From the human brain to coral diseases

What problems are solved by bioinformatics at Skoltech

Five young scientists, researchers at the Skoltech Center for Systems Biomedicine and Biotechnology, told TrV-Science about their research and whether it is possible to do science in Russia today. The questions were asked by Nadezhda Markina.

How to turn off life and start back up

Pavel Mazin, Jr. sci. co-author, scientific group of Prof. Philip Haitovich

– Pavel, I know that the two objects of your recent research are the brain and the mosquito. Let's start with the brain.

– OK, although about the mosquito, I think the article turned out to be more interesting. As for the brain, our work is not so much about the brain as about alternative splicing. Two words about what it is. Information about the structure of proteins in eukaryotes is encoded in the gene not continuously, but in pieces, exons; the areas between them – introns – must be cut out when copied into RNA. This process is called splicing, and it can happen in different ways - some exons are sometimes skipped. This is an alternative splicing.

– Is this the mechanism that allows one gene to encode several proteins?

– One of the mechanisms. When they began to compare splicing in different species, it turned out that it is more similar in the human brain and liver than in the human and chimpanzee brains, although the brain and liver physiologically have nothing in common. This means that splicing evolves very quickly, even if it differs greatly between close species. But if it is not conservative, then it does not carry a lot of semantic load? We compared the splicing in the cerebral cortex in humans, chimpanzees and macaques from birth and at different ages. Of course, we hoped to see some specific human features, but it turned out that the splicing changes with age in three species occur approximately the same. And it turns out that, despite the rapid overall evolution of splicing, regulated alternative splicing evolves slowly.

– And what interesting things did the mosquito tell you?

– In this work, a completely unexpected result turned out. Our object is an African mosquito, its larva lives in puddles that dry up. The mosquito dries up completely, falling into a state called "anhydrobiosis" – literally "life without water". And then you can soak him, and he lives on quite normally. It turns out that you can "turn off" life in it and start it back. Our Japanese colleagues have read the mosquito genome and found genes that start working when drying out. Analyzing this data, I was looking for an answer to the question of how these genes are activated. And I found that many of them have the same motif of seven nucleotides before the start of the gene. Such a clear signal is a great success. Assuming that this is the binding site of the transcriptional regulator, we started looking for this regulator, and we found it! It turned out to be a heat stress regulator. This is a well-studied and very conservative system. But the mosquito took it and adapted it for another purpose – when drying, not only heat stress occurs, but also a lot of other things. Hundreds of different genes have come under the regulation of this factor – this is an excellent example of the plasticity of regulatory systems.

– Can there be any practical way out of this discovery?

– Well, the way out to practice here is just clear. It would be very interesting to learn how to store cell lines "on the shelf", without freezing, and in the future – whole organs, although this, of course, is still far away.

– Pavel, tell us about your scientific biography.

– I graduated from the MSU FBB, postgraduate studies at MSU, worked under the guidance of Mikhail Gelfand. With the data obtained in Philip Haitovich's alternative splicing group, I worked in China. And when Philip decided to return to Russia, he invited me to the laboratory being created at Skoltech, which is engaged in brain research. Now we have slightly changed the focus of our work to the study of the lipid composition of the brain. This requires mass spectrometers, and we have them.

– Working here, do you feel involved in the global scientific community? Have you ever thought of leaving Russia?

– Well, only if to gain new experience, so as not to work all the time in one place. But now everything suits me. As for engagement, it does not depend on the place of work, it is enough to read, publish and go to conferences.

"We have promoted an incomprehensible protein into a global regulator"

Maria Tutukina, sci. co-author, scientific group of Prof. Michael Gelfand

– Maria, tell us about your research at Skoltech.

