CRISPR has been one of the most influential breakthroughs of the last decade, but it’s still imperfect. While the gene editing tool is already helping people with genetic foods, scientists are also trying to improve on it.
The effort expanded the CRISPR family to include less harmful, more precise and smaller versions of the gene editor. But in the bacterial world, where CRISPR was originally discovered, we’re only scratching the surface. Two new papers suggest that an even more powerful gene editor may be around the corner — if it can be proven to work in cells like our own.
In one of the papers, Arc Institute scientists say they have discovered a new CRISPR-like gene-editing tool in bacterial “jumping genes.” Another article, written independently, covers the same instrument and extends the work to a similar one in a different family.
Jumping genes move within genomes and even between individuals. They have long been known to do this by cutting out and inserting their own DNA, but none of the machines have been shown to be programmable like CRISPR. In recent studies, scientists describe hopping gene systems that, in a process the teams alternatively call bridge and searchRNA editing, can be engineered to cut, insert and flip any DNA sequence.
Crucially, unlike CRISPR, the system does all this without breaking DNA strands or relying on the cell to repair them, a process that can be messy and unpredictable. The various molecules involved are also fewer and smaller than those in CRISPR, potentially making the tool safer and easier to deliver into cells and can handle much longer sequences.
“Bridge recombination can universally modify genetic material through sequence-specific insertion, excision, inversion, and more, enabling a word processor for the living genome beyond CRISPR,” said Berkeley’s Patrick Hsu, lead author of one of the studies and a core member of the Arc Institute. investigator in a press release.
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ToggleThe CRISPR shot
Scientists first discovered CRISPR in bacteria that fight off viruses. In nature, the Cas9 protein pairs with a guide RNA molecule to seek out viral DNA and, when located, cleaves it. Researchers have learned to look for this reengineering system none DNA sequences, including sequences found in human genomes, and break DNA strands at these locations. The cell’s natural machinery then repairs these breaks, sometimes using a provided strand of DNA.
CRISPR gene editing is powerful. It is being investigated in clinical trials as a treatment for various genetic diseases and received its first clinical approval late last year as a therapy for sickle cell disease and beta thalassemia. But it’s not perfect.
Because the system breaks DNA and relies on the cell to repair those breaks, it can be imprecise and unpredictable. The tool also works primarily on short stretches of DNA. While many genetic diseases are caused by point mutations where a single “letter” of DNA has been changed, the ability to work with longer sequences would expand the potential uses of this technology in both synthetic biology and gene therapy.
Over the years, scientists have developed new CRISPR-based systems that address these shortcomings. Some systems break only one strand of DNA or swap individual genetic “letters” to increase accuracy. Studies are also looking for other CRISPR-like systems by screening the entire bacterial universe; others have found naturally occurring systems in eukaryotic cells like ours.
The new work expands the quest by adding jumping genes into the mix.
The RNA bridge
Jumping genes are a fascinating feat of genetic magic. These DNA sequences can be moved between locations in the genome by machines that cut and paste. In bacteria, they even move between individuals. This gene sharing could be one way bacteria acquire resistance to antibiotics—one cell that has evolved to avoid a drug can share its genetic defenses with an entire population.
In the Arc Institute study, researchers looked at a specific jumping gene in bacteria called IS110. They discovered that when the gene is in motion, it calls upon an RNA sequence—like the guide RNA in CRISPR—to facilitate the process. The RNA contains two loops: One binds the gene itself, and the other locates and binds to the gene’s target site in the genome. It acts as a bridge between the DNA sequence and the specific site where it is to be inserted. Unlike CRISPR, once found, the sequence can be added without breaking the DNA.
“Bridge editing (cuts and inserts the DNA) in a one-step mechanism that recombines and religates the DNA so it leaves it completely intact,” Hsu said Wild biotechnology in email. “This is very different from CRISPR editing, which creates exposed DNA breaks that require DNA repair and has been shown to create adverse DNA damage responses.”
Significantly, the researchers found that both loops of RNA can be reprogrammed. This means that scientists can specify where the genome is and what sequence should be there. In theory, the system could be used to exchange long genes or even multiple genes. As a proof of concept in E-coli the team programmed IS110 to insert a DNA sequence nearly 5,000 bases long. They also cut and flipped another DNA sequence.
The study was joined by another paper written independently by another team of scientists at the University of Sydney, which details both IS110 and a related enzyme from another family, IS111, which they say is similarly programmable. In their paper, they called these systems “seekRNA”.
The tools rely on a single protein half the size of CRISPR. This means it may be easier to wrap them in harmless viruses or lipid nanoparticles – these are also used in Covid vaccines – and transfer them to cells where they can start working.
Another jump
This approach has great potential, but there is also a major caveat. So far, scientists have shown that it only works on bacteria. CRISPR, on the other hand, is incredibly versatile and has proven itself in countless cell types. They further hope to refine the approach further and adapt it to mammalian cells like ours. This may not be easy. The University of Tokyo’s Hiroshi Nishimasu says the IS110 family has not yet shown itself to be ready for such a task.
All this means that it is still early in the development of the technology. Scientists knew about CRISPR years before they showed it was programmable, and it wasn’t put to work in human cells until 2013. Although it has since moved relatively quickly from the lab to the clinic, the first CRISPR-based treatments took years longer than materialized.
At least the new work shows that we haven’t exhausted all that nature has to offer for gene editing. The technology could also be useful in the field of synthetic biology, where individual cells are engineered on a large scale to learn how life works in its most basic conditions and how we can reengineer it. And if the new system can be adapted to human cells, it would be a useful new option in the development of safer and more effective gene therapies.
“If it works in other cells, it will be a game changer,” said Sandro Fernandes Ataide, a structural biologist at the University of Sydney and author of the paper describing IS111. Nature. “It opens up a new field in gene editing.”
Image credit: The Arc Institute
