The "master list" totals 3,581 sites in which the enzyme ADAR might swap an "A" nucleotide for a "G" in an RNA molecule. Such a seemingly small tweak means a lot because it changes how genetic instructions in DNA are put into action in the fly body, affecting many fundamental functions including proper neural and gender development. In humans, perturbed RNA editing has been strongly implicated in the diseases ALS and Acardi-Gutieres disease.
The two two dark humps in the top row (representing fly RNA) are missing in flies that lack the enzyme to edit RNA (middle row) and missing in the DNA of normal flies. The analysis suggests they are sites of RNA editing.
Credit: Reenan lab/Brown University
The new list of editing sites could therefore help thousands of researchers studying the RNA molecules that are transcribed from DNA, the so-called "transcriptome," by providing reliable information about the thousands of editing changes that can occur.
"Drosophila serves as a model for all the organisms where people are studying transcriptomes," said the paper's corresponding author Robert Reenan, professor of biology in the Department of Molecular Biology, Cell Biology, and Biochemistry at Brown. "But in the early days of RNA editing research, the catalog of these sites was determined completely by chance – people working on genes of interest would discover a site. The number of sites grew slowly."
In fact, Reenan was co-author of a paper in Science 10 years ago that made a splash with only 56 new editing sites which at the time, more than doubled the number of known sites in the entire field.
Validation means accuracy
Several more recent attempts to catalog RNA editing sites have yielded larger catalogs, but those contained many errors (the paper provides a comparison between the new list and previous efforts such as ModENCODE).
To avoid such mistakes, Reenan and colleagues, including lead author and graduate student Georges St. Laurent, painstakingly validated 1,799 of the sites. They worked with Charles Lawrence, professor of applied mathematics and the paper's co-senior author, to predict another 1,782 sites and validated a statistically rigorous sampling of those.
In all, the team's methodology allowed them to estimate that the combined list of 3,581 directly observed and predicted sites is 87 percent accurate.
"The sites that we validated, for anyone who wants to do the same experiment under the same conditions, the sites should be there," said co-author and postdoctoral researcher Yiannis Savva. "In other papers, they just did sequencing to say there is an editing site there, but when you check, it's not there."
The researchers used the tried-and-true, decades-old Sanger method of sequencing to double-check all the candidate editing sites that they had found using the high-throughput technology called single molecule sequencing. They compared the sequenced RNA of a population of fruit flies to their sequenced DNA and to the RNA of another population of flies engineered to lack the ADAR editing enzyme. By comparing these three sequences they were able to see the A-to-G changes that could not be attributed to anomalies in DNA (i.e., mutations, or single-nucleotide polymorphisms) and that never occurred in flies incapable of editing.
As they conducted their validations, they fed the results back into their prediction algorithm. Over several iterations, that computer model "learned" to make better and better predictions. They ultimately found 77 different variables that helped them to distinguish real editing sites from nucleotides that were conclusively not editing sites.
The researchers then examined the implications of the patterns they saw in their data and gained several insights.
One was that a considerable amount of editing occurs in sections of RNA that do not code for making proteins. Editing is concentrated in a small number of RNAs, raising the question, Lawrence said, of what accounts for that selectivity.
"How does the cell go about choosing which ones are going to get edited and which aren't is an interesting question this opens," he said.
Where editing is found, the researchers discovered, there is usually more alternative splicing, which means the body is more often assembling a different recipe from its genetic instructions to make certain proteins.
The researchers also found that the RNAs that are most heavily edited tend to be expressed to a lesser extent, decreasing how often they are put into action in the body.
RNA editing helps explain why organisms are even more different from each other – and from themselves at different times — than DNA differences alone would suggest.
"RNA editing has emerged as a way to diversify not just the proteome but the transcriptome overall," Reenan said.
In addition to Reenan, Lawrence, Savva, and St. Laurent, who is also affiliated with the St. Laurent Institute in Cambridge, Mass., the paper's other authors are Michael Tackett, Sergey Nechkin, Dimitry Shtokalo, and Philipp Kapranov of the St. Lawernce Institute, Denis Antonets of the State Research Center of Virology and Biotechnology in Russia, and Rachael Maloney, a Brown graduate now at the University of Massachusetts Medical School.
Reenan received funding from the Ellison Medical Research Foundation.
David Orenstein | EurekAlert!
Colorectal cancer risk factors decrypted
13.07.2018 | Max-Planck-Institut für Stoffwechselforschung
Algae Have Land Genes
13.07.2018 | Julius-Maximilians-Universität Würzburg
For the first time ever, scientists have determined the cosmic origin of highest-energy neutrinos. A research group led by IceCube scientist Elisa Resconi, spokesperson of the Collaborative Research Center SFB1258 at the Technical University of Munich (TUM), provides an important piece of evidence that the particles detected by the IceCube neutrino telescope at the South Pole originate from a galaxy four billion light-years away from Earth.
To rule out other origins with certainty, the team led by neutrino physicist Elisa Resconi from the Technical University of Munich and multi-wavelength...
For the first time a team of researchers have discovered two different phases of magnetic skyrmions in a single material. Physicists of the Technical Universities of Munich and Dresden and the University of Cologne can now better study and understand the properties of these magnetic structures, which are important for both basic research and applications.
Whirlpools are an everyday experience in a bath tub: When the water is drained a circular vortex is formed. Typically, such whirls are rather stable. Similar...
Physicists working with Roland Wester at the University of Innsbruck have investigated if and how chemical reactions can be influenced by targeted vibrational excitation of the reactants. They were able to demonstrate that excitation with a laser beam does not affect the efficiency of a chemical exchange reaction and that the excited molecular group acts only as a spectator in the reaction.
A frequently used reaction in organic chemistry is nucleophilic substitution. It plays, for example, an important role in in the synthesis of new chemical...
Optical spectroscopy allows investigating the energy structure and dynamic properties of complex quantum systems. Researchers from the University of Würzburg present two new approaches of coherent two-dimensional spectroscopy.
"Put an excitation into the system and observe how it evolves." According to physicist Professor Tobias Brixner, this is the credo of optical spectroscopy....
Ultra-short, high-intensity X-ray flashes open the door to the foundations of chemical reactions. Free-electron lasers generate these kinds of pulses, but there is a catch: the pulses vary in duration and energy. An international research team has now presented a solution: Using a ring of 16 detectors and a circularly polarized laser beam, they can determine both factors with attosecond accuracy.
Free-electron lasers (FELs) generate extremely short and intense X-ray flashes. Researchers can use these flashes to resolve structures with diameters on the...
13.07.2018 | Event News
12.07.2018 | Event News
03.07.2018 | Event News
13.07.2018 | Event News
13.07.2018 | Materials Sciences
13.07.2018 | Life Sciences