More accurate, powerful genetic analysis tool opens new gene-regulation realms

Apr 22, 2013

Researchers from Huntsman Cancer Institute (HCI) at the University of Utah have developed a novel and powerful technique to identify the targets for a group of enzymes called RNA cytosine methyltransferases (RMTs) in human RNA. They applied their technique to a particular RMT, NSUN2, which has been implicated in mental retardation and cancers in humans, finding and validating many previously unknown RMT targets—an indication of the technique's power. The research results were published online in the journal Nature Biotechnology on April 21.

"Although RMTs have been known for many years, virtually nothing is known about the majority of these enzymes in humans," said Bradley R. Cairns, co-author of the study and Senior Director of Basic Science at HCI. "This new technique will now allow the very rapid identification of the direct target RNAs for each human RMT, and we are excited about conducting that work."

Within all living cells, RNA acts as a critical intermediate in transmitting genetic information from DNA—RNA is made from DNA and then used to encode proteins called enzymes that control . A process called cytosine methylation attaches molecules to cytosine bases in DNA and . RMTs act as catalysts to allow methylation at particular locations in RNA molecules. Methylation can regulate the flow of (from RNA to ) in cells, and it can change the way RNA interacts with proteins.

RNA methylation is currently poorly understood, partly because of limitations in the technique currently used to identify which RNA molecules and cytosine bases are RMT targets. As each cell contains thousands of different types of RNA molecules, often with only a small percentage being targets for a specific RMT, the first step in a study of RNA methylation is to sort out and concentrate the precise target RNA molecules for a particular RMT, in a process called enrichment.

The work involved a novel enrichment method, which applied a special "chemical cross-linker" to physically join the RMT to an RNA that it is trying to methylate, said Vahid Khoddami, the study's co-author and a member of the Cairns Lab. "Our new technique takes advantage of the mechanism of the enzyme. The drug/crosslinker we used looks like cytosine, so it is incorporated in place of the cytosine in the RNA. The RMT tries to methylate this drug— thinking it is a normal target cytosine—but instead becomes crosslinked to the RNA, defining the precise location of the intended methylation. As our reaction-based method requires that the enzyme both bind the RNA and commit to the act of methylation, it greatly increases our identification of true positives," said Khoddami.

"This technique gives us 200-fold enrichment, when two-fold enrichment has been considered a great result in the past," said Khoddami. "In fact, for some RNA types, the enrichment is more than 700-fold."

After the enrichment process, high-throughput gene sequencing is used to analyze the RNA samples obtained.

"Our enrichment results were fantastic by themselves, but in the sequencing process we made another important discovery," Khoddami said. "We found that after sequencing, the target cytosine in the modified RNA instead appeared as an alternative molecule, guanosine, more than 50% of the time. After sequencing, you can look for these cytosine to guanosine transversions and know you have the precise target—in a single experiment."

According to Khoddami, ten RMTs are known in humans, and only two of them have been partially characterized. "None of the other eight have been studied in the laboratory," he explained, "although some of them have been shown to have connections to cancer, infertility, and particular genetic disorders in humans.

"These diseases have been puzzling because previously we did not have the tools to analyze the RNA. Now we have beautiful tools," said Khoddami.

Explore further: Heaven scent: Finding may help restore fragrance to roses

Related Stories

Gene's function may give new target for cancer drugs

Sep 12, 2012

(Phys.org)—Purdue University scientists have determined that a gene long known to be involved in cancer cell formation and chemotherapy resistance is key to proper RNA creation, an understanding that could one day lead ...

R-loops break walls of gene silencing

Mar 02, 2012

(Medical Xpress) -- Researchers at the University of California, Davis, have figured out how the human body keeps essential genes switched “on” and silences the vast stretches of genetic repeats and “junk” ...

Human cells can copy not only DNA, but also RNA

Aug 10, 2010

Single-molecule sequencing technology has detected and quantified novel small RNAs in human cells that represent entirely new classes of the gene-translating molecules, confirming a long-held but unproven hypothesis that ...

Process important to brain development studied in detail

Nov 07, 2011

Knowledge about the development of the nervous system is of the greatest importance for us to understand the function of the brain and brain disorders. Researchers at Uppsala University have examined the key step when genes ...

Recommended for you

Study on pesticides in lab rat feed causes a stir

Jul 02, 2015

French scientists published evidence Thursday of pesticide contamination of lab rat feed which they said discredited historic toxicity studies, though commentators questioned the analysis.

International consortium to study plant fertility evolution

Jul 02, 2015

Mark Johnson, associate professor of biology, has joined a consortium of seven other researchers in four European countries to develop the fullest understanding yet of how fertilization evolved in flowering plants. The research, ...

Making the biofuels process safer for microbes

Jul 02, 2015

A team of investigators at the University of Wisconsin-Madison and Michigan State University have created a process for making the work environment less toxic—literally—for the organisms that do the heavy ...

Why GM food is so hard to sell to a wary public

Jul 02, 2015

Whether commanding the attention of rock star Neil Young or apparently being supported by the former head of Greenpeace, genetically modified food is almost always in the news – and often in a negative ...

The hidden treasure in RNA-seq

Jul 01, 2015

Michael Stadler and his team at the Friedrich Miescher institute for Biomedical Research (FMI) have developed a novel computational approach to analyze RNA-seq data. By comparing intronic and exonic RNA reads, ...

User comments : 0

Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.