Mitochondria contain circular DNA. Research on mitochondrial DNA. Properties and functions of DNA

Why do mitochondria need their own DNA? Although why shouldn't symbionts have their own DNA in them, producing everything they need on the spot? Why then transfer part of mitochondrial DNA into the cell nucleus, creating the need to transport gene products into mitochondria? Why are mitochondria passed down from only one parent? How do the mitochondria obtained from the mother get along with the cell's genome, made up of the DNA of the mother and father? The more people learn about mitochondria, the more questions arise.

However, this applies not only to mitochondria: in any area of ​​any science, the expansion of the sphere of knowledge only leads to an increase in its surface in contact with the unknown, causing more and more questions, the answers to which will expand the same sphere with the same predictable result.

So, the DNA of modern mitochondria is distributed very strangely: a small part of genes is contained directly in mitochondria on the circular chromosome (more precisely, in several copies of the same chromosome in each mitochondrion), and most of the blueprints for the production of mitochondrial constituents are stored in the cell nucleus. Therefore, the copying of these genes occurs simultaneously with the copying of the genome of the whole organism, and the products produced by them go a long way from the cytoplasm of the cell into the mitochondria. Nevertheless, it is in many ways convenient: the mitochondria is relieved of the need to copy all these genes during reproduction, to read them and build proteins and other components, focusing on its main function of producing energy. Why, then, is there a small DNA in mitochondria, for the maintenance of which all these mechanisms are required, getting rid of which mitochondria could throw even more resources on the main purpose of their existence?

At first, it was assumed that the DNA remaining in the mitochondria is an atavism, the legacy of a pro-mitochondria absorbed by methanogen, which has a complete bacterial genome. At the beginning of their symbiosis, despite the existence in the nucleus of those mitochondrial genes ( m-genes), which were necessary to maintain a comfortable environment for pro-mitochondria inside the methanogen (this is described in detail in the section on mitochondria), the same genes were stored in each of the mitochondria. The pro-mitochondria at the beginning of its life as a symbiont looked about the same as the modern bacterium in the diagram to the left of this paragraph.

And very slowly, due to lack of demand, these genes disappeared from the mitochondrial chromosome as a result of a variety of mutations. But the cell nucleus accumulated more and more m-genes that entered the cytoplasm from the destroyed symbionts-mitochondria and integrated into the genome of the eukaryotic chimera. As soon as the newly built m-gene began to be read, the cellular mechanisms produced the products necessary for mitochondria, freeing the symbionts from their independent creation. This means that the mitochondrial analogue of the gene that passed into the nucleus was no longer maintained in working order by natural selection and was erased by mutations in the same way as all the previous ones. Therefore, it would be logical to assume that soon those genes that are still in the mitochondria will pass into the nucleus, which will lead to a great energy benefit for eukaryotes: after all, cumbersome mechanisms of copying, reading and correcting DNA can be removed from each mitochondrion, and so everything you need to create proteins.

Having come to this conclusion, the scientists calculated how long it took for all genes to migrate through natural drift from the mitochondria to the nucleus. And it turned out that this period has long passed. At the time of the appearance of the eukaryotic cell, mitochondria had a common bacterial genome of several thousand genes (scientists establish what this genome was by studying the m-genes transferred to the nucleus in different organisms), and now mitochondria of all types of eukaryotes have lost from 95 to 99.9% of their genes. No one has left more than a hundred genes in mitochondria, but no one has a geneless mitochondria either. If chance played a key role in this process, then at least a few species would have already gone through the path of gene transfer to the nucleus to the end. But this did not happen, and the mitochondria of different species studied at the moment, losing their genes independently of each other, retained the same set of them, which directly indicates the need for the presence of precisely these genes in mitochondria.

Moreover, other energy-producing organelles of cells, chloroplasts, also have their own DNA, and in the same way chloroplasts of different species evolved in parallel and independently, each with the same set of genes.

This means that all those significant inconveniences in maintaining its own genome in each cell mitochondrion (and on average, one cell contains several hundred!) And the bulky apparatus for copying-correcting-translating it (the main, but not all! ) are outweighed by something.

And at the moment there is a consistent theory of this "something": the ability to produce certain parts of the mitochondrion directly inside it is necessary to regulate the respiration rate and adjust the processes occurring in the mitochondria to the every minute changing needs of the whole organism.

Imagine that one of the hundreds of mitochondria of a cell suddenly lacks the elements of the respiratory chain (see details about it in), or there are not enough ATP synthases in it. It turns out to be either overloaded with food and oxygen and cannot process them quickly enough, or its intermembrane space is bursting with protons that have nowhere to go - a complete disaster in general. Of course, all these deviations from the ideal life situation trigger multiple signals aimed at leveling the roll of the sinking ship.

These signals trigger the production of exactly those parts that are missing in the mitochondria at the moment, activating the reading of the genes by which proteins are built. As soon as the mitochondrion has enough components of the respiratory chain or ATPases, the "roll will even out", the signals about the need to build new parts will cease to come, and the genes will be turned off again. This is one of the surprisingly elegant in its simplicity necessary mechanisms of self-regulation of the cell, the slightest violation of it leads to a serious illness or even non-viability of the organism.

Let's try to logically determine where the genes necessary to respond to this distress signal should be located. Imagine the situation that these genes are located in the nucleus of a cell containing a couple of hundred mitochondria. In one of the mitochondria, for example, a lack of NADH dehydrogenase: the first enzyme in the respiratory chain, whose role is to strip two electrons from the NADH molecule, transfer them to the next enzyme, and pump 2-4 protons across the membrane.

In fact, such deficiencies of any enzyme occur quite often, because they periodically fail, the amount of food consumed is constantly changing, the needs of the cell for ATP also jump after the jumps or felting of the body containing this cell. Therefore, the situation is very typical. And so the mitochondrion emits a signal: "We need to build more NADH dehydrogenase!" By cellular standards, the transit time of this signal is very significant, and in fact it is also necessary to pull the constructed messenger RNA from the nucleus into the cytoplasm, create proteins from it, send them to the mitochondria ...

And this is where a problem arises that is much more significant than wasting extra time: when creating specialized mitochondrial proteins, they are marked with a signal "deliver to the mitochondria", but which one? Unknown. Therefore, proteins that they do not need begin to enter each of a couple of hundred mitochondria. The cell spends resources on their production and delivery, mitochondria are filled with extra respiratory chains (which leads to inefficiency of the respiratory processes), and the only mitochondria that needs these proteins does not receive them in sufficient quantities, because it gets, at best, a hundredth of the produced. Therefore, she continues to send out distress signals and the chaos continues. Even from this lyrical and superficial description of what is happening, it is clear that such a cell is not viable. And that there are genes that must be read and translated directly into the mitochondria in order to regulate the processes occurring in it, and not rely on the plan for the production of nails launched by the nucleus party ... that is, the proteins of the respiratory chain for all mitochondria at once.

After checking what exactly is produced by the different organisms remaining in the mitochondria (and therefore moving m-genes into the nucleus independently of each other), we found that these are the elements for the construction of respiratory chains and ATPase, as well as ribosomes (that is, the main part broadcasting apparatus).

You can read more about this (and not only) from Lane in "Energy, Sex, Suicide: Mitochondria and the Meaning of Life"... Well, you can simply compare the mitochondrial DNA diagram, where the encoded products are decoded (to the right of this paragraph), with the respiratory chain diagram (above), so that it becomes clear what exactly is produced in the mitochondria. Of course, not every protein inserted into this chain is produced locally; some of them are built in the cytoplasm of the cell. But the main “anchors” that the rest of the parts cling to are created inside the mitochondria. That allows you to produce exactly as many enzymes as needed, and exactly where they are needed.

How mitochondria are associated with sex and how different genomes coexist in one cell, I will write in one of the next chapters of this line.

Introduction

A quarter of a century passed since the discovery of DNA molecules in mitochondria before molecular biologists and cytologists became interested in them, as well as geneticists, evolutionists, as well as paleontologists and forensic scientists. Such a wide interest was provoked by the work of A. Wilson of the University of California. In 1987, he published the results of a comparative analysis of the DNA of mitochondria taken from 147 representatives of different ethnic groups of all human races inhabiting five continents. By the type, location and number of individual mutations, it was established that all mitochondrial DNA arose from one ancestral nucleotide sequence by divergence. In the pseudo-scientific press, this conclusion was interpreted in an extremely simplified way - all of humanity descended from one woman named mitochondrial Eve (since both daughters and sons receive mitochondria only from their mother), who lived in Northeast Africa about 200 thousand years ago ... After another 10 years, it was possible to decipher a fragment of mitochondrial DNA isolated from the Neanderthal wasps, and to estimate the lifetime of the last common ancestor of man and Neanderthal at 500 thousand years ago.

Today, human mitochondrial genetics is intensively developing both in the population and in the medical aspect. A relationship has been established between a number of severe hereditary diseases and defects in mitochondrial DNA. The genetic changes associated with aging are most pronounced in mitochondria. What is the mitochondrial genome, which differs in humans and other animals from that in plants, fungi and protozoa in size, shape, and genetic capacity? What is the role, how does it work and how did the mitochondrial genome arise in different taxa in general and in humans in particular? This is what will be discussed in my "smallest and most modest" essay.

In addition to DNA, the mitochondrial matrix contains its own ribosomes, which differ in many characteristics from the eukaryotic ribosomes located on the membranes of the endoplasmic reticulum. However, on the ribosomes of mitochondria, no more than 5% of all proteins that make up their composition are formed. Most of the proteins that make up the structural and functional components of mitochondria are encoded by the nuclear genome, synthesized on the ribosomes of the endoplasmic reticulum, and transported through its channels to the assembly site. Thus, mitochondria are the result of the combined efforts of two genomes and two transcription and translation machines. Some subunit enzymes of the mitochondrial respiratory chain consist of different polypeptides, some of which are encoded by the nuclear and some by the mitochondrial genome. For example, the key enzyme of oxidative phosphorylation, cytochrome c oxidase in yeast, consists of three subunits encoded and synthesized in mitochondria, and four encoded in the cell nucleus and synthesized in the cytoplasm. The expression of most mitochondrial genes is controlled by specific genes in the nucleus.

Symbiotic theory of the origin of mitochondria

The hypothesis about the origin of mitochondria and plant plastids from intracellular bacteria-endosymbionts was expressed by R. Altman back in 1890. During the century of rapid development of biochemistry, cytology, genetics and molecular biology that appeared half a century ago, the hypothesis grew into a theory based on a large amount of factual material ... Its essence is as follows: with the appearance of photosynthesizing bacteria, oxygen accumulated in the Earth's atmosphere - a by-product of their metabolism. With an increase in its concentration, the life of anaerobic heterotrophs became more difficult, and some of them, to obtain energy, passed from anoxic fermentation to oxidative phosphorylation. Such aerobic heterotrophs could, with a higher efficiency than anaerobic bacteria, break down organic matter formed as a result of photosynthesis. Some of the free-living aerobes were captured by anaerobes, but not “digested”, but preserved as energy stations, mitochondria. Mitochondria should not be viewed as slaves taken prisoner to supply cells with ATP molecules that are unable to breathe. Rather, they are “creatures” who, back in the Proterozoic, found the best of shelters for themselves and their offspring, where you can spend the least effort without risking being eaten.

