Searching for the Gene-Editing Off-Switch

CRISPR is revolutionary, but some scientists are anti-CRISPR. Ironically, their work will actually help improve the gene-editing tool.


No scientist in the field will deny it – biology is in a CRISPR revolution. One recent review called the gene-editing technique “one of the most disruptive tools in biology”. In spite of this, two Toronto-based researchers are decidedly anti-CRISPR.

CRISPR-based systems offer the ability to edit genomes with an accuracy and efficiency unparalleled by any other gene-editing system. The problem is that as long as the system is active it will be searching for DNA to edit, leading to possible off-target DNA cuts and edits. These potential off-target effects – which could lead to cancer formation among other unintended consequences – represent a major hurdle in translating the CRISPR gene-editing system to a viable therapy for genetic diseases such as cystic fibrosis and muscular dystrophy.

What we need is a CRISPR off-switch – something that could stop the CRISPR system once the intended gene editing has occurred, avoiding any off-target effects.

The anti-CRISPR scientists from the University of Toronto, Alan Davidson and Karen Maxwell, have been searching for this off-switch.

The CRISPR gene-editing system is borrowed from nature, adapted from bacteria’s natural defence systems against invading viruses, called phages. Davidson and Maxwell are looking to nature again for their CRISPR off-switch, asking “how do phages fight back against the CRISPR defense system?” Does a CRISPR off-switch exist in the defensive arsenal of phage proteins? It turns out that the answer is yes!

Davidson and Maxwell, working alongside one of the pioneers of CRISPR gene-editing, Jennifer Doudna from the University of California in Berkeley, and others, recently characterized two phage-derived proteins, which they called anti-CRISPR proteins. These proteins are capable of inactivating the CRISPR effector enzyme Cas9, effectively stopping CRISPR from working.

The two proteins prevent CRISPR function in distinct ways: one protein binds to and blocks the site on the Cas9 enzyme that cuts the DNA, while the other protein causes pairs of the Cas9 enzyme to bind to each other, which prevents the enzyme from binding to the target DNA.

The work was published last month in the journal Cell. In the article, the authors suggest that these two anti-CRISPR proteins likely represent only a tiny subset of the anti-CRISPR proteins employed by phages. After all, the battle between bacteria and phages represents an evolutionary arms race that dates back millions of years.

By learning more about these anti-CRISPR proteins, scientists will be able to better control CRISPR gene-editing. But avoiding off-target effects is only one of several possible applications of anti-CRISPR proteins. They could also be used to restrict gene-editing to a specific tissue or organ, or to only allow gene-editing at specific times during the cell-cycle. Ultimately, anti-CRISPR proteins provide a mechanism to fine-tune CRISPR gene-editing.

By being anti-CRISPR, Davidson and Maxwell are actually leading the way to a much more powerful CRISPR gene-editing system.

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Steven is a PhD candidate in the department of Molecular Genetics at the University of Toronto. He is passionate about CRISPR, computer programming, and science communication. Along with Research2Reality, Steven regularly contributes to the Ontario Institute for Regenerative Medicine as a writer for the Expression.