– We are investigating various aspects of the regulation of bacterial metabolism in order to understand how the human microbiome can be modified in a targeted way. Moreover, not with the help of antibiotics, which kill both harmful and beneficial bacteria, and not with genetic modification, but with certain additives. We want to find such substances that the right bacteria will be happy to eat and colonize the intestines and at the same time not let harmful bacteria live. It may be some sugars or other food sources. But in order to choose an agent, you first need to understand how bacteria utilize different substrates.

One of our recent articles is that in E. coli, the same gene cassette can ensure the utilization of two sugars at once – lactose and sulfoglucose. Sulfoglucose is a very exotic sugar, a person does not have it, it is found only in plants. Why E. coli should eat sulfoglucose is unclear. We analyzed a cassette of sulfoglucose metabolism genes using comparative genomics methods, and it turned out that it is very similar to a cassette of lactose metabolism genes. Then it was confirmed experimentally. The conclusion follows from this: the gene cassette can be multifunctional, depending on the substrate in the nutrient mixture, switch to one or another pathway of metabolism. And, perhaps, by giving the bacteria a certain additive, we can set this path. In this work, which was done under the guidance of Prof. Mikhail Gelfand, schoolchildren also participated, now students. They used pipettes and test tubes to help experimentally test our hypotheses: they themselves selected primers to study gene expression, isolated RNA themselves.

– And how did you attract schoolchildren to work?

– They were engaged in this work within the framework of the School of Molecular and Theoretical Biology (molbioschool.com ). We have been doing it for six years for high school students who want to try themselves in science. Another work grew out of the School – about a protein regulator, about which it was only known that it somehow participates in the regulation of sugar metabolism genes. But we found out that it affects not only the metabolism of sugars, but also the ability of E. coli cells to move, form colonies and form biofilms. And if we act directly on it, we can influence the ability of bacteria to attach to the walls of the intestine. Now this is a big project that we are doing together with the laboratory of Fyodor Kondrashov in IST Austria. It turned out that there are three forms of this protein encoded by one gene – they bind to different parts of DNA. One form – with sites that regulate the metabolism of sugars (and it is important for colonization by bacteria of the host organism), the other – with sites encoding enzymes and transporters of iron metabolism (this is the first thing that changes during infection). Now we are trying to understand how to specifically modify the action of this protein and thus change something in the microbiome. So we turned a small incomprehensible protein into a global regulator.

– Do you do experimental work on the basis of Skoltech?

– In cooperation with the Institute of Cell Biophysics of the Russian Academy of Sciences. But now we are actively developing an experimental base here. It is very good that we have a sequencer of one of the last generations of Illumina, and now with the help of Maria Logacheva we can quickly and well sequence what we need. In addition, we have been friends with the University of Birmingham (England) for more than eight years, where there are opportunities to work with pathogenic strains and select optimal targeted ligands.

– Maria, what is your scientific biography that led you to Skoltech?

– I graduated from the Biofactory and Chemical Faculty of Voronezh University, postgraduate studies in Pushchino, at the Institute of Cell Biophysics.

– Did you have the opportunity to leave Russia?

– Yes, I still have it. When I finished graduate school, there was a feeling that everything was bad, that there was no science in Russia. But now, having worked in different places, I can say that the creativity of scientists, the organization of science, and the atmosphere in good Russian laboratories are not much worse than in many European countries.

– Well, probably Skoltech is an oasis where young scientists can work in Russia, because their work is paid at a different level than in the institutes of the Russian Academy of Sciences?

– Of course, working with a decent salary makes it possible to think about science, and not about how to survive. Although money is not the main thing.

"We are looking for laboratories and clinics that would like to cooperate with us"

Elena Nabieva, sci. co-author, scientific group of Prof. George Bazykin

– Elena, is your research related to medical issues?