Numerous facts speak in favor of the symbiotic theory:

The sizes and shapes of mitochondria and free-living aerobic bacteria are the same; both contain circular DNA molecules that are not associated with histones (as opposed to linear nuclear DNA);

In terms of nucleotide sequences, ribosomal and transport RNAs of mitochondria differ from nuclear ones, while demonstrating surprising similarity with analogous molecules of some aerobic gram-negative eubacteria;

Mitochondrial RNA polymerases, although encoded in the cell nucleus, are inhibited by rifampicin, like bacterial ones, and eukaryotic RNA polymerases are insensitive to this antibiotic;

Protein synthesis in mitochondria and bacteria is suppressed by the same antibiotics that do not affect the ribosomes of eukaryotes;

The lipid composition of the inner membrane of mitochondria and bacterial plasmalemma is similar, but very different from that of the outer membrane of mitochondria, which is homologous to other membranes of eukaryotic cells;

Cristae, formed by the inner mitochondrial membrane, are evolutionary analogs of the mesosomal membranes of many prokaryotes;

Until now, organisms have survived that mimic intermediate forms on the path to the formation of mitochondria from bacteria (primitive amoeba Pelomyxa does not have mitochondria, but always contains endosymbiotic bacteria).

There is an idea that different kingdoms of eukaryotes had different ancestors and endosymbiosis of bacteria arose at different stages of the evolution of living organisms. This is also evidenced by the differences in the structure of mitochondrial genomes of protozoa, fungi, plants and higher animals. But in all cases, the main part of the genes from the promitochondria entered the nucleus, possibly with the help of mobile genetic elements. When a part of the genome of one of the symbionts is included in the genome of the other, the integration of the symbionts becomes irreversible. The new genome can create metabolic pathways leading to the formation of useful products that cannot be synthesized by any of the partners individually. Thus, the synthesis of steroid hormones by cells of the adrenal cortex is a complex chain of reactions, some of which occur in the mitochondria, and some in the endoplasmic reticulum. By capturing the genes of the promitochondria, the nucleus was able to reliably control the functions of the symbiont. The nucleus encodes all proteins and lipid synthesis of the outer membrane of mitochondria, most of the proteins of the matrix and the inner membrane of organelles. Most importantly, the nucleus encodes the enzymes of replication, transcription, and translation of mtDNA, thereby controlling the growth and reproduction of mitochondria. The growth rate of symbiotic partners should be approximately the same. If the host grows faster, then with each generation the number of symbionts per individual will decrease, and, in the end, descendants without mitochondria will appear. We know that in every cell of an organism that reproduces sexually, there are many mitochondria, which replicate their DNA in the interval between divisions of the host. This ensures that each of the daughter cells will receive at least one copy of the mitochondrial genome.

The role of the cell nucleus in mitochondrial biogenesis

A certain type of mutant yeast has an extensive deletion in mitochondrial DNA, which leads to a complete cessation of protein synthesis in mitochondria; as a result, these organelles are unable to perform their function. Since, when growing on a medium with a low glucose content, such mutants form small colonies, they are called cytoplasmic mutantamipetite.

Although petite mutants do not have mitochondrial protein synthesis and therefore do not form normal mitochondria, such mutants nevertheless contain promitochondria, which, to a certain extent, are similar to ordinary mitochondria, have a normal outer membrane and an inner membrane with poorly developed cristae. In the promitochondria there are many enzymes encoded by nuclear genes and synthesized on the ribosomes of the cytoplasm, including DNA and RNA polymerases, all enzymes of the citric acid cycle and many proteins that make up the inner membrane. This clearly demonstrates the predominant role of the nuclear genome in mitochondrial biogenesis.

It is interesting to note that although the lost DNA fragments account for 20 to more than 99.9% of the mitochondrial genome, the total amount of mitochondrial DNA in the petite mutants always remains at the same level as in the wild type. This is due to the still poorly studied process of DNA amplification, as a result of which a DNA molecule is formed, consisting of tandem repeats of the same region and equal in size to a normal molecule. For example, mitochondrial DNA of the petite mutant, which retains 50% of the nucleotide sequence of wild-type DNA, will consist of two repeats, while a molecule that retains only 0,1% the wild-type genome will be built from 1000 copies of the remaining fragment. Thus, petite mutants can be used to obtain in a large number of certain sections of mitochondrial DNA, which, one might say, are cloned by nature itself.

Although the biogenesis of organelles is controlled mainly by nuclear genes, the organelles themselves, judging by some data, have some kind of regulatory influence on the principle of feedback; in any case, this is the case with mitochondria. If you block protein synthesis in the mitochondria of intact cells, then enzymes involved in mitochondrial synthesis of DNA, RNA and proteins begin to form in excess in the cytoplasm, as if the cell is trying to overcome the effect of a blocking agent. But, although the existence of some kind of signal from the mitochondria is beyond doubt, its nature is still not known.

For a number of reasons, the mechanisms of mitochondrial biogenesis are now studied in most cases in cultures Saccharomyces carlsbergensis(brewer's yeast and S. cerevisiae(baker's yeast). First, when growing on glucose, these yeasts exhibit a unique ability to exist only through glycolysis, that is, to do without mitochondrial function. This makes it possible to study mutations in mitochondrial and nuclear DNA that prevent the development of these organelles. Such mutations are lethal in almost all other organisms. Secondly, yeast - simple unicellular eukaryotes - is easy to cultivate and biochemically test. Finally, yeast can multiply in both the haploid and diploid phases, usually by asexual budding process (asymmetric mitosis). But yeast also has a sexual process: from time to time, two haploid cells merge, forming a diploid zygote, which then either divides by mitosis or undergoes meiosis and again gives haploid cells. By controlling the alternation of asexual and sexual reproduction during the experiment, you can learn a lot about the genes responsible for the function of mitochondria. Using these methods, it is possible, in particular, to find out whether such genes are localized in nuclear or mitochondrial DNA, since mutations of mitochondrial genes are not inherited according to Mendel's laws, which govern the inheritance of nuclear genes.

Mitochondrial transport systems

Most of the proteins contained in mitochondria and chloroplasts are imported into these organelles from the cytosol. This raises two questions: how does the cell direct proteins to the proper organelle, and how do these proteins enter it?

A partial answer was obtained when studying the transport of the small subunit (S) of the enzyme into the chloroplast stroma ribulose-1,5-bisphosphate-carboxylazy. If mRNA isolated from the cytoplasm of a unicellular alga Chlamydomonas or from pea leaves, introduced as a matrix into a protein-synthesizing system in vitro, then one of the many proteins formed will be bound by a specific anti-S-antibody. The S-protein synthesized in vitro is called pro-S, since it is larger than the regular S-protein by about 50 amino acid residues. When the pro-S protein is incubated with intact chloroplasts, it penetrates the organelles and is converted there by peptidase into the S-protein. Then the S-protein binds to the large subunit of ribulose-1,5-bisphosphate carboxylase, synthesized on the ribosomes of the chloroplast, and forms an active enzyme with it in the stroma of the chloroplast.

The mechanism of S-protein transfer is unknown. It is believed that pro-S binds to a receptor protein located on the outer membrane of the chloroplast or at the point of contact between the outer and inner membranes, and is then transported to the stroma through transmembrane channels as a result of an energy-intensive process.

Proteins are transported into mitochondria in a similar way. If purified yeast mitochondria are incubated with a cell extract containing newly synthesized radioactive yeast proteins, it can be observed that mitochondrial proteins encoded by the nuclear genome are separated from non-mitochondrial proteins of the cytoplasm and are selectively incorporated into mitochondria, just as they do in an intact cell. In this case, proteins of the outer and inner membranes, matrix and intermembrane space find their way to the corresponding mitochondrial compartment.

Many of the newly synthesized proteins intended for the inner membrane, matrix, and intermembrane space have a leader peptide at their N-terminus, which is cleaved by a specific protease in the matrix during transportation. The transfer of proteins to these three mitochondrial compartments requires the energy of an electrochemical proton gradient created across the inner membrane. The mechanism of protein transfer for the outer membrane is different: in this case, neither energy expenditure nor proteolytic cleavage of a longer precursor protein is required. These and other observations suggest that all four groups of mitochondrial proteins are transported into the organelle using the following mechanism: it is assumed that all proteins, except for those intended for the outer membrane, are included in the inner membrane as a result of a process that requires energy consumption and occurs in contact points of the outer and inner membranes. Apparently, after this initial incorporation of the protein into the membrane, it undergoes proteolytic cleavage, which leads to a change in its conformation; depending on how the conformation changes, the protein is either fixed in the membrane or "pushed" into the matrix or into the intermembrane space.

The transfer of proteins through the membranes of mitochondria and chloroplasts is, in principle, analogous to their transfer through the membranes of the endoplasmic reticulum. However, there are several important differences here. First, when transported to the matrix or stroma, the protein passes through both the outer and inner membranes of the organelle, while when transported into the lumen of the endoplasmic reticulum, the molecules pass through only one membrane. In addition, the transfer of proteins into the reticulum is carried out using the mechanism directional removal(vectorial discharge) - it begins when the protein has not yet completely left the ribosome (co-translational import), and the transfer to mitochondria and chloroplasts occurs after the synthesis of the protein molecule is completely completed (post-translational import).

Despite these differences, in both cases, the cell synthesizes precursor proteins containing a signal sequence that determines which membrane a given protein is directed to. Apparently, in many cases, this sequence is cleaved from the precursor molecule after the completion of the transport process. However, some proteins are immediately synthesized in their final form. It is believed that in such cases the signal sequence is contained within the polypeptide chain of the final protein. Signal sequences are still poorly understood, but there must probably be several types of such sequences, each of which determines the transfer of a protein molecule to a specific region of the cell. For example, in a plant cell, some of the proteins, the synthesis of which begins in the cytosol, are then transported to mitochondria, others to chloroplasts, still others to peroxisomes, and still others to the endoplasmic reticulum. The complex processes leading to the correct intracellular distribution of proteins are only now becoming understood.

In addition to nucleic acids and proteins, lipids are needed to build new mitochondria. Unlike chloroplasts, mitochondria receive most of their lipids from the outside. In animal cells, phospholipids synthesized in the endoplasmic reticulum are transported to the outer membrane of mitochondria with the help of special proteins, and then incorporated into the inner membrane; this is believed to occur at the point of contact between the two membranes. The main reaction of lipid biosynthesis, catalyzed by the mitochondria themselves, is the conversion of phosphatidic acid into the phospholipid cardiolipin, which is found mainly in the inner mitochondrial membrane and makes up about 20% of all its lipids.

