The strategy promises to clarify the longstanding mystery of the role played by vast stretches of DNA sequence that do not code for the functional units—genes—that nevertheless may have a powerful regulatory influence. The research is described in the 12 February edition of the journal Nature.
"Our approach employs next generation sequencing technology to find regulatory regions, the 'switches' on a genome-wide scale and much more cost effectively," said Pennacchio. "It's the next layer of knowledge that's been missing."
The DOE JGI was founded in 1997 to accelerate the completion of the HGP and completed the DOE's commitment to sequence three (5, 16, 19) of the 23 chromosomes, totaling 11 percent of the human genome, and published the analysis in Nature back in 2004.
In this newly published study, Pennacchio, lead authors DOE scientist Axel Visel and postdoctoral fellow Matthew Blow, and their colleagues, describe a shortcut for identifying gene regulatory regions or the molecular switches that turn on or off gene expression.
Using what's called ChIP-Sequencing or ChIP-Seq, chromatin immunoprecipitation (ChIP) is combined with massively parallel DNA sequencing to identify binding sites of DNA-associated proteins.
Traditionally researchers have relied on evolution to guide them to non-coding sequences that are likely to have a function—such as enhancing the expression of genes. Via the public genome databases, they would align the entire human genome code with that of other vertebrate species (e.g. other mammals, birds, frogs, fish) and then look for sequences that are conserved in evolution.
"Most protein-coding sequences show signs of conservation between species, but there is also a large number of non-coding sequences that have been surprisingly well conserved for tens or even hundreds of millions of years," said Visel. "This suggests that these regions, formerly thought to be "junk" DNA, actually have some functional relevance and are under selection because sequence changes reduce fitness of affected individuals. Using such sequence conservation, we have in previous studies identified enhancer candidate regions and shown in transgenic mouse experiments that these conserved non-coding regions are in fact often enhancers that are active during embryonic development. Conservation-based methods are relatively good at finding enhancers in the genome, but an important limitation is that they don't tell us where and when that particular enhancer would be active and thereby drive the expression of its neighboring target gene(s).
The older methods lacked specificity, Blow said. "For example, if we have a gene that is important both for brain and for limb development, we would not have been able to specifically identify the enhancer sequences near that gene that would drive the expression in the brain or limb, the only way to find out was to test these activities in experiments one-by-one, which is slow and can't be done on a genomic scale.
"Using this new method, we can directly identify a genome-wide set of enhancers that are active in a particular anatomical region or tissue at a particular time-point, which is an important advantage over conservation-based methods because in addition to telling us where an enhancer is located in the genome, it also provides an initial experimental characterization where we should expect this enhancer to be active."
The team used ChIP linked with a particular enhancer-associated protein, p300, then directed DOE JGI's massively parallel next generation sequencing capacity to map several thousand sites in mouse embryonic forebrain, midbrain and limb tissue. Over 80 of these fragments were tested in transgenic mouse experiments indicating an almost perfect success rate of p300-ChIP-Seq for identifying enhancers active in vivo.
"Enhancers are especially important for regulating genes during embryonic development," said Pennacchio. "They can regulate genes over long distances and switch on their target genes during very specific time-points and in very specific anatomical structures during development. There are several examples of mutations in such enhancers that cause disease in humans because genes are not expressed at the right time or in the right place anymore. A fundamental problem in studying such enhancers is that until recently we did not have effective tools to even find them in the genome on a large scale.
Pennacchio said that this new method will prove useful to the greater genomics and biomedical community for characterizing the role of the vast non-coding regions—dubbed genome "dark matter"—about which little is known.
"These datasets will also help to identify mutations in enhancers that play a role in human disease," Pennacchio said. "Human genetic studies indicate that in many cases disease is caused by mutations in non-coding sequences, but it has been difficult to study this in detail because the function of most non-coding sequences is poorly understood. Eventually, this will be useful for purposes including disease detection and personalized medicine."
With the rapidly increasing efficiency and cost-savings of the next generation sequencing technologies, a deluge of data from individual human genomes are being to come to light, to the point where whole-genome sequencing of patients may soon become a standard diagnostic tool.
"While progress is being made towards this goal, it is important to keep in mind that our current understanding of the genome has focused on protein-coding sequences," said Pennacchio. "Datasets like the one provided through this study will be important to understand the remaining 98 percent of the genome and what its role in health and disease is."
The published study provides an important proof of principle to establish and validate a new method in three different mouse tissues at a single embryonic time-point, Pennacchio said. "We can now generate genome-wide enhancer datasets directly from human tissues and compare genome-wide sets of enhancer activities between healthy people and people suffering from disease, which may reveal how enhancer activities change on a global scale in these disease states."
David Gilbert | EurekAlert!
Further reports about: > ChIP-Seq > ChIP-Sequencing > DNA > DNA sequence > DNA-associated proteins > Genom > Nature Immunology > biofuels production > chromatin immunoprecipitation > embryonic development > human genome > molecular tools > multicellular organisms > non-coding sequence > on-off switches in genomes > protein-coding genes
Cancer diagnosis: no more needles?
25.05.2018 | Christian-Albrechts-Universität zu Kiel
Less is more? Gene switch for healthy aging found
25.05.2018 | Leibniz-Institut für Alternsforschung - Fritz-Lipmann-Institut e.V. (FLI)
The more electronics steer, accelerate and brake cars, the more important it is to protect them against cyber-attacks. That is why 15 partners from industry and academia will work together over the next three years on new approaches to IT security in self-driving cars. The joint project goes by the name Security For Connected, Autonomous Cars (SecForCARs) and has funding of €7.2 million from the German Federal Ministry of Education and Research. Infineon is leading the project.
Vehicles already offer diverse communication interfaces and more and more automated functions, such as distance and lane-keeping assist systems. At the same...
A research team led by physicists at the Technical University of Munich (TUM) has developed molecular nanoswitches that can be toggled between two structurally different states using an applied voltage. They can serve as the basis for a pioneering class of devices that could replace silicon-based components with organic molecules.
The development of new electronic technologies drives the incessant reduction of functional component sizes. In the context of an international collaborative...
At the LASYS 2018, from June 5th to 7th, the Laser Zentrum Hannover e.V. (LZH) will be showcasing processes for the laser material processing of tomorrow in hall 4 at stand 4E75. With blown bomb shells the LZH will present first results of a research project on civil security.
At this year's LASYS, the LZH will exhibit light-based processes such as cutting, welding, ablation and structuring as well as additive manufacturing for...
There are videos on the internet that can make one marvel at technology. For example, a smartphone is casually bent around the arm or a thin-film display is rolled in all directions and with almost every diameter. From the user's point of view, this looks fantastic. From a professional point of view, however, the question arises: Is that already possible?
At Display Week 2018, scientists from the Fraunhofer Institute for Applied Polymer Research IAP will be demonstrating today’s technological possibilities and...
So-called quantum many-body scars allow quantum systems to stay out of equilibrium much longer, explaining experiment | Study published in Nature Physics
Recently, researchers from Harvard and MIT succeeded in trapping a record 53 atoms and individually controlling their quantum state, realizing what is called a...
25.05.2018 | Event News
02.05.2018 | Event News
13.04.2018 | Event News
25.05.2018 | Event News
25.05.2018 | Machine Engineering
25.05.2018 | Life Sciences