A systematic search of viral genomes has unearthed a wealth of potential CRISPR-based genome editing tools.
CRISPR-Cas systems are widespread in the microbial world of bacteria and archaea, where they often help cells fight off viruses. But an analysis1 published on 23.11 cell finds CRISPR-Cas systems in 0.4% of publicly available genome sequences of viruses capable of infecting these microbes. Researchers believe the viruses use CRISPR-Cas to compete with each other — and possibly also to manipulate gene activity in their host to their advantage.
Some of these viral systems have been able to edit plant and mammalian genomes, and possess properties – such as compact structure and efficient editing – that could make them useful in the laboratory.
“This is a significant advance in discovering the tremendous diversity of CRISPR-Cas systems,” says computational biologist Kira Makarova of the US National Center for Biotechnology Information in Bethesda, Maryland. “There are many new things to discover here.”
Although CRISPR-Cas is best known as a tool for altering genomes in the laboratory, it can function as a rudimentary immune system in nature. About 40% of the sampled bacteria and 85% of the sampled archaea have CRISPR-Cas systems. Often these microbes can capture parts of an invading virus’ genome and store the sequences in a region of their own genome called a CRISPR array. CRISPR arrays then serve as templates to generate RNAs that instruct CRISPR-associated (Cas) enzymes to cut the appropriate DNA. This may allow microbes carrying the array to cut up the viral genome and potentially stop viral infections.
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Viruses sometimes pick up snippets of their hosts’ genomes, and researchers had previously found isolated examples of CRISPR-Cas in virus genomes. If these stolen pieces of DNA give the virus a competitive advantage, they could be preserved and incrementally modified to better serve the viral lifestyle. For example a virus that infects the bacterium Vibrio cholera uses CRISPR-Cas to cut and disable the DNA in the bacterium that encodes antiviral defenses2.
University of California, Berkeley molecular biologist Jennifer Doudna and microbiologist Jillian Banfield and their colleagues decided to conduct a broader search for CRISPR-Cas systems in viruses that infect bacteria and archaea called phages. To their surprise, they found about 6,000 of them, including representatives of all known types of CRISPR-Cas systems. “Evidence suggests these are systems that are useful for phages,” says Doudna.
The team found a variety of variations on the common CRISPR-Cas structure, with some systems missing components and others being unusually compact. “Even though phage-encoded CRISPR-Cas systems are rare, they are very diverse and widespread,” says Anne Chevallereau, who studies phage ecology and evolution at the French National Center for Scientific Research in Paris. “Nature is full of surprises.”
Small but efficient
Viral genomes tend to be compact, and some of the viral Cas enzymes were remarkably small. This could offer a particular advantage for genome editing applications, as smaller enzymes can be more easily introduced into cells. Doudna and her colleagues focused on a particular cluster of small Cas enzymes called Casλ and found that some of them could be used to edit the genomes of laboratory-grown thale cress cells (Arabidopsis thaliana), wheat and human kidney cells.
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The results suggest that viral Cas enzymes could become part of a growing collection of gene-editing tools discovered in microbes. Although researchers have discovered other small Cas enzymes in nature, many of these have so far been relatively inefficient for genome editing applications, Doudna says. In contrast, some of the viral Casλ enzymes combine both small size and high efficiency.
In the meantime, researchers will continue to scour microbes for potential improvements to known CRISPR-Cas systems. Makarova expects scientists will also look for CRISPR-Cas systems taken up by plasmids — pieces of DNA that can be transferred from microbe to microbe.
“Thousands of new genomes become available every year, and some of them come from very different environments,” she says. “So it gets really interesting.”
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