A consortium of scientists working at MIT, Harvard, Tsinghua University, Columbia University, and the Rockefeller University announced in the January 3 online edition of Science that they had devised a way to enable simultaneous editing of several sites within the mammalian genome.
Precise genome editing would have, the authors say, powerful applications across basic science, biotechnology and medicine. The technology, based on a bacterial defense system against viruses, could offer an easy-to-use, less-expensive way to engineer organisms that produce biofuels, to design animal models to study human disease, and to develop new therapies, among other potential applications.
They further note that it could be used to treat human diseases, either inserting missing or dysfunctional genes, or removing harmful genetic elements with much more precision than any available techniques.
To create their new genome-editing technique, the researchers modified a set of bacterial proteins that function as part of the regularly interspaced short palindromic repeats or CRISPR system. Using this system, scientists can alter several genome sites simultaneously and can achieve much greater control over where new genes are inserted, said Feng Zhang, an assistant professor of brain and cognitive sciences at MIT and leader of the research team.
Although genome-editing technologies such as designer zinc fingers (ZFs), transcription activator-like effectors (TALEs) and homing meganucleases have begun to enable targeted genome modifications, the authors noted, a need remains for new technologies that scalable, affordable, and easy to engineer. The scientists developed a new class of precision genome engineering tools they said, based on the RNA-guided Cas9 nuclease from the prokaryotic CRISPR adaptive immune system.
Unlike previous “hit or miss” methods to insert foreign DNA into mammalian genomes, or more complex methods like transcription activator-like effector nucleases (TALENs) that can also cut the genome in specific locations, but are expensive and difficult to assemble, the new system is much more user-friendly, Dr. Zhang says.
Making use of naturally occurring bacterial protein-RNA systems that recognize and snip viral DNA, the researchers created DNA-editing complexes that include a nuclease enzyme, Cas9, bound to short RNA sequences. These sequences can be designed to target specific locations in the genome; when they encounter a match, the Cas9 nuclease cuts the DNA.
And each of the RNA segments can target a different sequence. “That’s the beauty of this—you can easily program a nuclease to target one or more positions in the genome,” Zhang says, noting that it can be used either to disrupt the function of a gene or to replace it with a new one. To replace the gene, the researchers must also add a DNA template for the new gene, which would be copied into the genome after the DNA is cut.
The method is also very precise; if there is a single base-pair difference between the RNA targeting sequence and the genome sequence, Cas9 is not activated.
While the scientists have to date studied the editing system in mouse and human cells, the system could be used to design new therapies for diseases such as Huntington’s disease, caused by a single abnormal gene. Clinical trials that use zinc-finger nucleases to cut out the gene and replace it with the normal version are now under way, and this new technology could offer an efficient alternative. The investigators also pointed out that system might also be useful for treating HIV by removing patients’ lymphocytes and mutating the CCR5 receptor, through which the virus enters cells. After being put back in the patient, such cells would resist HIV infection.
The investigators plan to apply the new technology to study brain function and diseases.