"We researched how the yogurt bacterium protects itself against viruses, we did not plan in advance to develop a gene editing system"

This is what Prof. Virginios Shikshinis, one of the first scientists who isolated Crispr/Cas9 from the yogurt bacterium's immune system and turned it into a gene editing tool that is about to transform the health and agriculture systems, says in an interview with the scientist website. The interview took place during the LIFE SCIENCES BALTICS 2016 conference

Virginijus Siksnys is a molecular biologist at Vilnius University in Lithuania. He became interested in CRISPR in 2007, when the scientists in the group he led, who were working with the yogurt bacterium, first noticed the strange patches in the bacteria's DNA in short clusters that repeated themselves regularly - the same repeats that gave CRISPR its name are actually part of an ancient immune system of a bacterium designed to fight viruses. The pieces of DNA between the repeats were sequences of the virus, which actually marked the pathogens so the bacteria could fight them. The bacterium has Crispr-associated proteins - the CAS in CAS9, which seem to use these segments to cut the genetic material of the invading virus."

Prof. Shikshinis was naturally one of the guests of honor at the Life Sciences Baltics conference that took place about two weeks ago in Vilnius, the capital of Lithuania, and where this interview for the science website also took place.

"In my laboratory we didn't know how to make cheese or yogurt, but we knew how to work with E.coli" says Shikshinis in a lecture he gave at the LIFE SCIENCES Baltics conference plenary. That's why we took the Crispr and CAS sequences from the yogurt bacteria and attached them to E.coli cells, which became The bacterium is resistant to several types of viruses. In Koli Island, the CAS genes could be deleted one after the other and in 2012 Shikshinis identified that one of these proteins CAS9 is solely responsible for cutting the DNA. "

"In fact, every living thing can be edited, and by chance, just a few days before the interview, the US Department of Agriculture announced that plants or animals used in agriculture, and which were treated with Crispr/Cas9, are not considered genetically modified because no foreign DNA was inserted into them. According to Prof. Shikshinis, in a conversation with the science site, it remains to be seen whether this recognition will lead to widespread use, and that there will still be enough opponents that it will be difficult to convince them. "

“Crispr/Cas9 is a tool that actually uses a pair of molecular numbers to edit DNA. It is so accurate and easy to use that it has taken the biology world by storm. Hundreds if not thousands of labs are now using Crispr/Cas9 for everything from developing super-muscular pigs, to extracting HIV genes from infected cells, to growing transgenic monkeys for neuroscience experiments. "

As Prof. Shikshinis pointed out in his lecture, he stands on the shoulders of giants, when he described the entire development of the discovery that in its first versions had not yet revealed the promise inherent in the tool. The discovery did not come from the brilliance of one genius but from the gradual work of an entire body of research. "

"In May 2012, they submitted a paper detailing exactly how CAS9 cuts DNA to the journal PNAS. The peer review editors returned the article repeatedly and asked additional questions. The problem is that while the peer review process was taking place, a German American group (Dodna and Scherpentier) submitted a similar article to the journal Science that was published much faster (the main difference is that the two, unlike Shikishins, showed how two parts of the RNA that CAS9 needs to function can connect to a chimeric fragment). The article by Shikshinis and his group was published a second time and did not receive all the fame. Also a third group – Zhang and Church from Howard published a paper in February 2013 showing how Crispr/Cas9 modifies the DNA of human cells in culture. "

"According to Prof. Shikshinis, the article was also submitted to two other journals CELL and CELL REPORTS but they rejected it and then they moved to PNAS, which cost them the delay and the fame of the discoverers."

"In an interview with the science website, he explains: "My background is in chemistry and then I switched to research in biochemistry. It was important for me to understand how it works in vitro from the chemical point of view, how CAS9 works. In the first step, we isolated the proteins and studied the substances that bind to these proteins. We decided that it is not enough to do genetic experiments and we need to approach the issue from the biochemical angle - to extract proteins from the cell and show how the proteins work. When we learned how they work, we realized that we had a powerful system in our hands."

"This was a discovery that resulted from a search for an answer to basic questions. Our first goal was not to discover a gene editing system but to discover how the bacteria protect themselves against viruses, then we found CAS9, which protects the bacteria and understood how it works. Then we figured out how to reprogram it easily. I worked with Restriction Enzyme enzymes that cut DNA many years before CRISPR and they do the same action as CAS9 although through different mechanisms, however they were difficult to program. I left all other interrupting enzymes and switched to working with CAS9. Absurdly now I'm back to work with some of them. I am investigating how other CRISPR systems work, hoping that this understanding will lead to the development of new tools and applications that may be difficult for CAS9 to perform. In other words, I went back to basic science.”

What are you really doing today?
I work as part of an international team of universities from Western Europe, Ukraine and three universities in Lithuania - University of Vilnius, University of Kaunas and the School of Medical Sciences in Kaunas that deals with the field of aging, and in particular the genetic mutations that cause what we call senile diseases. I deal with the basic science, there are also doctors on the team who are looking for faster applications, among other things, using CRISPER/CAS9.

What do you think about biohackers' use of CRISPR kits?
Prof. Shikshinis": "It depends on what they do with them. I am always in favor of expanding scientific pursuits outside the laboratories."

Will the method lead to the end of genetic diseases?
"In the first step, the method will be good for diseases related to mutations in a single gene, but unfortunately, many diseases are caused by mutations in many genes, so it will be necessary to edit the genome in many places and it will be difficult. "

Is there not playing God here?
"CRISPER/CAS9 is a tool that opens up millions of possibilities. At a certain point, you stop thinking about these red lines. No one would object to correcting mutations that cause dangerous diseases. Conversely, there will be applications that are limited by ethics. For example, genetic improvement. It would be possible but I don't think it's the right way to go. "

Prof. Shikshinis, who today can work and research wherever he wants around the world says that he prefers to stay in the villa, especially in light of the huge investment in the Integrated Center for Life Sciences, as I described in the visit notes there in the article The paradise of life science researchers.