Mitochondrial DNA and family history. ☵ Mitochondrial DNA Why is mitochondrial DNA inherited maternally?

Surprises of the mitochondrial genome

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 of the Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences. Co-author and editor of four school textbooks on general biology.

A quarter of a century has passed since the discovery of DNA molecules in mitochondria before not only molecular biologists and cytologists became interested in them, but also geneticists, evolutionists, as well as paleontologists and criminologists, historians and linguists. Such widespread interest was provoked by the work of A. Wilson from the University of California. In 1987, he published the results of a comparative analysis of mitochondrial DNA taken from 147 representatives of different ethnic groups of all human races inhabiting five continents. Based on the type, location and number of individual mutations, it was established that all mitochondrial DNA arose from one ancestral nucleotide sequence through divergence. In the pseudo-scientific press, this conclusion was interpreted in an extremely simplified way - all of humanity descended from one woman, called mitochondrial Eve (both daughters and sons receive mitochondria only from their mother), who lived in North-East Africa about 200 thousand years ago. Another 10 years later, it was possible to decipher a fragment of mitochondrial DNA isolated from the remains of a Neanderthal, and estimate the existence of the last common ancestor of humans and Neanderthals at 500 thousand years ago.

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

Mitochondria are called the energy stations of the cell. In addition to the outer smooth membrane, they have an inner membrane that forms numerous folds - cristae. They contain 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 its energy needs. In the cells of green plants, in addition to mitochondria, there are also other energy stations - chloroplasts. They work on “solar batteries”, but 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 also contains its own ribosomes, which differ in many characteristics from eukaryotic ribosomes located on the membranes of the endoplasmic reticulum. However, no more than 5% of all proteins included in their composition are formed on the ribosomes of mitochondria. 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 site of assembly. Thus, mitochondria are the result of the combined efforts of two genomes and two transcription and translation apparatuses. Some subunit enzymes of the mitochondrial respiratory chain consist of different polypeptides, some of which are encoded by the nuclear genome 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 nuclear genes.

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 completely determined, vary greatly among different species of animals, plants, fungi and protozoa. The largest number of genes was found in the mitochondrial genome of flagellated protozoa 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 respiratory chain proteins, 22 for tRNA and two for rRNA (for the large ribosomal subunit 16S rRNA and for the small 12S rRNA). In plants and protozoa, unlike animals and most fungi, the mitochondrial genome also encodes some proteins that make up the ribosomes of these organelles. Key enzymes of template polynucleotide synthesis, such as DNA polymerase (replicating mitochondrial DNA) and RNA polymerase (transcribing the mitochondrial genome), are encrypted in the nucleus and synthesized on ribosomes in the cytoplasm. This fact indicates the relativity of mitochondrial autonomy in the complex hierarchy of the eukaryotic cell.

The mitochondrial genomes 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 the 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, such as ciliates, only linear DNA is found in the mitochondria.

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

The mitochondrial genome of plants typically consists of several molecules of varying sizes. One of them, the “main chromosome,” contains most of the genes, and smaller circular forms, 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 along repeated regions (indicated in blue).

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

The size of the mitochondrial genome of different organisms ranges from less than 6 thousand base pairs in the falciparum plasmodium (in addition to two rRNA genes, it contains only three protein-coding genes) to hundreds of thousands of base pairs in land plants (for example, Arabidopsis thaliana from the cruciferous family 366924 nucleotide pairs). Moreover, 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 - 16569 nucleotide pairs, in pigs - 16350, in dolphins - 16330, in clawed frogs 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, mtDNA contains up to 80% non-coding sequences. The order of genes in mitochondrial genomes differs among species.

The high concentration of reactive oxygen species in mitochondria and a weak repair system increase the frequency of mtDNA mutations by an order of magnitude compared to nuclear DNA. Oxygen radicals cause specific substitutions C®T (cytosine deamination) and G®T (oxidative damage to guanine), as a result of which mtDNA is possibly rich in AT pairs. In addition, all mtDNA 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.

Replication and transcription of mammalian mitochondrial DNA

In most animals, the complementary chains in mtDNA vary significantly in specific density, since they contain unequal amounts of “heavy” purine and “light” pyrimidine nucleotides. So they are called - H (heavy - heavy) and L (light - light) chain. At the beginning of replication of the mtDNA molecule, a so-called D-loop is formed (from the English displacement loop - displacement loop). This structure, visible under an electron microscope, consists of a double-stranded and a single-stranded (extended part of the H-chain) region. The double-stranded region is formed by part of the L-chain and a newly synthesized DNA fragment complementary to it, 450-650 nucleotides long (depending on the type of organism), having a ribonucleotide primer at the 5" end, which corresponds to the starting point of H-chain synthesis (ori H). Synthesis 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 DNA synthesis enzymes only in a single-stranded state, and therefore, only in an untwisted double helix during H synthesis -chains 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-strand is synthesized,
then the synthesis of the daughter L-chain begins.

In mitochondria, the total number of molecules with a D-loop significantly exceeds the number of fully replicating molecules. This is due to the fact that the D-loop has additional functions - attachment of mtDNA to the inner membrane and initiation of transcription, since 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 transcribed to form a single RNA molecule, starting in the ori H region. In addition to these two long RNA molecules, complementary to the H- and L-chains, more are formed short sections of the H-chain that begin 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 is formed from them and 16S rRNA, involved in the formation of mitochondrial ribosomes, as well as phenylalanine and valine tRNA. The remaining tRNAs are excised from 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. Splicing does not occur 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-strand transcripts. tRNA and mRNA are formed as a result of processing from transcripts of both strands of DNA. tRNA genes are indicated in light green.

