For genome editing: details
A meeting in a cafe that changed the life of mankind: some details of the invention of genetic scissors
Julia Rudy, Vesti
The 2020 Nobel Prize in Chemistry was awarded to two women scientists Emmanuelle Charpentier from the Max Planck Institute in Germany and Jennifer Dudna from the University of California at Berkeley, USA.
The official wording of the Nobel Committee is "for the development of genome editing techniques" (for rewriting the code of life, as the host of the ceremony Yeran Hansson put it).
To be more precise, we are talking about the CRISPR/Cas9 method, which we have repeatedly told readers about, and which representatives of the Nobel Committee call genetic scissors. Today, this revolutionary technique has already made it possible to overcome oncological diseases, severe genetic blood diseases in the course of experimental studies, helped to create a powerful weapon against RNA viruses and obtain tuberculosis-resistant cows. And this is only a small part of the opportunities that have opened up to researchers thanks to the work of Charpentier and Dudna.
Biochemists and cell biologists can now easily investigate the functions of various genes and their possible role in the progression of diseases. Breeders can now not spend many years developing the necessary traits by plants or animals, but can immediately endow representatives of flora and fauna with specific characteristics, such as the ability to withstand drought in warmer climates and the invasion of insect pests. In medicine, the CRISPR/Cas9 gene editor is used to develop new cancer treatments and in research aimed at treating previously incurable hereditary diseases.
Initially, Emmanuel Charpentier investigated the immune system of bacteria of the genus streptococci. These pathogens are very common and often infect people. The researcher hoped to find a new form of antibiotics to fight these microbes.
In 2002, when Emmanuelle Charpentier founded her own research group at the University of Vienna, she focused on one of the bacteria that causes great harm to humanity: Streptococcus pyogenes.
Every year, this pathogen infects millions of people, often initiating easily treatable infections such as tonsillitis and streptoderma. However, it can also cause life-threatening sepsis and destroy the soft tissues of the body.
But how can S. pyogenes be neutralized? Charpentier began her work with a thorough study of how the genes of this bacterium work.
Meanwhile, Jennifer Dudna is studying RNA molecules, the molecular sister of DNA. In 2006, she became the head of a research group at the University of California, Berkeley, with twenty years of experience working with RNA.
Jennifer's group is studying RNA interference, which plays an important role in protecting cells from viruses.
Bacteria and their ancient immune system
One day Dudna learns from a microbiologist colleague about a new discovery. Researchers who studied the genetic material of completely different bacteria, as well as archaea, found repeating fragments in DNA. The same code appears over and over again, but there are unique sequences between these repetitions that differ from each other.
Scientists called the find CRISPR (short for clustered regularly interspaced short palindromic repeats). Interestingly, the non-repeating sequences were similar to the genetic code of various viruses. It was as if bacteria and archaea were making archival records in their DNA about the attacks of pathogens, interspersing them with repetitive code.
This led scientists to believe that CRISPR is one of the parts of the ancient immune system that protects bacteria and archaea from viruses.
Dudna's colleague noted that the mechanism used by bacteria to neutralize the virus is very similar to what Jennifer's group is studying (the same RNA interference).
Complex machinery
Plunging into this topic, Dudna found out that other researchers also discovered special genes, which they called CRISPR-associated (abbreviated Cas).
Jennifer understands that these genes are very similar to the genes that are responsible for the work of proteins that specialize in unwinding and cutting DNA strands. Maybe Cas proteins have the same function? Maybe they break down the DNA of viruses?
The subsequent years of work of her scientific group allowed to study in detail the work of Cas proteins.
Meanwhile, Charpentier is moving to a university in Sweden and is studying small RNAs found in bacteria of the species S. pyogenes.
She discovers that small unknown RNA molecules are present in large quantities in this bacterium. And for some reason, these RNAs are very similar in structure to the CRISPR sequences in the bacterial genome. Later it will turn out that these RNA molecules play a very important role.
In the meantime, it becomes clear that S. pyogenes needs the Cas9 protein to cleave viral DNA.
Charpentier shows that an unknown RNA molecule is necessary for an ordinary "long" RNA (created according to the CRISPR template) to become active.
In the spring of 2011, at a scientific conference in Puerto Rico, she meets Jennifer, after which the scientists decide to study the functions of Cas9 in S. pyogenes together?
They suspect that CRISPR is necessary to identify the virus, and Cas9 is a kind of scissors that cut the DNA molecule of the virus and thereby neutralize it.
However, when scientists conduct experiments "in vitro", nothing happens. The pathogen's DNA molecule remains intact.
After a long brainstorming session and many unsuccessful experiments, the researchers add the same unknown RNA molecules to the test tube. And... the system is doing the right job!
Researchers discover a weapon that streptococci have developed to protect against viruses, simple and effective, even brilliant.
The story of genetic scissors could have ended there. But luck accompanies prepared minds.
Epochal experiment
The researchers decided to use the genetic scissors they discovered to change the genetic code. Using their new knowledge, they obtained a molecule that allowed them to "set" scissors on a certain section of DNA (the so-called RNA guide).
The results of the experiment were stunning. The DNA molecules were split in exactly the right places.
Shortly after Emmanuel Charpentier and Jennifer Doudna published an article in 2012 about the discovery of the CRISPR/Cas9 genetic scissors, several research groups demonstrated that this tool could be used to change the genome in both mice and human cells.
Previously, changing genes in a cell, plant or organism took a long time, and sometimes it was simply impossible. Now, the researchers could potentially cut any DNA molecule.
This tool was so easy to use that it is now used in many fundamental research.
Animal experiments have shown that it is possible to deliver genetic scissors to the right cells.
At the same time, the development of genetic scissors has caused serious ethical disputes. After all, potentially with their help it is quite possible to create a child "to order" (for example, with blue eyes, blond hair or other necessary signs). Therefore, this area requires separate regulation.
However, all this does not negate the fact that the huge long-term work of Emmanuelle Charpentier and Jennifer Dudny allowed us to develop a chemical instrument that brought the life sciences to a completely new stage of development.
Let's add that the CRISPR-Cas9 technique does not work perfectly yet. But today, many scientists in the world offer ways to improve it, which will increase its effectiveness (for example, increase the number of edited genes). In 2017, scientists also proposed a tool that allows treating diseases without changing DNA.
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