In a new study, researchers from at the Massachusetts Institute of Technology McGovern Institute have discovered a bacterial enzyme that they say can expand the CRISPR toolbox used by scientists. It can easily cut and edit RNA , and before that, this accuracy was only applicable to DNA editing. This bacterial enzyme, which they eventually named Cas7-11, modifies RNA targets without harming cells, indicating that in addition to being a valuable research tool, it also provides a fertile platform for therapeutic applications. related research results were published online in Nature on September 6, 2021. The title of the paper is " Programmable RNA targeting with the single-protein CRISPR effector Cas7-11 ".
Co-corresponding author of the paper and McGovern Institute researcher Omar Abudayyeh mentioned the DNA-cutting enzyme Cas9 that has made modern biology faster, cheaper and more precise, and said, "This new bacterial enzyme is like Cas9 that edits RNA." He added, "It cuts twice on the target with precision and does not destroy the cell during the cutting process like other enzymes."
So far, only one other RNA targeting enzyme family, namely Cas13, has been widely developed for RNA targeting applications. However, when Cas13 recognizes its target, it crushes any RNA in the cell, thereby destroying the cell by the way. Like Cas9, Cas7-11 is part of a programmable system; it can be directed to specific RNA targets through the CRISPR guide. Abudayyeh, McGovern researcher Jonathan Gootenberg (another co-corresponding author of the paper) and their colleagues discovered Cas7-11 through in-depth exploration of the CRISPR system found in the microbial world.
Explore natural diversity
Like other CRISPR proteins, Cas7-11 is used by bacteria as a defense mechanism against viruses. After encountering a new virus, bacteria using the CRISPR system record viral infections in the form of small fragments of viral genetic material. If the virus reappears, the CRISPR system will be activated, guided by a small piece of RNA, to destroy the viral genome and eliminate the infection. These ancient immune systems are extensive and diverse, and different bacteria deploy different proteins to fight their viral invaders.
paper co-author and evolutionary biologist Eugene Koonin of the National Center for Biotechnology Information Center said, “Some CRISPR proteins target DNA, and some CRISPR proteins target RNA. Some CRISPR proteins are very effective at cleaving targets. But some are toxic, while others are not. They introduce different types of cleavage, and they can be different in specificity---and so on."
Abudayyeh, Gootenberg, and Koonin have been studying genome sequences, To understand the natural diversity of the CRISPR system and analyze them as a potential tool. Abudayyeh said the idea is to take advantage of what evolution has already done to transform protein machinery.
When these authors used public databases to study the components of different bacterial defense systems, a protein of the bacterium Desulfonema ishimotonii isolated from Tokyo Bay, Japan, caught their attention. Its amino acid sequence indicates that it belongs to a class of CRISPR systems that use large multi-protein complexes to find and cut their targets. But this protein seems to have everything it needs,This task can be done independently. Koonin said that other known single-protein Cas enzymes, including the Cas9 protein that has been widely used for DNA editing, belong to a separate category of the CRISPR system, but Cas7-11 blurs the boundaries of the CRISPR classification system. They eventually named this protein Cas7-11.
Image source: doi:10.1038/s41586-021-03886-5
Cas7-11 is very attractive from an engineering point of view because a single protein is easier to deliver Into the cell, and is easier to become an editing tool than complex multi-protein complexes. But its composition also marks an unexpected evolutionary history. These authors found evidence that through evolution, the components of the more complex Cas machine fuse together to form the Cas7-11 protein. Gootenberg is equivalent to discovering bats when you previously thought that birds were the only animals that could fly, thus realizing that there are multiple evolutionary paths to achieve flight. He said, "This completely changes the way people think about the CRISPR system, both in terms of function and evolution." When they came out of the Cas7-11 protein and began to experiment with it, they realized that this unusual enzyme provided a powerful means of manipulating and studying RNA. When they introduced it into the cell together with the guide RNA (gRNA), it cut its RNA target precisely, while other RNAs were not interfered. This means that they can use Cas7-11 to change the specific base in the RNA code to correct errors caused by genetic mutations. They can also program Cas7-11,To stabilize or destroy specific RNA molecules in cells, which gives them the ability to adjust the level of proteins encoded by these RNAs.
Abudayyeh and Gootenberg also found that the ability of Cas7-11 to cleave RNA can be inhibited by a protein that seems to be also involved in triggering programmed cell death, suggesting that there may be a relationship between CRISPR defense and a more extreme response to infection connect.
These authors discovered that a gene therapy vector can deliver a complete Cas7-11 editing system to cells, and Cas7-11 does not harm the health of the cells. They hope that with further development, this enzyme may one day be used to edit disease-causing sequences in patients' RNA so that their cells can produce healthy proteins or down-regulate the levels of proteins that cause damage due to genetic diseases.
Gootenberg said, "We believe that the unique cutting method of Cas7-11 can achieve many interesting and diverse applications. No other CRISPR tool can cut RNA with such precision. This is another good example of these foundations. Biology-driven exploration can produce new tools for treatment and diagnosis. We must have only touched the surface of natural diversity." (Bioon.com) Ahsen Özcan et al. Programmable RNA targeting with the single-protein CRISPR effector Cas7-11. Nature, 2021, doi:10.1038/s41586-021-03886-5.
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