- Researchers have uncovered two distinct anti-CRISPR proteins — AcrIIA2 and AcrIIA4 — that worked to inhibit the ability of SpyCas9 to target specific genomes in bacteria.
- The discovery could lead to an “off switch” for CRISPR that would give scientists greater control over the gene editing tool when using it in human subjects.
As it stands, CRISPR is a rather impressive gene editing tool and already the most precise method we have available for genetic manipulation. Studies like this recent one from UC Berkeley are helping us refine the CRISPR-Cas9 system, and now a new study published in Cell from UC San Francisco (UCSF) is offering a way to deal with some of its greatest remaining downsides.
Researchers discovered a way to switch off this gene-editing system using recently identified proteins discovered in the lab of Joseph Bondy-Denomy from UCSF’s Department of Microbiology and Immunology. These anti-CRISPR proteins could eventually improve the safety and accuracy of already very accurate CRISPR applications, and the researchers relied on a nifty little trick to discover them.
“Just as CRISPR technology was developed from the natural anti-viral defense systems in bacteria, we can also take advantage of the anti-CRISPR proteins that viruses have sculpted to get around those bacterial defenses,” explains the leader of the study, Benjamin Rauch.
In their research, the team looked for bacterial strains that had been infected by a known virus. They reasoned that their existence would be evidence that a bacteria’s Cas9 was not functioning properly.
“Cas9 isn’t very smart,” according to Bondy-Denomy. “It’s not able to avoid cutting the bacterium’s own DNA if it is programmed to do so. So we looked for strains of bacteria where the CRISPR-Cas9 system ought to be targeting its own genome — the fact that the cells do not self-destruct was a clue that the whole CRISPR system was inactivated.”
After examining nearly 300 strains of Listeria using Rauch’s bioinformatic approach, the team found three strains that showed this evidence. From those, they isolated four distinct anti-CRISPR proteins, and of these four, test showed that two — dubbed AcrIIA2 and AcrIIA4 — worked to inhibit the ability of SpyCas9 to target specific genomes.
A Better System
“These natural Cas9-specific ‘anti-CRISPRs’ present tools that can be used to regulate the genome engineering activities of CRISPR-Cas9,” the researchers write. They believe that with these proteins, it’s possible to avoid unintended or “off-target” gene modifications caused by keeping CRISPR’s machinery active in the body.
Of course, the next step would be to see how these proteins function in human cells. “We also want to understand exactly how the inhibitor proteins block Cas9’s gene targeting abilities, and continue the search for more and better CRISPR inhibitors in other bacteria,” Raunch explained.
Ultimately, this “off-switch” for CRISPR could prove almost as important as the system itself. “Researchers and the public are reasonably concerned about CRISPR being so powerful that it potentially gets put to dangerous uses,” Bondy-Denomy said. “These inhibitors provide a mechanism to block nefarious or out-of-control CRISPR applications, making it safer to explore all the ways this technology can be used to help people.”