Surprises of the mitochondrial genome

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 the corresponding sequences in humans, yeast mRNAs additionally have a 5" leader and a 3" noncoding region, like most nuclear mRNAs. A number of genes also contain introns. Thus, the box gene encoding cytochrome oxidase b has two introns. A copy of most of the first intron is excised from the primary RNA transcript autocatalytically (without the participation of any proteins). 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 the 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 this phenomenon forced us to reconsider the idea of introns as “non-coding sequences.”

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

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 cytochrome c oxidase subunits is 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).

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

Mitochondria presented the greatest surprise to scientists in 1979. Until that time, it was believed that the genetic code was universal and the same triplets encode the same amino acids in bacteria, viruses, fungi, plants and animals. 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 turned out that the genetic code of mitochondria in cattle (as well as in humans) not only differs from the universal one, it is “ideal”, i.e. 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 - U, C), then they code for the same amino acid.” In the universal code there are two exceptions to this rule: the AUA triplet encodes isoleucine and the AUG codon encodes 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 concern fundamental aspects of protein synthesis: the AUG codon is the initiating one, and the stop codon UGA stops the synthesis of the polypeptide. The ideal code is not inherent in all described mitochondria, but none of them has a universal code. We can say that mitochondria speak different languages, but never the language of the nucleus.

As already mentioned, there are 22 tRNA genes in the vertebrate mitochondrial genome. How does such an incomplete set serve all 60 codons for amino acids (the ideal code of 64 triplets has four stop codons, the universal code has three)? The fact is that during protein synthesis in mitochondria, codon-anticodon interactions are simplified - two out of three anticodon nucleotides are used for recognition. Thus, one tRNA recognizes all four members of a codon family, differing only in the third nucleotide. For example, leucine tRNA with the GAU anticodon stands on the ribosome opposite the codons TsU, TsUC, TsUA and Tsug, ensuring the error-free incorporation of leucine into the polypeptide chain. Two other 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 encoding one amino acid.

It is important that aminoacyl-tRNA synthetase enzymes, responsible for the addition of amino acids to the corresponding mitochondrial tRNAs, 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 encrypted in the nucleus. In this case, protein synthesis 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 about the origin of mitochondria and plant plastids from intracellular endosymbiont bacteria was expressed by R. Altman back in 1890. Over the century of rapid development of biochemistry, cytology, genetics and molecular biology, which appeared half a century ago, the hypothesis has grown into a theory based on a large amount of factual material. Its essence is this: with the advent of photosynthetic bacteria, oxygen accumulated in the Earth's atmosphere - a by-product of their metabolism. As its concentration increased, the life of anaerobic heterotrophs became more complicated, and some of them switched from oxygen-free fermentation to oxidative phosphorylation to obtain energy. Such aerobic heterotrophs could break down organic substances resulting from photosynthesis with greater efficiency than anaerobic bacteria. Some of the free-living aerobes were captured by anaerobes, but not “digested”, but stored as energy stations, mitochondria. Mitochondria should not be viewed as slaves, taken captive to supply ATP molecules to cells that are not capable of respiration. They are rather “creatures” that, back in the Proterozoic, found for themselves and their offspring the best of shelters, where they could expend the least amount of effort without running the risk of being eaten.

Numerous facts speak in favor of the symbiotic theory:

- the sizes and shapes of mitochondria and free-living aerobic bacteria coincide; both contain circular DNA molecules not associated with histones (unlike linear nuclear DNA);
In terms of nucleotide sequences, ribosomal and transfer RNAs of mitochondria differ from nuclear ones, while demonstrating surprising similarity with similar 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 the bacterial plasmalemma is similar, but is very different from that of the outer membrane of mitochondria, which is homologous to other membranes of eukaryotic cells;
The cristae formed by the inner mitochondrial membrane are the evolutionary analogues of the mesosomal membranes of many prokaryotes;
There are still organisms that imitate intermediate forms on the way 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 either partner alone. Thus, the synthesis of steroid hormones by cells of the adrenal cortex is a complex chain of reactions, some of which occur in mitochondria, and some in the endoplasmic reticulum. By capturing the promitochondrial genes, 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 mtDNA replication, transcription and translation, thereby controlling the growth and reproduction of mitochondria. The growth rate of symbiosis partners should be approximately the same. If the host grows faster, then with each generation the number of symbionts per individual will decrease, and, eventually, descendants without mitochondria will appear. We know that each cell of a sexually reproducing organism contains many mitochondria that replicate their DNA between divisions of the host. This ensures that each of the daughter cells receives at least one copy of the mitochondrial genome.

Cytoplasmic inheritance

In addition to encoding the key components of the respiratory chain and its own protein synthesizing apparatus, the mitochondrial genome in some cases is involved in the formation of some morphological and physiological characteristics. These traits include NCS syndrome (non-chromosomal stripe, non-chromosomal encoded leaf spot) and cytoplasmic male sterility (CMS), characteristic of a number of species of higher plants, which leads to disruption of the normal development of pollen. The manifestation of both signs 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 entire genes. Such changes can cause not only damage to existing genes, but also the emergence of new working genes.