Size and shape of mitochondrial genomes

To date, more than 100 different mitochondrial genomes have been read. The set and the number of their genes in mitochondrial DNA, for which the nucleotide sequence is fully determined, differ greatly in different species of animals, plants, fungi and protozoa. The largest number of genes found in the mitochondrial genome of the flagellate protozoan Rectinomo-nas americana- 97 genes, including all protein-coding genes found in the mtDNA of other organisms. In most higher animals, the mitochondrial genome contains 37 genes: 13 for the proteins of the respiratory chain, 22 for tRNA, and two for rRNA (for the large ribosome subunit 16S rRNA and for the small 12S rRNA). In plants and protozoa, unlike animals and most fungi, some proteins that make up the ribosomes of these organelles are encoded in the mitochondrial genome. Key enzymes of template polynucleotide synthesis, such as DNA polymerase (replicating mitochondrial DNA) and RNA polymerase (transcribing the mitochondrial genome), are encoded in the nucleus and synthesized on the cytoplasmic ribosomes. This fact indicates the relativity of mitochondrial autonomy in the complex hierarchy of the eukaryotic cell.

The genomes of mitochondria of different species differ not only in the set of genes, the order of their location and expression, but in the size and shape of DNA. The overwhelming majority of the mitochondrial genomes described today are circular supercoiled double-stranded DNA molecules. In some plants, along with circular forms, there are linear ones, and in some protozoa, for example, ciliates, only linear DNA is found in mitochondria.

As a rule, each mitochondrion contains several copies of its genome. So, in the cells of the human liver there are about 2 thousand mitochondria, and in each of them there are 10 identical genomes. In mouse fibroblasts there are 500 mitochondria containing two genomes, and in yeast cells S. cerevisiae- up to 22 mitochondria with four genomes.

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Fig 2. Scheme of formation of linear (A), circular (B), chain (C) mtDNA oligomers. ori - region of the beginning of DNA replication.

The genome size of mitochondria of different organisms ranges from less than 6 thousand base pairs in the malaria plasmodium (in addition to two rRNA genes, it contains only three genes encoding proteins) to hundreds of thousands of nucleotide pairs in terrestrial plants (for example, in Arabidopsis thaliana from the cruciferous family 366924 base pairs). At the same time, 7-8-fold differences in the size of mtDNA of higher plants are found even within the same family. The length of mtDNA of vertebrates differs slightly: in humans - 16,569 base pairs, in a pig - 16350, in a dolphin - 16330, in a clawed frog Xenopus laevis- 17533, in carp - 16400. These genomes are also similar in the localization of genes, most of which are located end-to-end; in some cases they even overlap, usually by one nucleotide, so that the last nucleotide of one gene is the first in the next. Unlike vertebrates, in plants, fungi, and protozoa, mtDNAs contain up to 80% of non-coding sequences. The order of genes in mitochondrial genomes differs from species to species.

A high concentration of reactive oxygen species in mitochondria and a weak repair system increase the frequency of mtDNA mutations in comparison with the nuclear one by an order of magnitude. Oxygen radicals cause specific substitutions of C®T (deamination of cytosine) and GT®T (oxidative damage to guanine), as a result of which, possibly, mtDNA is rich in AT-pairs. In addition, all mtDNAs have an interesting property - they are not methylated, unlike nuclear and prokaryotic DNA. It is known that methylation (temporary chemical modification of the nucleotide sequence without disrupting the coding function of DNA) is one of the mechanisms of programmed gene inactivation.

The size and structure of DNA molecules in organelles

The relative amount of DNA organelles in some cells and tissues

Functioning of the mitochondrial genome

What is special about the mechanisms of replication and transcription of mammalian mitochondrial DNA?

Complementary "href =" / text / category / komplementarij / "rel =" bookmark "> complementary strands in mtDNA differ significantly in specific gravity, since they contain different amounts of“ heavy ”purine and“ light ”pyrimidine nucleotides. - H (heavy - heavy) and L (light - light) chain. At the beginning of the replication of the mtDNA molecule, the so-called D-loop (from the English Displace-ment loop) is formed. This structure, visible in the electronic micro- osp, consists of a double-stranded and single-stranded (retracted part of the H-chain) sections. the end of the ribonucleotide seed, which corresponds to the point of initiation of the synthesis of the H-chain (oriH). Synthesis of the L-chain begins only when the daughter H-chain reaches the point ori L. This is due to the fact that the region of initiation of replication of the L-chain is accessible for DNA synthesis enzymes only in a single-stranded state, and, consequently, only in an untwisted double helix in the synthesis of the H-chain. Thus, mtDNA daughter strands are synthesized continuously and asynchronously (Fig. 3).

Fig 3. Mammalian mtDNA replication scheme. First, the D-loop is formed, then the daughter H-chain is synthesized, then the synthesis of the daughter L-chain begins.

End of the 16S rRNA gene (Fig. 4). There are 10 times more such short transcripts than long ones. As a result of maturation (processing), 12S rRNA and 16S rRNA are formed from them, which are involved in the formation of mitochondrial ribosomes, as well as phenylalanine and valine tRNA. The remaining tRNAs are excised from the long transcripts and translated mRNAs are formed, to the 3 "ends of which polyadenyl sequences are attached. The 5" ends of these mRNAs are not capped, which is unusual for eukaryotes. No splicing (splicing) occurs, since none of the mammalian mitochondrial genes contain introns.

Fig 4. Transcription of human mtDNA containing 37 genes. All transcripts begin to be synthesized in the ori H region. Ribosomal RNAs are excised from the long and short H chain transcripts. tRNA and mRNA are formed as a result of processing from transcripts of both DNA strands. The tRNA genes are shown in light green.

Do you want to know what other surprises the mitochondrial genome can present? Fine! Read on! ..

The leader and 3'-noncoding regions, like most nuclear mRNAs. A number of genes also contain introns. Thus, in the box gene encoding cytochrome oxidase b, there are two introns. From the primary RNA transcript autocatalytically (without the participation of any or proteins) a copy of most of the first intron is cut out. The remaining RNA serves as a template for the formation of the enzyme maturase, which is involved in splicing. Part of its amino acid sequence is encoded in the remaining copies of introns. Maturase cuts them out, destroying its own mRNA, copies of exons are stitched together, and mRNA for cytochrome oxidase b is formed (Fig. 5.) The discovery of such a phenomenon forced us to reconsider the concept of introns as “nothing coding sequences”.

Fig 5. Processing (maturation) of cytochrome oxidase b mRNA in yeast mitochondria. At the first stage of splicing, mRNA is formed, by which maturase is synthesized, which is necessary for the second stage of splicing.

When studying the expression of mitochondrial genes Trypanosoma brucei discovered a surprising deviation from one of the basic axioms of molecular biology, which states that the sequence of nucleotides in mRNA exactly matches that in the coding regions of DNA. It turned out that the mRNA of one of the subunits of cytochrome c oxidase is being edited, that is, after transcription, its primary structure changes - four uracils are inserted. As a result, a new mRNA is formed, which serves as a template for the synthesis of an additional subunit of the enzyme, the sequence of amino acids in which has nothing to do with the sequence. Virus "href =" / text / category / virus / "rel =" bookmark "> viruses, fungi, The English researcher Burrell compared the structure of one of the calf mitochondrial genes with the sequence of amino acids in the cytochrome oxidase subunit encoded by this gene. it is “ideal”, that is, it obeys the following rule: “if two codons have two identical nucleotides, and the third nucleotides belong to the same class (purine - A, G, or pyrimidine - Y, C), then they encode the same amino acid. ”There are two exceptions to this rule in the universal code: the AUA triplet encodes isoleucine, and the AUG codon encodes methionine, while in the ideal mitochondrial code, both of these triplets to oder methionine; the UGG triplet encodes only tryptophan, and the UGA triplet encodes a stop codon. In the universal code, both deviations relate to the fundamental moments of protein synthesis: the AUG codon is the initiator, and the UGA stop codon stops polypeptide synthesis. The ideal code is not inherent in all mitochondria described, but none of them has a universal code. It can be said that mitochondria speak different languages, but never speak the language of the nucleus.

Differences between the “universal” genetic code and the two mitochondrial codes

As already mentioned, there are 22 tRNA genes in the mitochondrial genome of vertebrates. How, then, does such an incomplete set serve all 60 codons for amino acids (in an ideal code of 64 triplets, there are four stop codons, in a universal one - three)? The fact is that during protein synthesis in mitochondria, codon-anticodon interactions are simplified - two of the three anticodon nucleotides are used for recognition. Thus, one tRNA recognizes all four members of the codon family, which differ only in the third nucleotide. For example, leucine tRNA with the GAU anticodon stands on the ribosome on-against the codons CUU, CUC, CUA and CUG, ensuring the error-free incorporation of leucine into the polypeptide chain. The other two leucine codons, UUA and UUG, are recognized by tRNA with the anticodon AAU. In total, eight different tRNA molecules recognize eight families of four codons each, and 14 tRNAs recognize different pairs of codons, each of which encodes one amino acid.

It is important that the aminoacyl tRNA synthetase enzymes responsible for the attachment of amino acids to the corresponding tRNA of mitochondria are encoded in the cell nucleus and synthesized on the ribosomes of the endoplasmic reticulum. Thus, in vertebrates, all protein components of mitochondrial polypeptide synthesis are encoded in the nucleus. At the same time, protein synthesis in mitochondria is not suppressed by cycloheximide, which blocks the work of eukaryotic ribosomes, but is sensitive to antibiotics erythromycin and chloramphenicol, which inhibit protein synthesis in bacteria. This fact serves as one of the arguments in favor of the origin of mitochondria from aerobic bacteria during the symbiotic formation of eukaryotic cells.

The importance of having your own genetic system for mitochondria

Why do mitochondria need their own genetic system, while other organelles, such as peroxisomes and lysosomes, do not have it? This question is not at all trivial, since maintaining a separate genetic system is expensive for the cell, given the required number of additional genes in the nuclear genome. Here ribosomal proteins, aminoacyl tRNA synthetases, DNA and RNA polymerases, RNA processing and modification enzymes, etc. should be encoded. Most of the studied proteins from mitochondria differ in amino acid sequence from their analogs from other parts of the cell, and there is reason to believe that there are very few proteins in these organs that could be found elsewhere. This means that just to maintain the genetic system of mitochondria, the nuclear genome must have several tens of additional genes. The reasons for this “waste” are unclear, and the hope that a clue will be found in the nucleotide sequence of mitochondrial DNA was not justified. It is difficult to imagine why proteins formed in mitochondria must be synthesized there, and not in the cytosol.

Usually, the existence of a genetic system in energy organelles is explained by the fact that some of the proteins synthesized inside the organelle are too hydrophobic to pass through the mitochondrial membrane from the outside. However, the study of the ATP-synthetase complex showed that such an explanation is implausible. Although individual protein subunits of ATP synthetase are highly conserved in the course of evolution, the sites of their synthesis change. In chloroplasts, several rather hydrophilic proteins, including four of the five subunits of the F1-ATPase part of the complex, are formed on ribosomes within the organelle. On the contrary, at the mushroom Neurospora and in animal cells, a very hydrophobic component (subunit 9) of the membrane part of ATPase is synthesized on the cytoplasmic ribosomes and only after that passes into the organelle. The different localization of genes encoding subunits of functionally equivalent proteins in different organisms is difficult to explain with the help of any hypothesis postulating certain evolutionary advantages of modern genetic systems of mitochondria and chloroplasts.

