Mini-Cas
A "viral" version of the CRISPR/Cas9 editor was found in the genome of bacteriophages
Studying bacteriophages, the discoverers of the CRISPR/Cas9 genomic editor, which is based on the "antiviral" proteins of microbes, discovered its more compact and convenient analogue. The results of their research were published by the scientific journal Science (Pausch et al., CRISPR-CasΦ from huge phages is a hypercompact genome editor).
"This viral protein, CasΦ, copes with the assigned tasks no worse than its bacterial counterpart, Cas9. However, at the same time, it is about half its size. This is very important, since such an enzyme is much easier to deliver into the cell than Cas9. Now this is one of the main barriers to widely applying CRISPR/Cas9 in practice," she said Jennifer Dudna, professor at the University of California at Berkeley (USA) and one of the authors of the study.
Diseases and infections affect not only humans and other multicellular living beings, but also unicellular microbes. Viruses, as scientists suggest, appeared almost simultaneously with bacteria, and there has been a continuous war for survival between them for several hundred million years.
Over millions of years of evolution, viruses have gradually learned to bypass the attention of the protective systems of microbes. In response to this, the bacteria have developed a kind of genetic "antivirus" – the CRISPR/Cas9 system. It finds traces of viral DNA in the genome of the microbe, cuts it out or causes the bacterium to self-destruct. Bacteriophages, in turn, have learned to suppress the work of CRISPR/Cas9, and the biological "arms race" has entered a new round.
Studying the vicissitudes of this "war" allowed Dudna and her colleagues, as well as two other groups of scientists, to adapt CRISPR/Cas9 to solve completely different tasks – editing the genome of humans and other multicellular living beings. Over the past five years, scientists have discovered several other varieties of this bacterial "antivirus" that surpass CRISPR/Cas9 in editing accuracy or allow you to change the structure of RNA molecules, not DNA.
Virus "antivirus"
Studying the genomes of large bacteriophage viruses from the Biggiephage family, Dudna and her colleagues found an even more interesting version of this system. Dudna's colleagues recently found these bacteriophages in temporary drying lakes, in cowsheds and in the forest floor soaked with water.
Over the past few years, biologists have already found analogues of the Cas9 gene in the genomes of viruses, but their DNA or RNA did not have the second part of this genomic editor, the CRISPR "library". In contrast, the genome of several viruses from the Biggiepage family has a full–fledged, but at the same time very compact version of CRISPR/Cas, whose protein – CAS - is similar in structure and function to the bacterial enzyme Cas12a. It works more selectively and more accurately than Cas9. Φ in this case is not the Cyrillic letter "ef", but the Greek "phi" – a common abbreviation for bacteriophages.
After analyzing the contents of this very compact CRISPR library, biologists suggested that this system is not designed to fight bacteria, but to destroy "competitors" – other viruses that have already penetrated the cell and are trying to multiply inside it. The viral "antivirus" recognizes fragments of their genome and forces the microbe to destroy them, clearing its living space.
Dudna and her colleagues tested this hypothesis in practice by injecting small amounts of three varieties of CasΦ into ordinary E. coli. Experiments have shown that this protein really destroyed fragments of the genome, similar to viral RNA or DNA, if they had sequences that are present in a set of templates from the CRISPR library.
At the same time, CasΦ turned out to be more versatile than many of its bacterial "competitors". This enzyme can work not only with single-stranded DNA, but also with a double helix, which simplifies the genome editing procedure. Coupled with the small size of CasΦ, which allows it to fit inside retroviruses, this makes the protein especially interesting from a practical point of view, the scientists conclude.
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