Cytoplasmic inheritance, unlike nuclear inheritance, does not obey Mendel's laws. This is due to the fact that in higher animals and plants, gametes from different sexes contain disparate amounts of mitochondria. So, in a mouse egg there are 90 thousand mitochondria, but in a sperm there are only four. It is obvious that in a fertilized egg the mitochondria are predominantly or only from the female individual, i.e. Inheritance of all mitochondrial genes is maternal. Genetic analysis of cytoplasmic inheritance 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. 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, which follows the specific laws of population genetics, and the relationships 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 mutations in mitochondrial DNA or in nuclear genes that control mitochondria are associated with some hereditary diseases and human aging. Data are accumulating on the involvement 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 mtDNA deletions were found in patients with severe muscle weakness, ataxia, deafness, and mental retardation, inherited in an autosomal dominant manner. Sexual dimorphism has been established in the clinical manifestations of coronary heart disease, which is most likely due to the maternal effect - cytoplasmic inheritance. The development of gene therapy gives 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 drawings for the article.

Literature

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

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

3. Gvozdev V.A.// Soros. education magazine 1999. No. 10. P.11-17.

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

5. Skulachev V.P.// Soros. education magazine 1998. No. 8. P.2-7.

6. Igamberdiev A.U.// Soros. education magazine 2000. No. 1. P.32-36.

Genes that remained during evolution in the “energy stations of the cell” help to avoid management problems: if something breaks in the mitochondria, it can fix it itself, without waiting for permission from the “center.”

Our cells receive energy with the help of special organelles called mitochondria, which are often called the energy stations of the cell. Externally, they look like tanks with a double wall, and the inner wall is very uneven, with numerous strong indentations.

A cell with a nucleus (colored blue) and mitochondria (colored red). (Photo by NICHD/Flickr.com)

Mitochondria in section, outgrowths of the inner membrane are visible as longitudinal internal stripes. (Photo by Visuals Unlimited/Corbis.)

A huge number of biochemical reactions occur in mitochondria, during which “food” molecules are gradually oxidized and disintegrated, and the energy of their chemical bonds is stored in a form convenient for the cell. But, in addition, these “energy stations” have their own DNA with genes, which is served by their own molecular machines that provide RNA synthesis followed by protein synthesis.

It is believed that mitochondria in the very distant past were independent bacteria that were eaten by some other single-celled creatures (most likely archaea). But one day the “predators” suddenly stopped digesting the swallowed protomitochondria, keeping them inside themselves. A long rubbing of the symbionts with each other began; as a result, those who were swallowed greatly simplified their structure and became intracellular organelles, and their “hosts” were able, due to more efficient energy, to develop further into more and more complex forms of life, up to plants and animals.

The fact that mitochondria were once independent is evidenced by the remains of their genetic apparatus. Of course, if you live inside with everything ready-made, the need to contain your own genes disappears: the DNA of modern mitochondria in human cells contains only 37 genes - against 20-25 thousand of those contained in nuclear DNA. Over millions of years of evolution, many of the mitochondrial genes have moved to the cell nucleus: the proteins they encode are synthesized in the cytoplasm and then transported to the mitochondria. However, the question immediately arises: why did 37 genes still remain where they were?

Mitochondria, we repeat, are present in all eukaryotic organisms, that is, in animals, plants, fungi, and protozoa. Ian Johnston ( Iain Johnston) from the University of Birmingham and Ben Williams ( Ben P. Williams) from the Whitehead Institute analyzed more than 2,000 mitochondrial genomes taken from various eukaryotes. Using a special mathematical model, the researchers were able to understand which genes were more likely to remain in the mitochondria during evolution.

Introduction

A quarter of a century passed from the time DNA molecules were discovered in mitochondria before they became interested not only molecular biologists and cytologists, but also geneticists, evolutionists, as well as paleontologists and criminologists. Such widespread interest was provoked by the work of A. Wilson from the University of California. In 1987, he published the results of a comparative analysis of mitochondrial DNA taken from 147 representatives of different ethnic groups of all human races inhabiting five continents. Based on the type, location and number of individual mutations, it was established that all mitochondrial DNA arose from one ancestral nucleotide sequence by divergences. In the pseudo-scientific press, this conclusion was interpreted in an extremely simplified way - all of humanity descended from one woman, called mitochondrial Eve (since both daughters and sons receive mitochondria only from their mother), who lived in North-East Africa about 200 thousand years ago . Another 10 years later, it was possible to decipher a fragment of mitochondrial DNA isolated from the remains of a Neanderthal, and estimate the existence of the last common ancestor of humans and Neanderthals at 500 thousand years ago.

Today, human mitochondrial genetics is intensively developing both in population and medical aspects. A connection has been established between a number of severe hereditary diseases and defects in mitochondrial DNA. Genetic changes associated with aging are most pronounced in mitochondria. What is the mitochondrial genome that differs in humans and other animals from that of 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 will be discussed in my “small and most modest” essay.

In addition to DNA, the mitochondrial matrix also contains its own ribosomes, which differ in many characteristics from eukaryotic ribosomes located on the membranes of the endoplasmic reticulum. However, no more than 5% of all proteins included in their composition are formed on the ribosomes of mitochondria. 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 apparatuses. Some subunit enzymes of the mitochondrial respiratory chain consist of different polypeptides, some of which are encoded by the nuclear genome, 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 nuclear genes.

Symbiotic theory of the origin of mitochondria

The hypothesis about the origin of mitochondria and plant plastids from intracellular endosymbiont bacteria was expressed by R. Altman back in 1890. During a century of rapid development biochemistry , cytology, genetics and molecular biology, which appeared half a century ago, the hypothesis grew into a theory based on a large amount of factual material. Its essence is this: with the advent of photosynthetic bacteria, oxygen accumulated in the Earth's atmosphere - a by-product of their metabolism. As its concentration increased, the life of anaerobic heterotrophs became more complicated, and some of them switched from oxygen-free conditions to obtain energy. fermentation to oxidative phosphorylation. Such aerobic heterotrophs could, with greater efficiency than anaerobic bacteria, break down organic substances formed as a result of photosynthesis. Some of the free-living aerobes were captured by anaerobes, but not “digested”, but stored as energy stations, mitochondria. Mitochondria should not be viewed as slaves, taken captive in order to supply ATP molecules to cells that are not capable of respiration. They are rather “creatures” that, back in the Proterozoic, found for themselves and their offspring the best of shelters, where they could expend the least amount of effort without running the risk of being eaten.

