landscape, in 2018 we would imagine it more like a seabed; containing different layers, with each contributing a part to the overall topology on the surface. We can picture each undersea peak as a stem cell state. From here a rock would follow a trajectory mapped by the surface topology, ending in a hollow (a differentiated state). The layers of information are diverse and many have only begun to be investigated. In this short introduction, we’ll deal with some current threads and interesting new discoveries in the field, focusing on three broad themes of epigenetics.
Epigenetics are an emerging field, where do you see the field could go in the next 10 years?
The epigenome will be a key component of multiomics. Indeed epigenetics play major roles in the interaction between the environment, the genome, the proteome, and the transcriptome. In addition, researchers have unveiled a number of specific epigenetic modifications and mechanisms related to disease and continue to evaluate the role epigenetics plays in various disorders. Epigenetic anomalies have been linked to a staggering number of disorders such as cancer, diabetes, mental retardation, and anxiety to name a few. There is still more work to do, and scientists are still in the discovery phase of pinpointing the epigenetic mechanisms underlying many conditions. As these mechanisms become better understood, the prospect of clinical solutions and therapies targeting abnormal epigenetic modifications will become more promising within the next 10 years.
Philippe Cronet, Business Unit Manager at Diagenode
The cytosine bases in DNA can have a methyl group covalently added to them. Despite being the first described epigenetic modification1-3, DNA methylation represents the paradigm of epigenetic modification that can be heritably transmitted. The modification is initiated on unmethylated DNA by DNA Methyltransferase (DNMT) 3A and 3B 4,5, with the help of DNMT3L and maintained on hemimethylated DNA by DNMT16,7. CpG islands are regions of the genome which are rich in unmethylated CpG dinucleotides and appear to have a special immunity to methylation8. Their deregulation is linked to cancer9 and they were recently found to have a tight interplay with histone methylation10,11. Methylated cytosines can be converted into hydroxymethylated, formylated or carboxylated intermediates 12-14, on their way to being demethylated. These intermediates also play a role in gene regulation13,15-17. Despite the fact that DNA methylation represses gene expression, remarkably hydroxymethylation, corresponding to the addition of a single hydroxyl group, is enough to change the outcome of DNA methylation and generally lead to activation13. Much recent work has also focused on other DNA modifications, such as the methylation of adenine resides18-20.
Not long ago we still didn’t know much about DNA methylation, do you think there is still more to uncover about this particular DNA modification?
Researchers are still conducting intensive research around DNA methylation marks such as 5-mC, 5-hmC, 5-fC, or 5-caC. New technologies such as single time molecule sequencing promise to uncover marks that are still unknown. In addition, more research is needed and underway for identifying biomarkers for diseases related to abnormal DNA methylation patterns. The mechanisms of DNA methylation in a number of diseases is not yet fully understood – and only until research elucidates these mechanisms can appropriate therapies be devised.
As a last point, DNA is not the only nucleic acid of interest in epigenetics research. The study of RNA modifications will become increasingly important.
Philippe Cronet, Business Unit Manager at Diagenode
Each cell contains metres of DNA, which rather than getting tangled around the nucleus, is wrapped around protein structures known as nucleosomes21. These are the basic units of chromatin, which physically store our genetic information. Each nucleosome is made up of eight core histones (two H2A, two H2B, one H3 and one H4 histone) and a linker histone known as H122-24. All of these core histones can be chemically modified by the addition of a plethora of marks by dedicated enzymes. This functionally impacts on DNA-based processes like gene expression and can regulate them through two main mechanisms involving either the disruption of chromatin contacts (between nucleosomes) or the recruitment of non-histone proteins. Histone modifications include acetylation, methylation, phosphorylation, ubiquitylation, sumoylation, ADP-ribosylation, arginine deamination, arginine methylation and proline isomerisation25,26. Importantly most of these marks are dynamic and a host of enzymes are responsible for their removal, a mechanism that can also directly influence gene expression25. More recently, a host of acylations such as butyrylation and propionylation were discovered, all having discrete roles in the control of gene expression27. Modifications occur on the flexible tails issuing from each histone tail, but also on the lateral surface of the nucleosome28,29. Interestingly, there is a close interplay between histone modification and DNA methylation 10,11,30-32. Histone modification is another type of epigenetic modification which can transmit information to the next generation and many studies are currently trying to unravel the heritable nature of histone modifications and how this affects cell phenotype over time.
The plethora of histone modifications is overwhelming, how much do we understand about them and are we still discovering more of them?
Of course. Single events (e.g. a histone modification that is related to a physiological disorder) are easy to uncover, but understanding the complexity of interactions related to histone modifications is a big task. Most of the histone modifications have been identified, researchers have uncovered the various types of histone modifications such as methylation, acetylation, deacetylation, ubiquitination, phosphorylation, sumoylation, etc, but numerous modification sites have yet to be discovered in ongoing epigenetics research. Additionally, the large number of histone modifications make their functions complicated to understand. Researchers still need to understand how the presence of multiple modifications on the same histone, in combination with histones variants regulates gene expression. Mass spectrometry combined with ChIP and transcriptomics data and the emerging single cell technologies will provide with a great wealth of data to address this question in the coming years.
Philippe Cronet, Business Unit Manager at Diagenode
Epigenetics, as well as detailing the distribution of chemical modifications to DNA and its surrounding proteins, deals with all ways in which gene expression can be altered without changing coding sequences. This includes the packaging of DNA within the cell. The cell is broadly divided into regions of dense packing known as heterochromatin, and more loose packing known as euchromatin, each of them being characterised by a different set of histone modifications. Heterochromatic genes tend to be repressed, whereas euchromatic ones have higher expression33. However, the packing of DNA occurs on many levels. Regulatory elements such as enhancers and promoters interact with each other to control gene expression34-36. This also occurs on a much larger scale and interaction maps produced by HiC, a technique which fixes and then sequences the regions of chromosomes which interact with one another, can show these interactions in more detail, proving the existence of Topologically Associated Domains (TADs)37,38 and their role in the regulation of DNA-based processes. Entire chromosome regions can form ‘territories’ which pack together like items in a shopping bag, jostled by gene expression. Finally, some regions of the genome attach to the nuclear lamina, and these tend to be heterochromatic and late replicating39,40, showing a further association of positioning with gene expression regulation.
In conclusion, DNA methylation, histone methylation and its derivatives, and nuclear structure and organisation all have an influence on gene expression without altering the underlying coding sequence of the DNA. Collectively, they are known as epigenetics and form a multi-faceted gene control system, each layer forming the input to the next, responding to, initiating and cementing gene expression change. Much current work is focused on finding other epigenetic factors which could heritably regulate gene expression, for example RNA modification, which could form the next layer in the landscape, influencing gene expression and transmitting information over time.
Epigenetics is a multi-faceted field, how far are we from utilising what we have learned about its various facets, from applying epigenetics to treating disease?
We still have a lot of basic research and discovery to do before epigenetic therapy is a reality. Currently, 5 epigenetics drugs are available on the market today, mainly anticancer drugs. Epigenetic targets are increasingly gaining interest from the pharmaceutical sector, and a plethora of new compounds are under investigation, particularly histone modification reader modulators such as bromodomains. In 2017, there were over 30 ongoing clinical trials addressing epigenetic modulating compounds. In the shorter term, basic research continues to reveal epigenetic signatures and specific biomarkers related to disease, which will continue to be useful for clinical knowledge and drug discovery.
Philippe Cronet, Business Unit Manager at Diagenode
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