Given all of the above, it remains only to assume that the genetic system of mitochondria represents an evolutionary dead end. Within the framework of the endo-symbiotic hypothesis, this means that the process of transfer of endosymbiont genes into the host's nuclear genome stopped before it was completely completed.

Cytoplasmic inheritance

The consequences of cytoplasmic gene transfer for some animals, including humans, are more serious than for yeast. Two merging haploid yeast cells have the same size and contribute the same amount of mitochondrial DNA to the resulting zygote. Thus, in yeast, the mitochondrial genome is inherited from both parents, which make an equal contribution to the gene pool of the offspring (although, after several generations separate offspring will often contain mitochondria of only one of the parental types). In contrast to this, in higher animals, the egg cell introduces more cytoplasm into the zygote than sperm, and in some animals, sperm may not add cytoplasm at all. Therefore, one can think that in higher animals the mitochondrial genome will be transmitted only from one parent (namely, by maternal lines); indeed, this has been confirmed by experiments. It turned out, for example, that when rats of two laboratory lines are crossed with mitochondrial DNA slightly different in the sequence of nucleotides (types A and B), offspring is obtained containing

containing mitochondrial DNA only of the maternal type.

Cytoplasmic heredity, unlike nuclear heredity, does not obey Mendel's laws. This is due to the fact that in higher animals and plants gametes from different sexes contain incomparable amounts of mitochondria. So, in the mouse egg there are 90 thousand mitochondria, and in the sperm - only four. Obviously, in a fertilized egg, mitochondria are predominantly or only from a female, i.e., the inheritance of all mitochondrial genes is maternal. Genetic analysis of cytoplasmic heredity is difficult due to nuclear-cytoplasmic interactions. In the case of cytoplasmic male sterility, the mutant mitochondrial genome interacts with certain nuclear genes, the recessive alleles of which are necessary for the development of the trait. The dominant alleles of these genes, both in homo- and heterozygous states, restore plant fertility regardless of the state of the mitochondrial genome.

I would like to dwell on the mechanism of maternal gene inheritance by giving a specific example. In order to finally and irrevocably understand the mechanism of non-Mendelian (cytoplasmic) inheritance of mitochondrial genes, let us consider what happens to such genes when two haploid cells fuse to form a diploid zygote. In the case when one yeast cell carries a mutation that determines the resistance of mitochondrial protein synthesis to chloramphenicol, and the other, a wild-type cell, is sensitive to this antibiotic: mutant genes can be easily identified by growing yeast on a medium with glycerol, which can only be used by cells with intact mitochondria; therefore, in the presence of chloramphenicol, only cells carrying the mutant gene can grow on such a medium. Our diploid zygote will initially have both mutant and wild-type mitochondria. As a result of mitosis, a diploid daughter cell will sprout from the zygote, which will contain only a small number of mitochondria. After several mitotic cycles, eventually one of the new cells will receive all mitochondria, either mutant or wild type. Therefore, all the offspring of such a cell will have genetically identical mitochondria. Such a random process, which results in the formation of diploid offspring containing mitochondria of only one type, is called mitoticth beholdgreeceth. When a diploid cell with only one type of mitochondria undergoes meiosis, all four daughter haploid cells receive the same mitochondrial genes. This type of inheritance is called nemendea lion skim or cytoplasmic in contrast to Mendelian inheritance of nuclear genes. The transfer of genes by the cytoplasmic type means that the genes under study are located in the mitochondria.

The study of mitochondrial genomes, their evolution, proceeding according to the specific laws of population genetics, the relationship between nuclear and mitochondrial genetic systems, is necessary to understand the complex hierarchical organization of the eukaryotic cell and the organism as a whole.

Certain hereditary diseases and human aging are associated with certain mutations in mitochondrial DNA or in nuclear genes that control mitochondrial function. Data are accumulating on the participation of mtDNA defects in carcinogenesis. Therefore, mitochondria may be a target for cancer chemotherapy. There are facts about the close interaction of the nuclear and mitochondrial genomes in the development of a number of human pathologies. Multiple deletions of mtDNA were found in patients with severe muscle weakness, ataxia, deafness, mental retardation, inherited in an autosomal dominant manner. Established sexual dimorphism in the clinical manifestations of coronary heart disease, which is most likely due to the maternal effect - cytoplasmic heredity. The development of gene therapy gives hope for the correction of defects in the genomes of mitochondria in the foreseeable future.

As you know, in order to check the function of one of the components of a multicomponent system, it becomes necessary to eliminate this component, followed by an analysis of the changes that have occurred. Since the topic of this abstract is an indication of the role of the maternal genome for the development of the offspring, it would be logical to learn about the consequences of violations in the composition of the mitochondrial genome caused by various factors. The mutational process turned out to be a tool for studying the above role, and the consequences of its action of interest to us were the so-called. mitochondrial diseases.

Mitochondrial diseases are an example of cytoplasmic heredity in humans, or rather "organelle heredity". This clarification should be made because now the existence, at least in some organisms, of cytoplasmic hereditary determinants not associated with cell organelles - cytogens has been proven (Vechtomov, 1996).

Mitochondrial diseases are a heterogeneous group of diseases caused by genetic, structural, biochemical defects of mitochondria and impaired tissue respiration. For the diagnosis of mitochondrial disease, a comprehensive genealogical, clinical, biochemical, morphological, and genetic analysis is important. The main biochemical sign of mitochondrial pathology is the development of lactic acidosis, usually hyperlactatacidemia combined with hyperpyruvatacidemia is detected. The number of different variants has reached 120 forms. There is a stable increase in the concentration of lactic and pyruvic acids in the cerebrospinal fluid.

Mitochondrial diseases (MB) represent a significant problem for modern medicine. According to the methods of hereditary transmission, among MBs, diseases are distinguished that are inherited monogenously according to the Mendelian type, in which, due to the mutation of nuclear genes, either the structure and functioning of mitochondrial proteins are disrupted, or the expression of mitochondrial DNA changes, as well as diseases caused by mutations of mitochondrial genes, which in are mainly passed on to the offspring through the maternal line.

Data of morphological studies, indicating a gross pathology of mitochondria: abnormal proliferation of mitochondria, polymorphism of mitochondria with disturbances in shape and size, disorganization of cristae, accumulations of abnormal mitochondria under the sarcolemma, paracrystalline inclusions in mitochondria, the presence of interfibrillar vacuoles

Forms of mitochondrial diseases

1 ... Mitochondrial Diseases Caused by Mitochondrial DNA Mutations

1.1 Diseases caused by deletions of mitochondrial DNA

1.1.1. Kearns-Sayre Syndrome

The disease manifests itself at the age of 4-18 years, progressive external ophthalmoplegia, retinitis pigmentosa, ataxia, intentional tremor, atrioventricular heart block, increased protein level in cerebrospinal fluid of more than 1 g / l, "torn" red fibers in biopsies of skeletal muscles

1.1.2 Pearson's Syndrome

The debut of the disease from birth or in the first months of life, sometimes it is possible to develop encephalomyopathies, ataxia, dementia, progressive external ophthalmoplegia, hypoplastic anemia, impaired exocrine pancreatic function, progressive course

2 Diseases due to mitochondrial DNA point mutations

Maternal type of inheritance, acute or subacute decrease in visual acuity in one or both eyes, combination with neurological and osteoarticular disorders, retinal microangiopathy, progressive course with the possibility of remission or restoration of visual acuity, onset of the disease at the age of 20-30 years

2.2 NAPR syndrome (neuropathy, ataxia, retinitis pigmentosa)

Maternal type of inheritance, a combination of neuropathy, ataxia and retinitis pigmentosa, delayed psychomotor development, dementia, the presence of "torn" red fibers in muscle tissue biopsies

2.3. MERRF syndrome (myoclonus-epilepsy, "torn" red fibers)

Maternal type of inheritance, onset of the disease at the age of 3-65 years, myoclonic epilepsy, ataxia, dementia in combination with neurosensory deafness, atrophy of the optic nerves and impaired deep sensitivity, lactic acidosis, during the EEG examination, generaliza- bathroom epileptic complexes, "torn" red fibers in skeletal muscle biopsies, progressive course

2.4 MELAS syndrome (mitochondrial encephalomyopathy, lactic acidosis, stroke-like episodes)

Maternal type of inheritance, onset of the disease before the age of 40, exercise intolerance, migraine-like headaches with nausea and vomiting, stroke-like episodes, convulsions, lactic acidosis, "torn" red fibers in muscle biopsies, progressive course.

3 .Pathology associated with defects in intergenomic communication

3.1 Mitochondrial DNA multiple deletion syndromes

Blepharoptosis, external ophthalmoplegia, muscle weakness, sensorineural deafness, optic atrophy, progressive course, "torn" red fibers in skeletal muscle biopsies, decreased activity of respiratory chain enzymes.

3.2 Mitochondrial DNA deletion syndrome

Autosomal recessive inheritance

Clinical forms:

3.2.1.Fatal infantile

a) severe hepatic impairment b) hepatopathy c) muscle hypotension

Debut in the neonatal period

3.2.2.Congenital myopathy

Severe muscle weakness, generalized hypotension, cardiomyopathy and seizures, kidney damage, glucosuria, aminoacidopathy, phosphaturia

3.2.3.Infantile myopathy

occurs in the first 2 years of life, progressive muscle weakness, atrophy of proximal muscle groups and loss of tendon reflexes, rapidly progressive course, death in the first 3 years of life.

4 .Mitochondrial diseases due to nuclear DNA mutations

4.1 Diseases associated with defects in the respiratory chain

4.1.1 Complex 1 (NADH: CoQ reductase) deficiency

Onset of the disease before 15 years of age, myopathy syndrome, delayed psychomotor development, impaired cardiovascular system, seizures resistant to therapy, multiple neurological disorders, progressive course

4.1.2. Complex 2 (succinate-CoQ reductase) deficiency

It is characterized by the syndrome of encephalomyopathy, progressive course, sub-roads, the development of ptosis is possible

4.1.3. Complex 3 (CoQ-cytochrome C-oxidoreductase) deficiency

Multisystem disorders, damage to various organs and systems, with the involvement of the central and peripheral nervous system, endocrine system, kidneys, progressive course

4.1.4. Complex (cytochrome C-oxidase) deficiency

4.1.4.1 Fatal infantile congenital lactic acidosis

Mitochondrial myopathy with renal failure or cardiomyopathy, debut at neonatal age, severe respiratory disorders, diffuse muscle hypotension, progressive course, death in the first year of life.

