Researchers detect unwanted effects of important gene manipulation system

May 30, 2014 by Josh Barney
U.Va.’s new approach identifies ways to help prevent the potentially dangerous “off-target” effects of a popular new gene-editing system, allowing scientists to improve their results.

Researchers at the University of Virginia School of Medicine have devised a way to detect unintended side effects of manipulating genes using a revolutionary new system that is sweeping the scientific world by storm.

The gene targeting system, called CRISPR, allows editing of at specifically targeted sites in the genome. U.Va.'s new approach reveals the system has the potential to bind to unintended sites and cause gene mutations at some of these sites – mutations that could have serious consequences for research and efforts to develop medical treatments.

U.Va.'s new approach, however, also identifies ways to help prevent those potentially dangerous "off-target" effects, allowing scientists to improve their results with this important new gene-editing system.

Gene Mutations and How to Avoid Them

A primary goal of gene manipulation is to correct harmful mutations, so it is vital to avoid introducing mutations unintentionally, explained U.Va.'s Mazhar Adli, of the Department of Biochemistry and Molecular Genetics.

"We know that genetic mutations are hallmarks of disease. The whole aim is to change these apparent genetic mutations – to go and correct these mutations," he said. "We want to change this information only at the targeted space, at the targeted locus. If you change any other information, basically you are introducing mutations that you don't want. You are correcting one gene and potentially you might be introducing mutations in 10 other genes and maybe many other places in the genome."

CRISPR: The 'Holy Grail of Research'

Adli's new research sheds light on the potential off-target effects of the CRISPR/Cas9 gene editing system. The system has proved extremely popular because it allows scientists to manipulate specific sections of the genome of living mammals, making it a tool of tremendous importance for scientific research. It has been adopted quickly and widely, including for work in human cells, because it is comparatively simple and because many labs have the resources to use it.

"You can basically target any genomic region in living cells and change the genetic information, which has been the holy grail of research for the past several decades," Adli said. "To be able to go and change the genetic information in living cells was a dream, basically."

U.Va.'s new research helps explain the mechanism that underpins the CRISPR system – and why it is vulnerable to off-target gene mutations.

"We not only found where it binds in the genome, we also investigated why it goes to these regions in the genome," Adli said. "By analyzing specific sequences underlying these off-targets, we also found out the determinants – why it goes to the on-targets and also to these off-target regions – and our research shows that it goes there because of some sequence similarity to the original targeted regions.

"So our results will help improve the specificity of the system so that we can minimize the off-targets."

Major Mutation Implications

Adli's work also showed that the naturally occurring form of a key enzyme used in the CRISPR system introduces far more mutations than an altered form of that enzyme that is less commonly used. The former cuts both strands of DNA during gene editing, allowing mutations to occur, while the latter snips only one strand, allowing cells to repair the damage without introducing .

"Unfortunately the wild-type form [the naturally occurring form] is much easier to deal with. Everyone in the field is using the wild type," he said. "Now with this paper, and with additional papers coming out, they will have to stop using the wild-type form of the enzyme. They have to use the [altered form] to overcome the off-targets. It is much superior and the off-targets are very dangerous."

Published Online

The findings have been published online by the journal Nature Biotechnology and will appear in a forthcoming print edition. The paper was written by U.Va.'s Cem Kuscu and Sevki Arslan, sharing credit as the lead authors; Ritambhara Singh; Jeremy Thorpe; and Adli.

Explore further: Precise gene editing in monkeys paves the way for valuable human disease models

Related Stories

Precise gene editing in monkeys paves the way for valuable human disease models

January 30, 2014
Monkeys are important for modeling diseases because of their close similarities to humans, but past efforts to precisely modify genes in primates have failed. In a study published by Cell Press January 30th in the journal ...

Recommended for you

Gene variant activity is surprisingly variable between tissues

August 21, 2017
Every gene in almost every cell of the body is present in two variants called alleles—one from the mother, the other one from the father. In most cases, both alleles are active and transcribed by the cells into RNA. However, ...

Genome analysis with near-complete privacy possible, say researchers

August 17, 2017
It is now possible to scour complete human genomes for the presence of disease-associated genes without revealing any genetic information not directly associated with the inquiry, say Stanford University researchers.

Science Says: DNA test results may not change health habits

August 17, 2017
If you learned your DNA made you more susceptible to getting a disease, wouldn't you work to stay healthy?

Genetic variants found to play key role in human immune system

August 16, 2017
It is widely recognized that people respond differently to infections. This can partially be explained by genetics, shows a new study published today in Nature Communications by an international collaboration of researchers ...

Phenotype varies for presumed pathogenic variants in KCNB1

August 16, 2017
(HealthDay)—De novo KCNB1 missense and loss-of-function variants are associated with neurodevelopmental disorders, with or without seizures, according to a study published online Aug. 14 in JAMA Neurology.

Active non-coding DNA might help pinpoint genetic risk for psychiatric disorders

August 16, 2017
Northwestern Medicine scientists have demonstrated a new method of analyzing non-coding regions of DNA in neurons, which may help to pinpoint which genetic variants are most important to the development of schizophrenia and ...

1 comment

Adjust slider to filter visible comments by rank

Display comments: newest first

not rated yet May 30, 2014
Doesn't this also link nutrient-dependent DNA repair via CRISPER to the epigenetic effects of nutrient-stress and social stress on the stability of the organized genome via one-carbon metabolism and DNA methlyation and amino acid substitutions, like glycine in microbes (for 1.8 billion years) and glycine in the GnRH decapeptide of vertebrates (for 400 million years)?

Or, perhaps ecological adaptations occurred more rapidly.


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.