Numerous facts speak in favor of the symbiotic theory:

The sizes and shapes of mitochondria and free-living aerobic bacteria coincide; both contain circular DNA molecules not associated with histones (unlike linear nuclear DNA);

In terms of nucleotide sequences, ribosomal and transfer RNAs of mitochondria differ from nuclear ones, while demonstrating surprising similarity with similar 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 the bacterial plasmalemma is similar, but is very different from that of the outer membrane of mitochondria, which is homologous to other membranes of eukaryotic cells;

The cristae formed by the inner mitochondrial membrane are the evolutionary analogues of the mesosomal membranes of many prokaryotes;

There are still organisms that imitate intermediate forms on the way 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 bacterial endosymbiosis arose at different stages of the evolution of living organisms. This is also evidenced by differences in the structure of the mitochondrial genomes of protozoa, fungi, plants and higher animals. But in all cases, the main part of the genes from promitochondria entered the nucleus, possibly with the help of mobile genetic elements. When part of the genome of one of the symbionts is included in the genome of another, the integration of the symbionts becomes irreversible. The new genome can create metabolic pathways that lead to the formation of useful products that cannot be synthesized by either partner individually. Thus, the synthesis of steroid hormones by cells of the adrenal cortex is a complex chain of reactions, some of which occur in mitochondria, and some in the endoplasmic reticulum. By capturing the promitochondrial genes, 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 mtDNA replication, transcription and translation, thereby controlling the growth and reproduction of mitochondria. The growth rate of symbiosis partners should be approximately the same. If the host grows faster, then with each generation the number of symbionts per individual will decrease, and, eventually, descendants without mitochondria will appear. We know that each cell of a sexually reproducing organism contains many mitochondria that replicate their DNA between divisions of the host. This ensures that each of the daughter cells receives at least one copy of the mitochondrial genome.

The role of the cell nucleus in mitochondrial biogenesis

A certain type of mutant yeast has a large deletion in the mitochondrial DNA, which leads to a complete cessation of protein synthesis in the mitochondria; as a result, these organelles are unable to perform their function. Since such mutants form small colonies when growing on a low-glucose medium, 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 are to a certain extent similar to ordinary mitochondria, have a normal outer membrane and an inner membrane with poorly developed cristae. Promitochondria contain many enzymes encoded by nuclear genes and synthesized on cytoplasmic ribosomes, 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 petite mutants always remains at the same level as in the wild type. This is due to the still little studied process of DNA amplification, as a result of which a DNA molecule is formed, consisting of tandem repeats of the same section and equal in size to a normal molecule. For example, the mitochondrial DNA of a petite mutant that retains 50% of the nucleotide sequence of wild-type DNA will consist of two repeats, whereas a molecule that retains only 0,1% wild-type genome will be built from 1000 copies of the remaining fragment. Thus, petite mutants can be used to obtain large quantities of specific sections of mitochondrial DNA, which can be said to be cloned by nature itself.

Although the biogenesis of organelles is controlled mainly by nuclear genes, the organelles themselves, according to some data, also have some kind of regulatory influence on the feedback principle; at least this is the case with mitochondria. If protein synthesis is blocked in the mitochondria of intact cells, then enzymes involved in the 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 the blocking agent. But, although the existence of some signal from mitochondria is beyond doubt, its nature is still unknown.

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). Firstly, 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 interfere with the development of these organelles. Such mutations are lethal in almost all other organisms. Secondly, yeast - simple single-celled eukaryotes - are easy to cultivate and biochemically study. Finally, yeast can reproduce in both the haploid and diploid phases, usually by asexual budding (asymmetric mitosis). But in yeast, the sexual process also occurs: from time to time, two haploid cells fuse to form a diploid zygote, which then either divides by mitosis or undergoes meiosis and again produces haploid cells. By experimentally controlling the alternation of asexual and sexual reproduction, one can learn a lot about the genes responsible for mitochondrial function. Using these methods, it is possible, in particular, to find out whether such genes are localized in nuclear DNA or in mitochondrial DNA, since mutations of mitochondrial genes are not inherited according to Mendel's laws, which govern the inheritance of nuclear genes.

Transport systems of mitochondria

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 the cell?

A partial answer was obtained by studying the transport of the small subunit (S) of the enzyme into the chloroplast stroma ribulose-1,5-bisphosphate-carboxymanholes. If mRNA isolated from the cytoplasm of a single cell seaweed Chlamydomonas or from pea leaves, introduced as a matrix into a protein synthesizing system in vitro, then one of the many resulting proteins will be bound by a specific anti-S antibody. The S protein synthesized in vitro is called ppo-S because it is approximately 50 amino acid residues larger than the regular S protein. When the pro-S protein is incubated with intact chloroplasts, it penetrates into 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 junction of the outer and inner membranes, and is then transferred into the stroma through transmembrane channels as a result of a process that requires energy expenditure.

Protein transport into mitochondria occurs 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 in the cytoplasm and selectively incorporated into mitochondria - just as occurs in the intact cell. In this case, the proteins of the outer and inner membranes, matrix and intermembrane space find their way to the corresponding compartment of the mitochondrion.