4.1.4.2.Benign infantile muscle weakness

Atrophy, with adequate and timely treatment, rapid stabilization of the process and recovery by 1-3 years of age is possible

5 .Menkes syndrome (trichopolyodystrophy)

A sharp delay in psychomotor development, growth retardation, impaired growth and dystrophic changes in hair,

6 ... Mitochondrial encephalomyopathies

6.1.Leigh's syndrome(subacute neurotic encephalomyelopathy)

It manifests itself after 6 months of life, muscle hypotonia, ataxia, nystagmus, pyramidal symptoms, ophthalmoplegia, atrophy of the optic nerves, often associated with cardiomyopathy and mild metabolic acidosis

6.2.Alpers syndrome(progressive sclerosing polydystrophy)

Degeneration of the gray matter of the brain in combination with cirrhosis of the liver, deficiency of complex 5 (ATP synthetase), delayed psychomotor development, ataxia, dementia, muscle weakness, progressive course of the disease, unfavorable prognosis

6.3 Coenzyme-Q deficiency

Metabolic crises, muscle weakness and fatigue, ophthalmoplegia, deafness, decreased vision, stroke-like episodes, ataxia, myoclonus epilepsy, kidney damage: glucosuria, aminoacidopathy, phosphaturia, endocrine disorders, progressive course, decreased activity of respiratory enzymes

7 .Diseases associated with impaired metabolism of lactic and pyruvic acids

7.1. Deficiency of pyruvate carboxylase Autosomal recessive mode of inheritance, the onset of the disease in the neonatal period, the symptom complex of a "flaccid child", seizures resistant to therapy, high concentrations of ketone bodies in the blood, hyperammonemia, hyperlysinemia, decreased activity of pyruvate carboxylase in skeletal muscles

7.2 Pyruvate dehydrogenase deficiency

Manifestation in the neonatal period, craniofacial dysmorphia, seizures resistant to therapy, impaired breathing and sucking, symptom complex "flaccid child", brain dysginesia, severe acidosis with a high content of lactate and pyruvate

7.3 Decreased activity of pyruvate dehydrogenase

Manifestation in the first year of life, microcephaly, delayed psychomotor development, ataxia, muscular dystonia, choreoathetosis, lactic acidosis with a high pyruvate content

7.4 Dihydrolipoyltransacetylase Deficiency

Autosomal recessive type of inheritance, the onset of the disease in the neonatal period, microcephaly, delayed psychomotor development, muscle hypotension followed by an increase in muscle tone, optic disc atrophy, lactic acidosis, decreased activity of dihydrolipoyltrans-acetylase

7.5 Deficiency of dihydrolipoyl dehydrogenase

Autosomal recessive type of inheritance, the onset of the disease in the first year of life, the symptom complex of a "flaccid child", dysmetabolic crises with vomiting and diarrhea, delayed psychomotor development, optic disc atrophy, lactic acidosis, increased serum alanine, α- ketoglutarate, branched-chain α-keto acids, decreased dihydrolipoyl dehydrogenase activity

8 Diseases due to defects in beta-oxidation of fatty acids

8.1 Deficiency of long-chain acetyl-CoA dehydrogenase

Autosomal recessive mode of inheritance, onset of the disease in the first months of life, metabolic crises with vomiting and diarrhea, symptoms of a "flaccid child", hypoglycemia, dicarboxylic aciduria, decreased activity of acetyl-CoA dehydrogenase of fatty acids with a long carbon chain

8.2 Deficiency of Middle Carbon Acetyl CoA Dehydrogenase

Autosomal recessive mode of inheritance, onset of the disease in the neonatal period or the first months of life, metabolic crises with vomiting and diarrhea,

muscle weakness and hypotension, sudden death syndrome often develops, hypoglycemia, dicarboxylic aciduria, decreased acetyl-CoA dehydrogenase of medium carbon chain fatty acids

8.3. Deficiency of Acetyl-CoA Dehydrogenase of Short Carbon Chain Fatty Acids

Autosomal recessive inheritance, different ages of disease onset, decreased exercise tolerance, metabolic crises with vomiting and diarrhea, muscle weakness and hypotension, increased urinary excretion of methyl succinic acid, acetyl-CoA dehydrogenase of short carbon chain fatty acids

8.4 Multiple deficiency of acetyl-CoA dehydrogenases of fatty acids

Neonatal form: craniofacial dysmorphia, brain dysginesia, severe hypoglycemia and acidosis, malignant course, decreased activity of all acetyl-CoA dehydrogenases of fatty acids,

Infantile form: sluggish child symptom complex, cardiomyopathy, metabolic crises, hypoglycemia and acidosis

8.5 Decreased activity of all acetyl-CoA dehydrogenases of fatty acids

Late debut form: periodic episodes of muscle weakness, metabolic crises, hypoglycemia and acidosis are less pronounced, intelligence is preserved,

9 .Krebs cycle fermentopathy

9.1 Fumarase deficiency

Autosomal recessive mode of inheritance, onset of the disease in the neonatal or neonatal period, microcephaly, generalized muscle weakness and hypotension, episodes of lethargy, rapidly progressive en-cephalopathy, poor prognosis

9.2 Succinate dehydrogenase deficiency

A rare disease characterized by progressive encephalomyopathy

9.3 Alpha-ketoglutarate dehydrogenase deficiency

Autosomal recessive type of inheritance, neonatal onset of the disease, microcephaly, symptom complex of a "flaccid child", episodes of lethargy, lactic acidosis, rapidly progressing course, a decrease in the content of enzymes of the Krebs cycle in tissues

9.4.Syndromes of deficiency of carnitine and enzymes of its metabolism

Deficiency of carnitine-palmitoyltransferrase-1, autosomal recessive type of inheritance, early onset of the disease, episodes of non-ketonemic hypoglycemic coma, hepatomegaly, hypertriglyceridemia and moderate hyperammoniemia, decreased activity of carnitine-palmitoyltransferrase-1 in the liver

9.5 Carnitine acylcarnitine translocase deficiency

Early onset of the disease, cardiovascular and respiratory disorders, symptoms of a "flaccid child", episodes of lethargy and coma, an increase in the concentration of carnitine esters and a long carbon chain against the background of a decrease in free carnitine in the blood serum, a decrease in the activity of carnitine-acylcarnitine-translocase

9.6 Carnitine palmitoyltransferrase-2 deficiency

Autosomal recessive inheritance, muscle weakness, myalgia, myoglobinuria, decreased activity of carnitine palmitoyltransferrase-2 in skeletal muscles

Autosomal recessive mode of inheritance, myopathic symptom complex, episodes of lethargy and lethargy, cardiomyopathy, episodes of hypoglycemia, decreased serum carnitine levels and increased urinary excretion of carnitine.

After analyzing such a ‘terrible’ list of pathologies associated with certain changes in the functioning of the mitochondrial (and not only) genome, certain questions arise. What are the products of mitochondrial genes and in which super-mega-vital cellular processes do they take part?

As it turned out, some of the above pathologies can occur when the synthesis of 7 subunits of the NADH dehydrogenase complex, 2 subunits of ATP synthetase, 3 subunits of cytochrome c oxidase and 1 subunit of ubiquinol cytochrome c reductase (cytochrome b) , which are the gene products of mitochondria. Based on this, it can be concluded that there is a key role for these proteins in the processes of cellular respiration, fatty acid oxidation and ATP synthesis, electron transfer in the electron transport system of the inner MT membrane, the functioning of the antioxidant system, etc.

Judging by the latest data on the mechanisms of apoptosis, many scientists have come to the conclusion that there is a center for the control of apoptosis, namely ...

The role of mitochondrial proteins has also been shown with antibiotics blocking MT synthesis. If human cells in tissue culture are treated with an antibiotic, for example, tetracycline or chloramphenicol, then after one or two divisions their growth will stop. This is due to inhibition of mitochondrial protein synthesis, leading to the appearance of defective mitochondria and, as a consequence, to insufficient formation of ATP. Why, then, can antibiotics be used to treat bacterial infections? There are several answers to this question:

1. Some antibiotics (such as erythromycin) do not pass through the inner membrane of mammalian mitochondria.

2. Most of the cells in our body do not divide or divide very slowly, so the replacement of existing mitochondria with new ones occurs just as slowly (in many tissues, half of the mitochondria are replaced in about five days or even longer). Thus, the number of normal mitochondria will decrease to a critical level only if the blockade of mitochondrial protein synthesis is maintained for many days.

3. Certain conditions inside the tissue prevent the penetration of certain drugs into the mitochondria of the most sensitive cells. For example, a high concentration of Ca2 + in the bone marrow leads to the formation of a Ca2 + tetracycline complex, which cannot penetrate into rapidly dividing (and therefore the most vulnerable) precursors of blood cells.

These factors make it possible to use some drugs that inhibit mitochondrial protein synthesis as antibiotics in the treatment of higher animals. Only two of these drugs have side effects: long-term treatment with large doses of chloramphenicol can lead to disruption of the hematopoietic function of the bone marrow (suppress the formation of red blood cells and leukocytes), and long-term use of tetracycline can damage the intestinal epithelium. But in both cases it is not yet completely clear whether these side effects are caused by a blockade of mitochondrial biogenesis or some other reason.

Output

The structural and functional features of the mt genome are as follows. First, it was found that mtDNA is transmitted from the mother to all of her

descendants and from her daughters to all subsequent generations, but sons do not pass on their DNA (maternal inheritance). Maternal character

mtDNA inheritance is probably due to two circumstances: either the proportion of paternal mtDNA is so small (paternal

more than one DNA molecule per 25 thousand maternal mtDNA) that they cannot be detected by existing methods, or replication of paternal mitochondria is blocked after fertilization. Second, the absence of combinative variability - mtDNA belongs to only one of the parents, therefore, there are no recombination events characteristic of nuclear DNA in meiosis, and the nucleotide sequence changes from generation to generation only due to mutations. Third, mtDNA has no introns

(a high probability that a random mutation will affect the coding region of DNA), protective histones and an effective DNA repair system - all this determines a 10 times higher mutation rate than in nuclear DNA. Fourth, inside the same cell, normal and mutant mtDNA can coexist simultaneously - the phenomenon of heteroplasmy (the presence of only normal or only mutant mtDNA is called homoplasmy). Finally, both chains are transcribed and translated in mtDNA, and in a number of characteristics, the genetic code of mtDNA differs from the universal one (UGA encodes tryptophan, AUA encodes methionine, AGA and AGG are stop

codons).

These properties and the aforementioned functions of the mt genome made the study of the variability of the nucleotide sequence of mtDNA an invaluable tool for physicians, forensic physicians, evolutionary biologists,

representatives of historical science in solving their specific tasks.

Since 1988, when it was discovered that mutations in mtDNA genes underlie mitochondrial myopathies (JY Holt et al., 1988) and Leber's hereditary optic neuropathy (DC Wallace, 1988), further systematic identification of mutations in the human mt genome led to the formation of the concept of mitochondrial diseases (MB). Currently, pathological mutations of mtDNA are discovered in every type of mitochondrial genes.

Bibliography

1. Skulachev, mitochondria and oxygen, Soros. educated. zhurn.

2. Fundamentals of Biochemistry: In three volumes, M .: Mir,.

3. Nicholes D. G. Bioenergetics, An Introd. to the Chemiosm. Th., Acad. Press, 1982.

4. Stryer L. Biochemistry, 2nd ed. San Fransisco, Freeman, 1981.

5. Skulachev of biological membranes. M., 1989.

6., Chentsov reticulum: Structure and some functions // Results of science. General problems of biology. 1989

7. Chentsov cytology. M .: Publishing house of Moscow State University, 1995

8. , Sphere of competence of the mitochondrial genome // Vestn. RAMS, 2001. No. 10, pp. 31-43.