– Yes, we are engaged in the study of spontaneous termination of pregnancy in humans in cases that cannot be explained by known reasons. Known causes include, for example, chromosomal abnormalities. Some trisomies (extra chromosomes in the karyotype) are compatible with life, such as Down syndrome, but most of them are incompatible, so that a significant part of the cases of termination of pregnancy is due to chromosomal abnormalities. But we are interested in other genetic causes, perhaps some point mutations that led to pregnancy loss. These may be unfavorable mutations that both parents had in a recessive state, but if they combine in the child's genome, such a combination may be incompatible with life. There may be more tricky things, for example, not mutations in any one gene, but simply too many harmful mutations, or the fetus has new mutations that the parents did not have.

– Have you managed to understand something yet?

– This project is at the initial stage. Now our main task is to collect a sufficient number of samples. We cooperate with laboratories that analyze such tissues obtained from clinics. We work with those cases where there are no chromosomal abnormalities and they cannot be explained by methods that are widely used in practice. We are sequencing these samples, our colleague Maria Logacheva is doing this, and we are interested in the exome – the part of DNA encoding proteins. At the same time, we need to sequence the DNA of not only unborn children, but also their parents. We have collected a number of such "triples", and some of the material is already at the stage of bioinformatic analysis, but we need more samples.

– Are there similar studies in the world on such a vital issue?

– As far as I know, there are very few such studies in the world, and exactly like ours, they have not been done at all. There were exomic studies in which there was a clear fetal anomaly by ultrasound. But if there is a visible pathology, it narrows down the range of hypotheses and the list of genes that need to be studied. If there is no such information, then it is necessary to search more widely. So our task is more complicated.

– Are you originally a bioinformatician? How did you end up at Skoltech?

– I graduated from Princeton University with a degree in computer science, it's not even really bioinformatics, there's more algorithmics. I have known Georgy Bazykin since graduate school, then I worked with him at Moscow State University, and he invited me to this project.

I must say that we are now actively looking for laboratories and clinics that would like to cooperate with us. Today we are working with three laboratories, but we would like to expand their list.

That asexual reproduction does not always lead to extinction

Olga Vakhrusheva, Jr. sci. co-author, scientific group of Prof. George Bazykin

– Olga, I only know that the subject of your research is rotifers. What are these animals?

– I am engaged in several projects, but let me tell you about bdelloid rotifers. This is the name of a subclass (Bdelloidea), and in general rotifers are a type of multicellular animals. Bdelloid rotifers are interesting primarily because, as it was believed for many years, it is one of the few ancient groups of species that completely abandoned sexual reproduction. This is important in light of discussions about why sexual reproduction is needed at all and why it has become so widespread among eukaryotes. One of the common hypotheses suggests that it allows you to more effectively remove harmful mutations from the population. The proof of this hypothesis is the fact that groups of asexual organisms most often sit on the ends of branches of phylogenetic trees and include a small number of species. Apparently, this means that the transition to asexual reproduction leads to the rapid extinction of the group, and in the short time that this asexual group exists, a large number of species do not have time to arise in it. But there are counterexamples, and one of the most striking is the bdelloid rotifers. It is believed that they abandoned sexual reproduction several tens of millions of years ago, and during this time a lot of species appeared within this group. No one understands why, in this case, the transition to asexual reproduction did not lead to extinction. If it were confirmed that bdelloid rotifers reproduce exclusively asexually, this would be a serious counterargument against the need for sexual reproduction for the long-term success of the species.

– Where do they live?

– These are microscopic invertebrates that live in water, in moss or in soil where there is a lot of water. Scientists looked at several hundred thousand individuals of bdelloid rotifers and among them did not see a single male – only females. And no one has ever observed meiosis in them (this is cell division leading to the formation of germ cells). A few years ago, as part of a large international consortium, we sequenced the first genome of the bdelloid rotifer. The analysis showed that classical meiosis definitely cannot occur in them, since they lack paired chromosomes. Most genes, like ours, are represented as two copies, but these copies are scattered throughout the genome in a mosaic order.

– So these are females reproducing, you can say, by cloning?