Many of the newly synthesized proteins destined for the inner membrane, matrix and intermembrane space have a leader peptide at their N-terminus, which is cleaved off during transport by a specific protease located in the matrix. The transport of proteins into these three mitochondrial compartments requires the energy of an electrochemical proton gradient created at the inner membrane. The mechanism of protein transfer for the outer membrane is different: in this case, neither energy 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 by the following mechanism: it is assumed that all proteins, except those destined for the outer membrane, are incorporated into the inner membrane as a result of a process requiring energy expenditure and occurring in places of contact between 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 anchored in the membrane or “pushed” into the matrix or into the intermembrane space.

The transfer of proteins across the membranes of mitochondria and chloroplasts is, in principle, similar to their transfer through the membranes of the endoplasmic reticulum. However, there are several important differences. First, when transported into the matrix or stroma, the protein passes through both the outer and inner membrane of the organelle, whereas 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 targeted release(vectorial discharge) - it begins when the protein has not yet completely left the ribosome (cotranslational import), and 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 the protein is directed to. Apparently, in many cases this sequence is cleaved from the precursor molecule after 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 in the polypeptide chain of the finished protein. Signal sequences are still poorly understood, but there are likely to 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, others to peroxisomes, and others to the endoplasmic reticulum. The complex processes that lead 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 obtain most of their lipids from the outside. In animal cells, phospholipids synthesized in the endoplasmic reticulum are transported to the outer membrane of mitochondria using special proteins and then incorporated into the inner membrane; this is believed to occur at the point of contact between two membranes. The main reaction of lipid biosynthesis, catalyzed by mitochondria themselves, is the conversion of phosphatidic acid to the phospholipid cardiolipin, which is found mainly in the inner mitochondrial membrane and accounts for 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 number of their genes in mitochondrial DNA, for which the nucleotide sequence is completely determined, vary greatly among different species of animals, plants, fungi and protozoa. The largest number of genes was found in the mitochondrial genome of flagellated protozoa 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 respiratory chain proteins, 22 for tRNA and two for rRNA (for the large ribosomal subunit 16S rRNA and for the small 12S rRNA). In plants and protozoa, unlike animals and most fungi, the mitochondrial genome also encodes some proteins that make up the ribosomes of these organelles. Key enzymes of template polynucleotide synthesis, such as DNA polymerase (replicating mitochondrial DNA) and RNA polymerase (transcribing the mitochondrial genome), are encrypted in the nucleus and synthesized on ribosomes in the cytoplasm. This fact indicates the relativity of mitochondrial autonomy in the complex hierarchy of a eukaryotic cell.

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

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

The size of the mitochondrial genome of different organisms ranges from less than 6 thousand nucleotide pairs in the falciparum plasmodium (in addition to two rRNA genes, it contains only three protein-coding genes) to hundreds of thousands of nucleotide pairs in land plants (for example, Arabidopsis thaliana from the cruciferous family 366924 nucleotide pairs). Moreover, 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 - 16569 nucleotide pairs, in pigs - 16350, in dolphins - 16330, in clawed frogs 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, plants, fungi and protozoa contain up to 80% non-coding sequences. The order of genes in mitochondrial genomes differs among species.

The high concentration of reactive oxygen species in mitochondria and a weak repair system increase the frequency of mtDNA mutations by an order of magnitude compared to nuclear DNA. Oxygen radicals cause specific substitutions C®T (cytosine deamination) and G®T (oxidative damage to guanine), as a result of which mtDNA is possibly 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.

Size and structure of DNA molecules in organelles

	Structure	Weight, million dalton	Notes
ohon Dria	Animals	Ring		Each individual species has all molecules of the same size
Higher ra sthenia	Ring	Varies	All studied species have circular DNA of different sizes, in which the total content of genetic information corresponds to a mass of 300 to 1000 million daltons, depending on the species
Mushrooms: Protozoa	Ring Ring Ring Linear
Chlorine payment stov	Seaweed	Ring Ring
Higher plants	Ring		In each individual species, molecules of only one were found

Relative amounts of DNA organelles in some cells and tissues

Organism	Fabric or cell type	Number of mol-l DNA/organel-	Number of organ- nell in cage	The share of DNA organelles in the entire Cell DNA, %
ohon Dria
	Line L cells
	Egg

Chlorine payment stov		Vegetative diploid cells
Corn

Functioning of the mitochondrial genome

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

Complementary" href="/text/category/komplementarij/" rel="bookmark">Complementary chains in mtDNA differ significantly in specific density, since they contain unequal amounts of “heavy” purine and “light” pyrimidine nucleotides. That’s what 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 is formed (from the English Displacement loop). This structure, visible in an electron micro- skop, consists of a double-stranded and a single-stranded (retracted part of the H-chain) sections. The double-stranded section is formed by part of the L-chain and a complementary newly synthesized DNA fragment with a length of 450-650 (depending on the type of organism) nucleotides, having 5"- the end of the ribonucleotide primer, which corresponds to the starting point of H-chain synthesis (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 to DNA synthesis enzymes only in a single-stranded state, and therefore, only in an unbraided state double helix during the synthesis of the H-chain. Thus, the daughter strands of mtDNA 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.

The 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) from them 12S rRNA and 16S rRNA are formed, which are involved in the formation of mitochondrial ribosomes, as well as phenylalanine and valine tRNA. The remaining tRNAs are excised from 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. Splicing (fusion) does not occur, since none of the mammalian mitochondrial genes contains 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-strand transcripts. tRNA and mRNA are formed as a result of processing from transcripts of both strands of DNA. tRNA genes are indicated in light green.

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

The leader and 3" non-coding 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 this phenomenon forced us to reconsider the idea of introns as “non-coding sequences.”