9. Holt I. J, Harding A. E., Morgan-Hughes I. A. Deletion of muscle mitochondrial DNA in patients with mitochondrial myopathies. Nature 1988, 331: 717-719.

10. and etc. Human genome and genes of predisposition. SPb., 2000

11. , Mitochondrial genome. Novosibirsk, 1990.

12. // Soros. educated. zhurn. 1999. No. 10. S.11-17.

13. The role of symbiosis in cell evolution. M., 1983.

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15. // Soros. educated. zhurn. 2000. No. 1. S.32-36.

Kiev National University. Taras Shevchenko

Department of Biology

abstract

on the topic:

"The role of the maternal genome in offspring development"

withthudenta IVcourse

Department of Biochemistry

Frolova Artem

Kiev 2004

Plan:

Introduction................................................. ..............................1

Symbiotic theory of the origin of mitochondria ... 2

The role of the cell nucleus in mitochondrial biogenesis ................................... 5

Transport systems of mitochondria ............................................... ...... 7

Size and shape of mitochondrial genomes .................. 10

Functioning of the mitochondrial genome ............... 14

The importance of having your own genetic system for mitochondria ........................................... ...................................19

Cytoplasmic inheritance .............................. 20

Historically, the first study of this kind was conducted using mitochondrial DNA. Scientists took a sample from the aborigines of Africa, Asia, Europe, America and in this, at first small, sample compared the mitochondrial DNA of different individuals with each other. They found that the diversity of mitochondrial DNA is highest in Africa. And since it is known that mutational events can change the type of mitochondrial DNA, and it is also known how it can change, then, therefore, we can say which types of people from which could have been mutated. In all the people who had their DNA tested, it was in the Africans that much more variability was found. The types of mitochondrial DNA on other continents were less diverse. This means that Africans had more time to accumulate these changes. They had more time for biological evolution, if it is in Africa that ancient remnants of DNA are found that are not characteristic of European human mutations.

It can be argued that mitochondrial DNA geneticists were able to prove the origin of women in Africa. They also studied the Y chromosomes. It turned out that men also come from Africa.

Thanks to research on mitochondrial DNA, it is possible to establish not only that a person originated from Africa, but also to determine the time of his origin. The time of the appearance of the mitochondrial foremother of mankind was established through a comparative study of the mitochondrial DNA of chimpanzees and modern humans. Knowing the rate of mutational divergence - 2-4% per million years - it is possible to determine the time of separation of two branches, chimpanzees and modern humans. This happened about 5 - 7 million years ago. In this case, the rate of mutational divergence is considered constant.

Mitochondrial eve

When people speak of mitochondrial Eve, they do not mean an individual. They talk about the emergence by evolution of a whole population of individuals with similar traits. It is believed that mitochondrial Eve lived during a period of sharp decline in the number of our ancestors, to about ten thousand individuals.

Origin of races

Studying the mitochondrial DNA of different populations, geneticists suggested that even before leaving Africa, the ancestral population was divided into three groups, giving rise to three modern races - African, Caucasian and Mongoloid. It is believed that this happened about 60 - 70 thousand years ago.

Comparison of mitochondrial DNA of nonstandard and modern humans

Additional information about the origin of humans was obtained by comparing the genetic texts of the mitochondrial DNA of Neanderthal and modern humans. Scientists were able to read the genetic texts of the mitochondrial DNA of the bone remains of two Neanderthals. The bones of the first Neanderthal man were found in the Feldhover Cave in Germany. A little later, the genetic text of the mitochondrial DNA of a Neanderthal child was read, which was found in the North Caucasus in the Mezhmayskaya cave. When comparing the mitochondrial DNA of modern humans and Neanderthals, very large differences were found. If we take a piece of DNA, then 27 out of 370 nucleotides differ. And if we compare the genetic texts of a modern man, his mitochondrial DNA, then only eight nucleotides differ. It is believed that the Neanderthal and modern man are completely separate branches, the evolution of each of them proceeded independently of each other.

When studying the difference in the genetic texts of the mitochondrial DNA of Neanderthal and modern humans, the date of the separation of these two branches was established. This happened about 500 thousand years ago, and about 300 thousand years ago, their final separation took place. It is believed that the Neanderthals settled in Europe and Asia and were driven out by a modern-type man who left Africa 200 thousand years later. And finally, about 28 - 35 thousand years ago, the Neanderthals became extinct. Why this happened, in general, is not yet clear. Maybe they could not stand the competition with a modern-type person, or maybe there were other reasons for this.

© G.M. Dymshits

Mitochondrial genome surprises

G.M. Dymshits

Grigory Moiseevich Dymshits, Doctor of Biological Sciences, Professor of the Department of Molecular Biology, Novosibirsk State University, Head of the Genome Structure Laboratory at the Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences. Co-author and editor of four school textbooks in general biology.

Today, human mitochondrial genetics is intensively developing both in the population and in the medical aspect. A relationship has been established between a number of severe hereditary diseases and defects in mitochondrial DNA. The genetic changes associated with aging are most pronounced in mitochondria. What is the mitochondrial genome, which differs in humans and other animals from that in plants, fungi and protozoa in size, shape, and genetic capacity? How does the mitochondrial genome work and how did it appear in different taxa? This is what will be discussed in our article.

Mitochondria are called the power plants of the cell. In addition to the outer smooth membrane, they have an inner membrane that forms numerous folds - cristae. They have built-in protein components of the respiratory chain - enzymes involved in converting the energy of chemical bonds of oxidized nutrients into the energy of adenosine triphosphoric acid (ATP) molecules. With this “convertible currency,” the cell pays for all of its energy needs. In the cells of green plants, in addition to mitochondria, there are also other energy stations - chloroplasts. They run on "solar cells", but they also form ATP from ADP and phosphate. Like mitochondria, chloroplasts - autonomously reproducing organelles - also have two membranes and contain DNA.

In addition to DNA, the mitochondrial matrix contains its own ribosomes, which differ in many characteristics from the eukaryotic ribosomes located on the membranes of the endoplasmic reticulum. However, on the ribosomes of mitochondria, no more than 5% of all proteins that make up their composition are formed. Most of the proteins that make up the structural and functional components of mitochondria are encoded by the nuclear genome, synthesized on the ribosomes of the endoplasmic reticulum, and transported through its channels to the assembly site. Thus, mitochondria are the result of the combined efforts of two genomes and two transcription and translation machines. Some subunit enzymes of the mitochondrial respiratory chain are composed of different polypeptides, some of which are encoded by the nuclear and some by the mitochondrial genome. For example, the key enzyme of oxidative phosphorylation, cytochrome c oxidase, in yeast consists of three subunits encoded and synthesized in mitochondria, and four subunits encoded in the cell nucleus and synthesized in the cytoplasm. The expression of most mitochondrial genes is controlled by specific genes in the nucleus.

Sizes and shapes of mitochondrial genomes

To date, more than 100 different mitochondrial genomes have been read. The set and number of their genes in mitochondrial DNA, for which the nucleotide sequence is fully determined, differ greatly in different species of animals, plants, fungi and protozoa. The largest number of genes found in the mitochondrial genome of the flagellate protozoan Rectinomonas americana- 97 genes, including all protein-coding genes found in the mtDNA of other organisms. In most higher animals, the mitochondrial genome contains 37 genes: 13 for the proteins of the respiratory chain, 22 for tRNA, and two for rRNA (for the large 16S rRNA ribosome subunit and for the small 12S rRNA). In plants and protozoa, unlike animals and most fungi, some proteins that make up the ribosomes of these organelles are encoded in the mitochondrial genome. Key enzymes of template polynucleotide synthesis, such as DNA polymerase (replicating mitochondrial DNA) and RNA polymerase (transcribing the mitochondrial genome), are encoded in the nucleus and synthesized on the cytoplasmic ribosomes. This fact indicates the relativity of mitochondrial autonomy in the complex hierarchy of the eukaryotic cell.

The genomes of mitochondria of different species differ not only in the set of genes, the order of their location and expression, but in the size and shape of DNA. The vast majority of mitochondrial genomes described today are circular, supercoiled, double-stranded DNA molecules. In some plants, along with circular forms, there are also linear ones, and in some protozoa, for example, ciliates, only linear DNA is found in mitochondria.

Typically, each mitochondrion contains several copies of its genome. So, in the cells of the human liver there are about 2 thousand mitochondria, and in each of them there are 10 identical genomes. In mouse fibroblasts, there are 500 mitochondria containing two genomes, and in yeast cells S.cerevisiae- up to 22 mitochondria with four genomes.

The mitochondrial genome of plants, as a rule, consists of several molecules of different sizes. One of them, the “main chromosome”, contains most of the genes, and the ring forms of shorter length, which are in dynamic equilibrium both with each other and with the main chromosome, are formed as a result of intra- and intermolecular recombination due to the presence of repeated sequences (Fig. 1 ).

Fig 1. Scheme of the formation of circular DNA molecules of different sizes in plant mitochondria.
Recombination occurs over repeated areas (marked in blue).

Fig 2. Scheme of formation of linear (A), circular (B), chain (C) mtDNA oligomers.
ori - region of the beginning of DNA replication.

The genome size of mitochondria of different organisms ranges from less than 6 thousand base pairs in the malaria plasmodium (in addition to two rRNA genes, it contains only three genes encoding proteins) to hundreds of thousands of base pairs in terrestrial plants (for example, in Arabidopsis thaliana from the cruciferous family 366924 base pairs). At the same time, 7-8-fold differences in the size of mtDNA of higher plants are found even within the same family. The length of mtDNA of vertebrates differs slightly: in humans - 16,569 base pairs, in a pig - 16350, in a dolphin - 16330, in a clawed frog Xenopus laevis- 17533, in carp - 16400. These genomes are also similar in the localization of genes, most of which are located end-to-end; in some cases they even overlap, usually by one nucleotide, so that the last nucleotide of one gene is the first in the next. Unlike vertebrates, in plants, fungi, and protozoa, mtDNAs contain up to 80% of non-coding sequences. The order of genes in mitochondrial genomes differs from species to species.

A high concentration of reactive oxygen species in mitochondria and a weak repair system increase the frequency of mtDNA mutations by an order of magnitude in comparison with the nuclear one. Oxygen radicals cause specific substitutions C® T (deamination of cytosine) and G® T (oxidative damage to guanine), as a result of which, possibly, mtDNA is rich in AT-pairs. In addition, all mtDNAs have an interesting property - they are not methylated, unlike nuclear and prokaryotic DNAs. It is known that methylation (temporary chemical modification of the nucleotide sequence without disrupting the coding function of DNA) is one of the mechanisms of programmed gene inactivation.