– In a sense, yes. But the fact that they do not have classical sexual reproduction does not exclude the possibility that there may be some other way of exchanging genetic material. We sequenced the genomes of 11 rotifers collected in the Moscow region to study how their genetic variability is arranged. If rotifers do not have recombination and exchange of genetic material, then, for example, if mutation A occurred in the context of mutations B and C, these mutations will continue to remain linked. And if recombination occurs, then this coupling will break, and the more often the greater the physical distance between mutations. And we saw exactly such a picture – this coupling is breaking. So, some forms of exchange of genetic material in rotifers occur, although this is not a classic meiosis. Perhaps that is why they have managed to evolve so long and successfully.

– And by your methods, can you find out what is going on with them?

– It's quite difficult. Based on how the rate of uncoupling of mutations depends on the distance between them, you can try to understand what mechanism underlies this.

– Your project belongs to a purely fundamental science. Do you at Skoltech have the opportunity to engage in such research that does not yet have any innovative prospects?

– Yes, it turns out that it is.

"The most interesting people who are engaged in bioinformatics in Russia have gathered here"

Sofya Garushyants, Jr. sci. Co-author, Scientific group of comparative genomics Prof. Michael Gelfand

– Sofia, let's talk about your research with corals.

– In recent years, as a result of global warming in the world, a large number of corals have been affected by various diseases. Populations are simply tragically declining, in the case of the Great Barrier Reef – by at least 30-40%. Only skeletons remain, and the colonies themselves die out. Zoologists from the Department of Invertebrate Zoology of Moscow State University came to our laboratory and brought samples of diseased and healthy coral tissues. We use our own methods to try to understand which organisms cause diseases – we compare a sample of diseased and healthy tissue and see how they differ in bacterial composition.

– Are you sequencing the genome of coral and bacteria?

– We use an approach called metagenomics. We are totally sequencing sections of ribosomal RNA, and such sections that bacteria have and that corals do not. Thus, we get a set of all the bacteria that is in the sample.

This project was preceded by other work. We tested the hypothesis that the change in the appearance of corals (their growth) is associated with small copepod crustaceans that live inside the colony. These crustaceans secrete some substances, and as a result, something like galls is formed on the coral, as on plants, and the crustaceans feed on these growing tissues. We checked whether these growths are associated with a change in bacterial composition. There was an idea that crustaceans transfer some kind of bacteria that cause diseases to corals. Such a connection has been found, but not too obvious.

– Will you tell us about your other works?

– In our bacterial studies, the objects can be very different. At the end of last year, an article was published, in the writing of which we collaborated with medical microbiologists. The paper investigated the causes of Crohn's disease – this is a whole group of diseases that lead to intestinal inflammation. Some of the observed cases are clearly inherited, and some are spontaneous. There was a hypothesis that this disease may be associated with a change in the composition of the intestinal microbiota. In a healthy person, there are about 300 types of bacteria in the intestine, and in Crohn's disease, most of the microbiome is made up of E. coli. We sequenced the genomes of E. coli isolated from patients to understand what features of the genomes may be associated with their accumulation. And they found that in most strains there are special plasmids, which, apparently, provide intestinal seizure in Crohn's disease.

– Sofia, are you originally a bioinformatician? And what brought you to Skoltech?

– I studied at the MSU FBB, but for many years I worked in experimental molecular biology at the Department of Virology. And I was engaged in bioinformatics with Mikhail Gelfand at the Institute of Problems of Information Transmission (IPPI RAS), and he invited me and several other employees to Skoltech.

– Does working at Skoltech give you the opportunity to do science in Russia?

– Obviously, yes. Although there are different models. At an academic institute, you get paid little, but you actually have a permanent position. And here contracts are limited in time, but at the same time the salary is much higher. Although it is more difficult to receive grants. Well, I also like our center, because there is a very strong composition of professors here and it is very interesting to study bioinformatics. It seems to me that the most interesting people who are engaged in bioinformatics in Russia are now gathered here.

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