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

When studying the expression of mitochondrial genes Trypanosoma brucei discovered a surprising deviation from one of the main axioms 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 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 matrix for the synthesis of an additional subunit of the enzyme, the amino acid sequence in which has nothing in common with the sequence of viruses, fungi, plants and animals. The English researcher Burrell compared the structure of one of the mitochondrial genes of a calf with the sequence of amino acids 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) not only differs from the universal one, it is “ideal,” i.e., 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 - U, C), then they code for the same amino acid.” In the universal code there are two exceptions to this rule: the triplet AUA codes for isoleucine, and the codon AUG for methionine, while in the ideal mitochondrial code both of these triplets code for methionine; The UGG triplet encodes only tryptophan, and the UGA triplet encodes a stop codon. In the universal code, both deviations concern the fundamental aspects of protein synthesis: the AUG codon is the initiating one, and the stop codon UGA stops the synthesis of the polypeptide. The ideal code is not inherent in all described mitochondria, but none of them has a universal code. We can say that mitochondria speak different languages, but never the language of the nucleus.

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

Codon	Mitochondrial mammal code	Mitochondrial yeast code	“ Universal”

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 the ideal code of 64 triplets there are four stop codons, in the universal code there are three)? The fact is that during protein synthesis in mitochondria, codon-anticodon interactions are simplified - two out of three anticodon nucleotides are used for recognition. Thus, one tRNA recognizes all four members of the codon family, differing only in the third nucleotide. For example, leucine tRNA with the GAU anticodon stands on the ribosome opposite the codons TsU, TsUC, TsUA and Tsug, ensuring the unmistakable incorporation of leucine into the polypeptide chain. Two other 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 encoding one amino acid.

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? This issue 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. Ribosomal proteins, aminoacyl-tRNA synthetases, DNA and RNA polymerases, RNA processing and modification enzymes, etc. should be encoded here. Most of the studied proteins from mitochondria differ in amino acid sequence from their counterparts from other parts of the cell, and there is reason to believe that in these organs there are very few proteins that could be found elsewhere. This means that just to maintain the genetic system of mitochondria, the nuclear genome must contain several dozen additional genes. The reasons for this “wastefulness” are unclear, and the hope that the answer would be found in the nucleotide sequence of mitochondrial DNA did not materialize. It is difficult to imagine why proteins formed in mitochondria must necessarily be synthesized there, and not in the cytosol.

Typically, 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, studies of the ATP synthetase complex have shown that such an explanation is implausible. Although the individual protein subunits of ATP synthetase are highly conserved during evolution, the sites of their synthesis change. In chloroplasts, several fairly hydrophilic proteins, including four of the five subunits of the F1-ATPase part of the complex, are produced on ribosomes within the organelle. On the contrary, the mushroom Neurospora and in animal cells, a very hydrophobic component (subunit 9) of the membrane part of ATPase is synthesized on the ribosomes of the cytoplasm 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 using any hypothesis postulating certain evolutionary advantages of modern genetic systems of mitochondria and chloroplasts.

Considering all of the above, we can only assume that the mitochondrial genetic system represents an evolutionary dead end. Within the framework of the endosymbiotic hypothesis, this means that the process of transfer of endosymbiont genes into the host 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 are 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, who contribute equally to the gene pool of the offspring (although, after several generations separate offspring will often contain mitochondria of only one of the parent types). In contrast, in higher animals the egg contributes more cytoplasm to the zygote than the sperm, and in some animals the sperm may not contribute 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); and indeed, this has been confirmed by experiments. It turned out, for example, that when crossing rats of two laboratory strains with mitochondrial DNA slightly different in nucleotide sequence (types A and B), offspring are obtained containing

containing mitochondrial DNA of only the maternal type.

Cytoplasmic inheritance, unlike nuclear one, does not obey Mendel’s laws. This is due to the fact that in higher animals and plants, gametes from different sexes contain disparate amounts of mitochondria. So, in a mouse egg there are 90 thousand mitochondria, but in a sperm there are only four. It is obvious that in a fertilized egg the mitochondria are predominantly or only from the female individual, i.e., the inheritance of all mitochondrial genes is maternal. Genetic analysis of cytoplasmic inheritance 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. 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 inheritance of genes 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 merge 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 only cells with intact mitochondria can use; therefore, in the presence of chloramphenicol, only cells carrying the mutant gene can grow in 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 bud 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 the mitochondria, either mutant or wild type. Therefore, all offspring of such a cell will have genetically identical mitochondria. Such a random process, as a result of which diploid offspring is formed containing mitochondria of only one type, is called mitoticth segregationth. 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 the Mendelian inheritance of nuclear genes. Cytoplasmic gene transfer means that the genes being studied are located in mitochondria.

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

Some 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 involvement 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 mtDNA deletions were found in patients with severe muscle weakness, ataxia, deafness, and mental retardation, inherited in an autosomal dominant manner. Sexual dimorphism in clinical manifestations has been established ischemic heart disease, which is most likely due to the maternal effect - cytoplasmic inheritance. The development of gene therapy gives hope for correcting defects in mitochondrial genomes in the foreseeable future.

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

Mitochondrial diseases are an example of cytoplasmic heredity in humans, or more precisely, “organelle heredity”. This clarification should be made because the existence, at least in some organisms, of cytoplasmic hereditary determinant, not associated with cellular organelles - cytogens (Vechtomov, 1996).

