The CRISPR gene-editing system is one of the most important medical breakthroughs of recent years, but that doesn't mean it's perfect. The most common enzyme in use is Cas9, which acts like a pair of molecular scissors and enables precise cut-and-paste DNA edits. But now, researchers have developed a new genetic tool that, they say, acts more like a motorized DNA "shredder."
The gene-editing technology was inspired by the self-defense mechanisms of bacteria, which lets them snip out short sections of an invading virus's DNA so it remembers how to fight it off next time. The tool, known as CRISPR, uses certain enzymes to make those cuts, and scientists have since co-opted the method for easier genetic engineering.
For years Cas9 has been the standard enzyme for the procedure, but others have been experimented with recently, which are said to have their own advantages. CasX, for example, is the smallest so it can get into cells easier, and since it comes from a non-pathogenic bacteria, there's lower risk of an immune response. Cas12a and Cas12b, meanwhile, are apparently more precise and safer to use.
Enter Cas3. This latest enzyme is a different beast altogether, for a number of reasons. First and foremost is its ravenous appetite for DNA sequences, allowing scientists to shred large segments instead of making small cuts.
"Cas9 is a molecular scissor that goes where you want it and snips once," says Yan Zhang, lead researcher on the study. "But Cas3 goes where you want it, travels along the chromosome, and makes a spectrum of deletions tens of kilobases long. This could make it a powerful screening tool to determine what large areas of DNA are most important for a particular disease."
In tests, the researchers delivered CRISPR-Cas3 into both human embryonic stem cells and HAP1 cells. They managed to use the tool to delete long DNA sequences measuring between a few hundred base pairs right up to 100,000.
The researchers say the shredder system could allow them to conduct lab tests on what certain sections of DNA do, by essentially destroying long segments and watching what changes. Or the shredding function could be stripped out, and Cas3 could be made into a harmless, long-range delivery system for epigenetic drugs.
Along with its longer range, the researchers say Cas3 is potentially more precise than other CRISPR enzymes. That's because it has a longer guide RNA sequence, and keeps the searching and degradation steps well separated, which should cut back on unintended edits – a concern with CRISPR-Cas9 use in humans. Still, further research is needed to check that this is the case.
Unlike the other enzymes, Cas3 is what's known as a Type I CRISPR, which is much more common in bacteria than Type II such as Cas9. In fact, this is the first time a Type I has been used outside of the host bacteria.
With the CRISPR toolbox continuing to grow, the future of genetic engineering – and medicine in general – looks to be fascinating.
The research was published in the journal Molecular Cell.
Source: University of Michigan
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