Measure it eighteen times
Our genes will be cut more accurately
Alexey Aleksenko, Forbes, 06.08.2018
Researchers from Texas have proposed a key improvement to the famous CRISPR gene editing system. Perhaps this will push the introduction of technology into clinical practice
Today, the main hopes for the deliverance of mankind from hereditary diseases are associated with the method of gene editing, discovered at the end of the first decade of the XXI century. The scientific name of this technique – CRISPR-cas9 – was learned by heart by popular science journalists around the world. But perhaps now they will have to retrain. A study by molecular biologists from the University of Texas (Strohkendl et al., Kinetic Basis for DNA Target Specificity of CRISPR-Cas12a) gives serious reasons to believe that it is more correct to say CRISPR-cas12a. To be more precise, the researchers have shown that replacing the key component of the system – the cas9 protein – with another protein can solve many problems of the method, over which genetic engineers have been struggling in recent years. And the main of these problems is selectivity and accuracy.
The problem of accuracy
The human genome does not tolerate typos. If, correcting a harmful mutation, geneticists accidentally damage other points of the chromosome, the harm from this will be much more than good: a small inaccuracy can lead to the development of a cancerous tumor. Meanwhile, at the current level of accuracy, the risk of such a development is unacceptably high. A few years ago, the whole world was shocked by the news that Chinese geneticists edited the genome of the human germ line for the first time. The opinions of the scientific community were divided: some welcomed the unprecedented scientific breakthrough, others warned against potential ethical conflicts. However, the main conclusion for which Huang Junjiu and his colleagues wrote their work lay aside from these disagreements: their experiments showed how inaccurate and ineffective the classical CRISPR-cas9 method works.
Instead of correcting the mutation of the beta-globin gene (the one that leads to the most severe hereditary disease beta-thalassemia), the enzyme in many cells simply spoiled the beta-globin gene, rewriting it on the model of another, similar – delta-globin. In some cells, the system did not work at all, and almost everywhere it introduced many undesirable mutations into the genome. So if the work of daring Chinese geneticists has demonstrated anything, it is how far researchers are still from the exact and safe application of this technology.
The device of the CRISPR-cas9 system is encrypted in its name. The first part means "grouped regularly alternating short palindromic repetitions". This strange object was discovered in the genome of bacteria at the end of the twentieth century. It turned out to be nothing more than a catalog of all kinds of viruses that the ancestors of the bacterium had to meet in their lives. When a viral chromosome enters a bacterium, the bacterium has a chance to quickly write down for itself a characteristic sequence of base letters identifying the virus. This way, at the next meeting, she will easily be able to recognize him.
But it is not enough to know, it must be destroyed. This is where the second part of the system comes into play – cas proteins. They are the ones who carry samples of viral sequences with them, which they use as a stencil: if they come across a nucleic acid with a matching signature, they immediately make an incision into it and destroy it. This property was used in the CRISPR-cas9 system. Having found a mutation in the human genome according to a given template, the cas9 protein cuts the chromosome in this place. If the "correct" copies of the gene are introduced into the cell at the same time, it will close the gap itself according to a new sample, and the error will be corrected. Unfortunately, in practice, the cas9 protein does its job sometimes too zealously, and sometimes carelessly. This leads to numerous errors.
Solving the problem
Rick Russell, Ilya Finkelstein and their colleagues set out to replace the weak link of the system – the cas9 protein – with something more suitable. Another component of the same bacterial system, the cas12a protein, turned out to be a suitable candidate. He does the same job, but approaches it in his own way.
Rick Russell offers the following comparison: cas9 with its ribonuclein stencil adheres to the target DNA like glue. It is enough for him to match just a few letters-bases to identify the target and proceed to its destruction (making a cut). That is why it reacts so sensitively to random partial coincidences that may occur in those places of the genome that the researchers did not plan to change.
The cas12 protein is not like that: its connection with DNA is more like a Velcro fastener. Each of the tiny villi-hooks (in this case, the "letters" of DNA) can provide a very weak connection, and only all of them together create a reliable contact. Moreover, if the "Velcro" is not fastened properly, you can always unbutton it and repeat the operation – with glue, such a trick will not work. Therefore, cas12a distinguishes the target sequence much better (where the letter match is complete) from random places in the genome (where most likely some letters will not "stick together"). He can "unbutton" and "fasten" the DNA several times before making sure he has found the right place for his attack. This protein does not begin to cut the chromosome without checking the coincidence of eighteen letters. Its cas9 counterpart is limited to only seven to eight.
Currently, the Rick Russell group continues to work on artificially modifying the cas12a, making it even more accurate. It is quite possible that it is on this path that geneticists will succeed in turning CRISPR technology into a reliable tool for clinical medical genetics.
Portal "Eternal youth" http://vechnayamolodost.ru