Mitochondrial diseases are a heterogeneous group of diseases caused by genetic, structural, biochemical defects of mitochondria and impaired tissue respiration. To make a diagnosis of mitochondrial disease, complex genealogical, clinical, biochemical, morphological and genetic analysis. The main biochemical sign of mitochondrial pathology is the development of lactic acidosis; hyperlactic acidemia in combination with hyperpyruvatic acidemia is usually detected. The number of different options reached 120 forms. There is a stable increase in the concentration of lactic and pyruvic acids in the cerebrospinal fluid.

Mitochondrial diseases (MD) represent a significant problem for modern medicine. According to the methods of hereditary transmission, MDs include diseases inherited monogenically according to the Mendelian type, in which, due to 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 mainly transmitted to offspring through the maternal line.

Data from morphological studies indicating gross pathology of mitochondria: abnormal proliferation of mitochondria, polymorphism of mitochondria with disturbance of shape and size, disorganization 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 mutations in mitochondrial DNA

1.1.Diseases caused by mitochondrial DNA deletions

1.1.1.Cairns-Sayre syndrome

The disease manifests itself at the age of 4-18 years, progressive external ophthalmoplegia, retinitis pigmentosa, ataxia, intention tremor, atrioventricular heart block, increased protein levels in the cerebrospinal fluid more than 1 g/l, “ragged” red fibers in skeletal muscle biopsies

1.1.2.Pearson syndrome

The onset of the disease is from birth or in the first months of life, sometimes the development of encephalomyopathies, ataxia, dementia, progressive external ophthalmoplegia, hypoplastic anemia, violation of exocrine pancreatic function, progressive course

2 .Diseases caused by point mutations of mitochondrial DNA

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 sensorineural deafness, atrophy of the optic nerves and disorders of deep sensitivity, lactic acidosis, EEG examinations reveal generalized bathroom epileptic complexes, “ragged” 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, “ragged” red fibers in muscle biopsies, progressive course.

3 .Pathology associated with defects in intergenomic communication

3.1.Multiple mitochondrial DNA deletion syndromes

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

3.2.Mitochondrial DNA deletion syndrome

Autosomal recessive mode of inheritance

Clinical forms:

3.2.1.Fatal infantile

a) severe liver failure b) hepatopathy c) muscle hypotension

Debut in the neonatal period

3.2.2.Congenital myopathy

Severe muscle weakness, generalized hypotension, cardiomyopathy and convulsions, kidney damage, glycosuria, 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 caused by nuclear DNA mutations

4.1.Diseases associated with defects in the respiratory chain

4.1.1.Complex 1 deficiency (NADH:CoQ reductase)

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

4.1.2.Complex 2 deficiency (succinate-CoQ reductase)

Characterized by encephalomyopathy syndrome, progressive course, seizures, possible development of ptosis

4.1.3.Deficiency of complex 3 (CoQ-cytochrome C oxidoreductase)

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

4.1.4.Complex deficiency (cytochrome C oxidase)

4.1.4.1.Fatal infantile congenital lactic acidosis

Mitochondrial myopathy with renal failure or cardiomyopathy, onset in 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 life is possible

5 Menkes syndrome (trichopolyodystrophy)

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

6 . Mitochondrial encephalomyopathies

6.1.Leigh syndrome(subacute neurotizing encephalomyelopathy)

Appears after 6 months of life, muscle hypotonia, ataxia, nystagmus, pyramidal symptoms, ophthalmoplegia, optic nerve atrophy, the addition of cardiomyopathy and mild metabolic acidosis is often noted

6.2.Alpers syndrome(progressive sclerosing polydystrophy)

Degeneration of the gray matter of the brain in combination with liver cirrhosis, 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 chain enzymes

7 .Diseases associated with metabolic disorders of lactic and pyruvic acids

7.1. Pyruvate carboxylase deficiency Autosomal recessive type of inheritance, onset of the disease in the neonatal period, “flaccid child” symptom complex, convulsions resistant to therapy, high concentrations of ketone bodies in the blood, hyperammonemia, hyperlysinemia, decreased pyruvate carboxylase activity in skeletal muscles

7.2.Pyruvate dehydrogenase deficiency

Manifestation in the neonatal period, craniofacial dysmorphia, convulsions resistant to therapy, breathing and sucking disorders, “flaccid child” symptom complex, cerebral dysgynesia, severe acidosis with high levels of lactate and pyruvate

7.3.Decreased pyruvate dehydrogenase activity

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

7.4.Dihydrolipoyltransacetylase deficiency

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

7.5.Dihydrolipoyl dehydrogenase deficiency

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

8 .Diseases caused by defects in beta-oxidation of fatty acids

8.1.Long carbon chain acetyl-CoA dehydrogenase deficiency

Autosomal recessive type of inheritance, onset of the disease in the first months of life, metabolic crises with vomiting and diarrhea, “flaccid child” symptom complex, hypoglycemia, dicarboxylic aciduria, decreased activity of acetyl-CoA dehydrogenase of long-carbon-chain fatty acids

8.2. Medium-carbon chain acetyl-CoA dehydrogenase deficiency

Autosomal recessive type 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 activity of acetyl-CoA dehydrogenase of medium-carbon chain fatty acids

8.3. Short-chain fatty acid acetyl-CoA dehydrogenase deficiency

Autosomal recessive type of inheritance, different ages of onset of the disease, decreased exercise tolerance, metabolic crises with vomiting and diarrhea, muscle weakness and hypotension, increased urinary excretion of methylsuccinic 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 dysgynesia, severe hypoglycemia and acidosis, malignant course, decreased activity of all acetyl-CoA dehydrogenases of fatty acids,