Replication and transcription of mammalian mitochondrial DNA

In most animals, complementary strands in mtDNA differ significantly in specific gravity, since they contain an unequal amount of “heavy” purine and “light” pyrimidine nucleotides. So they are called - H (heavy - heavy) and L (light - light) chain. At the beginning of the replication of the mtDNA molecule, a so-called D-loop (from the English displacement loop) is formed. This structure, visible under an electron microscope, consists of double-stranded and single-stranded (the retracted part of the H-chain) sections. The double-stranded region is formed by a part of the L-chain and a complementary newly synthesized DNA fragment of 450-650 (depending on the type of organism) nucleotides in length, which has a ribonucleotide primer at the 5'-end, which corresponds to the start point of the synthesis of the H-chain (ori H). The L-chain begins only when the daughter H-chain reaches the point ori L. This is due to the fact that the region of initiation of replication of the L-chain is accessible to the enzymes of DNA synthesis only in a single-stranded state, and therefore only in an untwisted double helix during the synthesis of H Thus, the daughter chains of mtDNA are synthesized continuously and asynchronously (Fig. 3).

Fig 3. Mammalian mtDNA replication scheme.
First, a D-loop is formed, then a daughter H-chain is synthesized,
then the synthesis of the daughter L-chain begins.

In mitochondria, the total number of D-loop molecules significantly exceeds the number of fully replicating molecules. This is due to the fact that the D-loop has additional functions - the attachment of mtDNA to the inner membrane and the initiation of transcription, since the transcription promoters of both DNA strands are localized in this region.

Unlike most eukaryotic genes, which are transcribed independently of each other, each of the mammalian mtDNA strands is rewritten to form one RNA molecule starting in the ori H region. In addition to these two long RNA molecules complementary to the H and L chains, more short sections of the H-chain that start at the same point and end at the 3 'end of the 16S rRNA gene (Fig. 4). There are 10 times more such short transcripts than long ones. As a result of maturation (processing), 12S rRNA are formed from them and 16S rRNA, which are involved in the formation of mitochondrial ribosomes, as well as phenylalanine and valine tRNAs The remaining tRNAs are excised from the long transcripts and translated mRNAs are formed, to the 3 'ends of which polyadenyl sequences are attached. The 5 "ends of these mRNAs are not capped, which is unusual for eukaryotes. No splicing occurs because none of the mammalian mitochondrial genes contain introns.

Fig 4. Transcription of human mtDNA containing 37 genes. All transcripts begin to be synthesized in the ori H region. Ribosomal RNAs are excised from the long and short H chain transcripts. tRNA and mRNA are formed as a result of processing from transcripts of both DNA strands. The tRNA genes are shown in light green.

Despite the fact that the genomes of mammalian and yeast mitochondria contain approximately the same number of genes, the size of the yeast genome is 4-5 times larger - about 80 thousand base pairs. Although the coding sequences of yeast mtDNA are highly homologous to those in humans, yeast mRNAs additionally have a 5 "leader and 3" non-coding region, like most nuclear mRNAs. A number of genes also contain introns. Thus, the box gene encoding cytochrome oxidase b contains two introns. A copy of most of the first intron is autocatalytically excised from the primary RNA transcript (without the participation of any proteins). The remaining RNA serves as a template for the formation of the splicing enzyme maturase. Part of its amino acid sequence is encoded in the remaining copies of the introns. Maturase cuts them out, destroying its own mRNA, copies of the exons are stitched together, and mRNA for cytochrome oxidase b is formed (Fig. 5). The discovery of such a phenomenon forced us to reconsider the concept of introns as “nothing coding sequences”.

Fig 5. Processing (maturation) of cytochrome oxidase b mRNA in yeast mitochondria.
At the first stage of splicing, mRNA is formed, according to which maturase is synthesized,
required for the second stage of splicing.

When studying the expression of mitochondrial genes Trypanosoma brucei discovered a surprising deviation from one of the basic axioms of molecular biology, which states that the sequence of nucleotides in mRNA exactly matches that in the coding regions of DNA. It turned out that the mRNA of one of the subunits of cytochrome c oxidase is being edited, i.e. after transcription, its primary structure changes - four uracils are inserted. As a result, a new mRNA is formed, which serves as a template for the synthesis of an additional subunit of the enzyme, the amino acid sequence of which has nothing in common with the sequence encoded by the unedited mRNA (see table).

RNA editing, discovered for the first time in trypanosome mitochondria, is widespread in chloroplasts and mitochondria of higher plants. It was also found in somatic mammalian cells, for example, in the human intestinal epithelium, the mRNA of the apolipoprotein gene is edited.

The greatest surprise to scientists was presented by mitochondria in 1979. Until that time, it was believed that the genetic code is universal and the same triplets encode the same amino acids in bacteria, viruses, fungi, plants and animals. English researcher Burrell compared the structure of one of the calf mitochondrial genes with the amino acid sequence in the cytochrome oxidase subunit encoded by this gene. It turned out that the genetic code of mitochondria in cattle (as well as in humans) is not just different from the universal one, it is “ideal”, ie. obeys the following rule: “if two codons have two identical nucleotides, and the third nucleotides belong to the same class (purine - A, G, or pyrimidine - Y, C), then they encode the same amino acid”. There are two exceptions to this rule in the universal code: the AUA triplet encodes isoleucine, and the AUG codon - methionine, while in the ideal mitochondrial code both of these triplets encode methionine; the UGG triplet encodes only tryptophan, and the UGA triplet encodes a stop codon. In the universal code, both deviations relate to the fundamental moments of protein synthesis: the AUG codon is the initiator, and the UGA stop codon stops polypeptide synthesis. The ideal code is not inherent in all mitochondria described, but none of them has a universal code. We can say that mitochondria speak different languages, but never speak the language of the nucleus.

As already mentioned, there are 22 tRNA genes in the mitochondrial genome of vertebrates. How does such an incomplete set serve all 60 codons for amino acids (in an ideal code of 64 triplets, there are four stop codons, in a universal one - three)? The fact is that during protein synthesis in mitochondria, codon-anticodon interactions are simplified - two of the three anticodon nucleotides are used for recognition. Thus, one tRNA recognizes all four members of the codon family, which differ only by the third nucleotide. For example, leucine tRNA with the GAU anticodon stands on the ribosome opposite to the codons CUU, CUC, CUA, and CUG, ensuring the error-free incorporation of leucine into the polypeptide chain. The other two leucine codons, UUA and UUG, are recognized by tRNA with the anticodon AAU. In total, eight different tRNA molecules recognize eight families of four codons each, and 14 tRNAs recognize different pairs of codons, each of which encodes one amino acid.

It is important that the aminoacyl tRNA synthetase enzymes responsible for the attachment of amino acids to the corresponding tRNA of mitochondria are encoded in the cell nucleus and synthesized on the ribosomes of the endoplasmic reticulum. Thus, in vertebrates, all protein components of mitochondrial polypeptide synthesis are encoded in the nucleus. At the same time, the synthesis of proteins in mitochondria is not suppressed by cycloheximide, which blocks the work of eukaryotic ribosomes, but is sensitive to the antibiotics erythromycin and chloramphenicol, which inhibit protein synthesis in bacteria. This fact serves as one of the arguments in favor of the origin of mitochondria from aerobic bacteria during the symbiotic formation of eukaryotic cells.

Symbiotic theory of the origin of mitochondria

The hypothesis of the origin of mitochondria and plant plastids from intracellular bacteria-endosymbionts was expressed by R. Altman back in 1890. During the century of rapid development of biochemistry, cytology, genetics and molecular biology that appeared half a century ago, the hypothesis grew into a theory based on a large amount of factual material. Its essence is as follows: with the appearance of photosynthetic bacteria, oxygen accumulated in the Earth's atmosphere - a by-product of their metabolism. With an increase in its concentration, the life of anaerobic heterotrophs became more difficult, and some of them, to obtain energy, passed from anoxic fermentation to oxidative phosphorylation. Such aerobic heterotrophs could, with a higher efficiency than anaerobic bacteria, break down organic matter formed as a result of photosynthesis. Some of the free-living aerobes were captured by anaerobes, but not “digested”, but preserved as energy stations, mitochondria. Mitochondria should not be viewed as slaves taken prisoner to supply ATP molecules to cells that are unable to breathe. Rather, they are “creatures” who, back in the Proterozoic, found the best of shelters for themselves and their offspring, where you can spend the least effort without risking being eaten.

Numerous facts speak in favor of the symbiotic theory:

- the size and shape of mitochondria and free-living aerobic bacteria are the same; both contain circular DNA molecules that are not associated with histones (as opposed to linear nuclear DNA);

In terms of nucleotide sequences, ribosomal and transport RNAs of mitochondria differ from nuclear ones, while demonstrating surprising similarity with analogous molecules of some aerobic gram-negative eubacteria;

Mitochondrial RNA polymerases, although encoded in the cell nucleus, are inhibited by rifampicin, like bacterial ones, and eukaryotic RNA polymerases are insensitive to this antibiotic;

Protein synthesis in mitochondria and bacteria is suppressed by the same antibiotics that do not affect the ribosomes of eukaryotes;

The lipid composition of the inner membrane of mitochondria and bacterial plasmalemma is similar, but very different from that of the outer membrane of mitochondria, which is homologous to other membranes of eukaryotic cells;

Cristae, formed by the inner mitochondrial membrane, are evolutionary analogs of the mesosomal membranes of many prokaryotes;

Until now, organisms have survived that mimic intermediate forms on the path to the formation of mitochondria from bacteria (primitive amoeba Pelomyxa does not have mitochondria, but always contains endosymbiotic bacteria).

The new genome can create metabolic pathways that lead to the formation of useful products that cannot be synthesized by any of the partners individually. Thus, the synthesis of steroid hormones by cells of the adrenal cortex is a complex chain of reactions, some of which occur in the mitochondria, and some in the endoplasmic reticulum. By capturing the genes of the promitochondria, the nucleus was able to reliably control the functions of the symbiont. The nucleus encodes all proteins and lipid synthesis of the outer membrane of mitochondria, most of the proteins of the matrix and the inner membrane of organelles. Most importantly, the nucleus encodes enzymes for replication, transcription and translation of mtDNA, thereby controlling the growth and reproduction of mitochondria. The growth rate of symbiotic partners should be approximately the same. If the host grows faster, then with each generation the number of symbionts per individual will decrease, and, in the end, descendants without mitochondria will appear. We know that every cell of a sexually reproducing organism contains many mitochondria, which replicate their DNA in the interval between divisions of the host. This ensures that each of the daughter cells will receive at least one copy of the mitochondrial genome.

Cytoplasmic inheritance

In addition to coding the key components of the respiratory chain and its own protein-synthesizing apparatus, the mitochondrial genome in some cases participates in the formation of some morphological and physiological characteristics. These features include the NCS syndrome (non-chromosomal stripe, nonchromosomally encoded leaf spot) and cytoplasmic male sterility (CMS), which leads to disruption of the normal development of pollen, which are characteristic of a number of higher plant species. The manifestation of both features is due to changes in the structure of mtDNA. In CMS, rearrangements of mitochondrial genomes are observed as a result of recombination events leading to deletions, duplications, inversions or insertions of certain nucleotide sequences or whole genes. Such changes can cause not only damage to existing genes, but also the emergence of new working genes.

Cytoplasmic heredity, unlike nuclear, does not obey Mendel's laws. This is due to the fact that in higher animals and plants gametes from different sexes contain incomparable amounts of mitochondria. So, in the mouse egg there are 90 thousand mitochondria, and in the sperm - only four. Obviously, in a fertilized egg, mitochondria are predominantly or only from a female, i.e. the inheritance of all mitochondrial genes is maternal. Genetic analysis of cytoplasmic heredity is difficult due to nuclear-cytoplasmic interactions. In the case of cytoplasmic male sterility, the mutant mitochondrial genome interacts with certain nuclear genes, the recessive alleles of which are necessary for the development of the trait. The dominant alleles of these genes, both in homo- and heterozygous states, restore plant fertility regardless of the state of the mitochondrial genome.

The study of mitochondrial genomes, their evolution proceeding according to the specific laws of population genetics, the relationship between nuclear and mitochondrial genetic systems, is necessary to understand the complex hierarchical organization of the eukaryotic cell and the organism as a whole.

Certain hereditary diseases and human aging have been linked to certain mutations in mitochondrial DNA or in the nuclear genes that control mitochondrial function. Data are accumulating on the involvement of mtDNA defects in carcinogenesis. Hence, mitochondria may be the target of cancer chemotherapy. There are facts about the close interaction of the nuclear and mitochondrial genomes in the development of a number of human pathologies. Multiple deletions of mtDNA were found in patients with severe muscle weakness, ataxia, deafness, mental retardation, inherited in an autosomal dominant manner. Established sexual dimorphism in the clinical manifestations of coronary heart disease, which is most likely due to the maternal effect - cytoplasmic heredity. The development of gene therapy offers hope for correcting defects in mitochondrial genomes in the foreseeable future.

This work was supported by the Russian Foundation for Basic Research. Project 01-04-48971.
The author is grateful to graduate student M.K. Ivanov, who created the figures for the article.

Literature

1. Yankovsky N.K., Borinskaya S.A. Our history recorded in DNA // Nature. 2001. No. 6. S.10-18.

2. Minchenko A.G., Dudareva N.A. Mitochondrial genome. Novosibirsk, 1990.

3. Gvozdev V.A.// Soros. educated. zhurn. 1999. No. 10. S.11-17.

4. Margelis L. The role of symbiosis in cell evolution. M., 1983.

5. Skulachev V.P.// Soros. educated. zhurn. 1998. No. 8. S.2-7.

6. Igamberdiev A.U.// Soros. educated. zhurn. 2000. No. 1. S.32-36.

Mitochondrial DNA located in the matrix is ​​a closed circular double-stranded molecule in human cells having a size of 16569 nucleotide pairs, which is approximately 10 5 times smaller than the DNA localized in the nucleus. In general, mitochondrial DNA encodes 2 rRNA, 22 tRNA, and 13 subunits of the respiratory chain enzymes, which is no more than half of the proteins found in it. In particular, under the control of the mitochondral genome, seven subunits of ATP synthetase, three subunits of cytochrome oxidase, and one subunit of ubiquinol-cytochrome are encoded. with-reductases. In this case, all proteins, except one, two ribosomal and six tRNAs are transcribed from the heavier (outer) DNA strand, and 14 other tRNAs and one protein are transcribed from the lighter (inner) strand.

Against this background, the genome of plant mitochondria is much larger and can reach 370,000 nucleotide pairs, which is about 20 times larger than the human mitochondrial genome described above. The number of genes here is also about 7 times greater, which is accompanied by the appearance in plant mitochondria of additional electron transport pathways that are not associated with ATP synthesis.

Mitochondrial DNA replicates in interphase, which is partially synchronized with DNA replication in the nucleus. During the cell cycle, mitochondria divide in two by a constriction, the formation of which begins with an annular groove on the inner mitochondrial membrane. A detailed study of the nucleotide sequence of the mitochondrial genome made it possible to establish that in the mitochondria of animals and fungi, deviations from the universal genetic code are frequent. Thus, in human mitochondria, the TAT codon instead of isoleucine in the standard code encodes the amino acid methionine, the TCT and TCC codons, usually encoding arginine, are stop codons, and the AST codon, which is the stop codon in the standard code, encodes the amino acid methionine. As for plant mitochondria, it seems that they use a universal genetic code. Another feature of mitochondria is the recognition of tRNA codons, which consists in the fact that one such molecule is able to recognize not one, but three or four codons at once. This feature reduces the significance of the third nucleotide in the codon and leads to the fact that mitochondria require a smaller variety of tRNA types. In this case, only 22 different tRNAs are sufficient.

Having its own genetic apparatus, the mitochondrion also has its own protein-synthesizing system, a feature of which in the cells of animals and fungi are very small ribosomes characterized by a sedimentation coefficient of 55S, which is even lower than that of 70s ribosomes of the prokaryotic type. In this case, two large ribosomal RNAs are also smaller than in prokaryotes, and small rRNA is absent altogether. In plant mitochondria, on the contrary, ribosomes are more similar to prokaryotic ones in size and structure.

Properties and functions of DNA.

DNA, or deoxyribonucleic acid, is the main hereditary material present in all cells of the body and mainly involves the blue seal of cell functions, growth, reproduction and death. A DNA structure called double-stranded helical structure was first described by Watson and Crick in 1953.

From then on, tremendous progress has been made in the synthesis, sequencing and manipulation of DNA. DNA these days can be virtualized or analyzed for little things and even genes can be inserted to trigger changes in DNA function and structure.

The main purpose of the hereditary material is the storage of hereditary information, on the basis of which the phenotype is formed. Most of the characteristics and properties of the organism are due to the synthesis of proteins that perform various functions.Thus, information on the structure of extremely diverse protein molecules, the specificity of which depends on the qualitative and quantitative composition of amino acids, as well as on the order of their arrangement in the peptide chain, must be recorded in the hereditary material. Consequently, the amino acid composition of proteins must be encoded in nucleic acid molecules.
Back in the early 50s, it was suggested about a method for recording genetic information, in which the coding of individual amino acids in a protein molecule should be carried out using certain combinations of four different nucleotides in a DNA molecule. To encrypt more than 20 amino acids, the required number of combinations is provided only by a triplet code, that is, a code that includes three adjacent nucleotides. In this case, the number of combinations of four nitrogenous bases, three each is 41 = 64. The assumption about the tripletness of the genetic code was later experimentally confirmed, and for the period from 1961 to 1964 a cipher was found, with the help of which the order of amino acids in nucleic acid molecules is written in peptide.
From table. 6 that out of 64 triplets, 61 triplets encode one or another amino acid, and individual amino acids are encrypted by more than one triplet, or codon (phenylalanine, leucine, valine, serine, etc.). Several triplets do not encode amino acids, and their functions are associated with the designation of the terminal region of the protein molecule.
Reading of information recorded in a nucleic acid molecule is carried out sequentially, co-Don by codon, so that each nucleotide is part of only one triplet.
The study of the genetic code in living organisms with different levels of organization has shown the universality of this mechanism for recording information in living nature.
Thus, research in the middle of the 20th century revealed a mechanism for recording hereditary information in nucleic acid molecules using a biological code, which is characterized by the following properties: a) triplet - amino acids are encrypted by triplets of nucleotides - codons; b) specificity - each triplet encodes only a certain amino acid; c) universality - in all living organisms, the coding of the same amino acids is carried out by the same codons; d) degeneracy - many amino acids are encrypted with more than one triplet; e) non-overlapping - information is read out successively triplet by triplet: AAGTSTCTSAGTSTSAT.

In addition to recording and storing biological information, the function of the material of heredity is its reproduction and transmission to a new generation in the process of reproduction of cells and organisms. This function of the hereditary material is carried out by DNA molecules in the process of its reduplication, i.e., absolutely exact reproduction of the structure, thanks to the implementation of the principle of complementarity (see 2.1).
Finally, the third function of the hereditary material, represented by DNA molecules, is to provide specific processes during the implementation of the information contained in it. This function is carried out with the participation of various types of RNA, which provide the translation process, that is, the assembly of a protein molecule that occurs in the cytoplasm on the basis of information received from the nucleus (see 2.4). In the course of the realization of hereditary information stored in the form of DNA molecules in the chromosomes of the nucleus, several stages are distinguished.
1. Reading information from a DNA molecule in the process of mRNA synthesis - transcription, which is carried out on one of the strands of the double helix of the DNA-codogenic chain according to the principle of complementarity (see 2.4).
2. Preparation of the transcription product for release into the cytoplasm - mRNA maturation.
3. Assembly on the ribosomes of the peptide chain of amino acids based on the information recorded in the mRNA molecule, with the participation of transport tRNA - translation (see 2.4).
4. Formation of secondary, tertiary and quaternary protein structures, which corresponds to the formation of a functioning protein (simple feature).
5. Formation of a complex trait as a result of the participation of products of several genes (protein-enzymes or other proteins) in biochemical processes.

The structure of the double helix of DNA, held together by only hydrogen bonds, can be easily destroyed. The breaking of hydrogen bonds between polynucleotide DNA chains can be carried out in strongly alkaline solutions (at pH> 12.5) or by heating. After that, the DNA strands are completely separated. This process is called DNA denaturation or melting.

Denaturation changes some of the physical properties of DNA, such as its optical density. Nitrogenous bases absorb light in the ultraviolet region (with a maximum close to 260 nm). DNA absorbs light almost 40% less than a mixture of free nucleotides of the same composition. This phenomenon is called the hypochromic effect, and it is due to the interaction of bases when they are located in a double helix.

Any deviation from the double-stranded state affects the change in the magnitude of this effect, i.e. there is a shift in optical density towards the value characteristic of free bases. Thus, DNA denaturation can be observed by changing its optical density.

When DNA is heated, the average temperature of the range at which DNA strands are separated is called the melting point and is designated as T pl... In solution T pl usually lies in the range 85-95 ° C. The DNA melting curve always has the same shape, but its position on the temperature scale depends on the base composition and denaturation conditions (Fig. 1). G-C pairs connected by three hydrogen bonds are more refractory than AT pairs having two hydrogen bonds, therefore, with an increase in the G-C-nap content, the value of T pl increases. DNA, 40% composed of G-C (characteristic of the mammalian genome), denatures at T pl about 87 ° C, while DNA containing 60% G-C has T pl
about 95 ° C.

The temperature of DNA denaturation (except for the composition of the bases) is influenced by the ionic strength of the solution. Moreover, the higher the concentration of monovalent cations, the higher T pl... T value pl also changes greatly when substances such as formamide (formic acid amide HCONH2) are added to the DNA solution, which
destabilizes hydrogen bonds. Its presence allows you to reduce T pl, up to 40 ° C.

The denaturation process is reversible. The phenomenon of restoration of the structure of a double helix, based on two separations of complementary strands, is called DNA renaturation. To carry out renaturation, as a rule, it is sufficient to pressurize the denatured DNA solution.

Renaturation involves two complementary sequences that were separated during denaturation. However, any complementary sequences that are capable of forming a double-stranded structure can be renatlated. If together. single-stranded DNA originating from different points is annealed, then the formation of a double-stranded DNA structure is called hybridization.

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