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

8.5.Decreased activity of all fatty acid acetyl-CoA dehydrogenases

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

9 .Enzymopathies of the Krebs cycle

9.1.Fumarase deficiency

Autosomal recessive type of inheritance, onset of the disease in the neonatal or newborn period, microcephaly, generalized muscle weakness and hypotension, episodes of lethargy, rapidly progressing encephalopathy, 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, “flaccid child” symptom complex, episodes of lethargy, lactic acidosis, rapidly progressive course, decreased content of Krebs cycle enzymes in tissues

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

Carnitine palmitoyltransferase-1 deficiency, autosomal recessive type of inheritance, early onset of the disease, episodes of non-ketonemic hypoglycemic coma, hepatomegaly, hypertriglyceridemia and moderate hyperammonemia, decreased carnitine palmitoyltransferase-1 activity in fibroblasts and liver cells

9.5.Carnitine acylcarnitine translocase deficiency

Early onset of the disease, cardiovascular and respiratory disorders, the “flaccid child” symptom complex, episodes of lethargy and coma, increased concentrations of carnitine esters and long carbon chains against the background of a decrease in free carnitine in the blood serum, decreased activity of carnitine acylcarnitine translocase

9.6.Carnitine palmitoyl transferase-2 deficiency

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

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

Having analyzed 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 due to disturbances in 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, we can conclude that these proteins have a key role in the processes of cellular respiration, fatty acid oxidation and ATP synthesis, electron transfer in the electron transport system of the internal 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 control center for apoptosis...

The role of mitochondrial proteins has also been shown in the use of antibiotics that block mitochondrial synthesis. If human cells in tissue culture are treated with an antibiotic, such as tetracycline or chloramphenicol, their growth will stop after one or two divisions. This is due to inhibition of mitochondrial protein synthesis, leading to the appearance of defective mitochondria and, as a consequence, insufficient ATP formation. 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 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 within the tissue prevent certain drugs from entering 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 the rapidly dividing (and therefore most vulnerable) blood cell precursors.

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 and white blood cells), and long-term use of tetracycline can damage the intestinal epithelium. But in both cases, it is not yet entirely clear whether these side effects are caused by a blockade of mitochondrial biogenesis or by some other reason.

Conclusion

The structural and functional features of the mt genome are as follows. Firstly, it has been established that mtDNA is transmitted from the mother to all her

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

inheritance of mtDNA is probably associated with two circumstances: either the proportion of paternal mtDNA is so small (not transmitted through the paternal line)

more than one DNA molecule per 25 thousand maternal mtDNA), that they cannot be detected by existing methods, or after fertilization the replication of paternal mitochondria is blocked. Secondly, the absence of combinative variability - mtDNA belongs to only one of the parents, therefore, recombination events characteristic of nuclear DNA in meiosis are absent, and the nucleotide sequence changes from generation to generation only due to mutations. Thirdly, 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 mutation rate 10 times higher than in nuclear DNA. Fourthly, normal and mutant mtDNA can coexist simultaneously within the same cell - 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 above functions of the mt-genome have made the study of mtDNA nucleotide sequence variability an invaluable tool for doctors, forensic scientists, evolutionary biologists,

representatives of historical science in solving their specific problems.

Since 1988, when it was discovered that mtDNA gene mutations underlie mitochondrial myopathies (J. Y. Holt et al., 1988) and Leber’s hereditary optic neuropathy (D. C. Wallace, 1988), further systematic identification of mutations in the human mt genome led to the formation of the concept of mitochondrial diseases (MD). Currently, pathological mtDNA mutations have been discovered in every type of mitochondrial gene.

Bibliography

1. Skulachev, mitochondria and oxygen, Soros. education magazine

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 Francisco, Freeman, 1981.

5. Skulachev biological membranes. M., 1989.

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

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

8. , Scope of competence of the mitochondrial genome // Vestn. RAMS, 2001. ‹ 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 predisposition genes. St. Petersburg, 2000

11. , Mitochondrial genome. Novosibirsk, 1990.

12. // Soros. education magazine 1999. No. 10. P.11-17.

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15. // Soros. education magazine 2000. No. 1. P.32-36.

Kyiv National University named after. Taras Shevchenko

Department of Biology

Essay

on the topic of:

“The role of the maternal genome in the development of the offspring”

Withthereenta IVcourse

Department of Biochemistry

Frolova Artema

Kyiv 2004

Plan:

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

Symbiotic theory of the origin of mitochondria......2

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

Mitochondrial transport systems................................................................. ......7

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

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

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

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

Main article: Mitochondrial DNA

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 DNA localized in the nucleus. In total, mitochondrial DNA encodes 2 rRNA, 22 tRNA and 13 subunits of respiratory chain enzymes, which accounts for no more than half of the proteins found in it. In particular, under the control of the mitochondrial genome, seven ATP synthetase subunits, three cytochrome oxidase subunits, and one ubiquinol-cytochrome subunit are encoded. With-reductase. In this case, all proteins except one, two ribosomal and six transfer RNAs are transcribed from the heavier (outer) DNA chain, and 14 other tRNAs and one protein are transcribed from the lighter (internal) chain.

Against this background, the plant mitochondrial genome is much larger and can reach 370,000 nucleotide pairs, which is approximately 20 times larger than the human mitochondrial genome described above. The number of genes here is also approximately 7 times greater, which is accompanied by the appearance in plant mitochondria of additional electron transport pathways 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 into two by constriction, the formation of which begins from a circular groove on the inner mitochondrial membrane. A detailed study of the nucleotide sequence of the mitochondrial genome has revealed that deviations from the universal genetic code are common in the mitochondria of animals and fungi. 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 a stop codon in the standard code, encodes the amino acid methionine. As for plant mitochondria, they apparently use a universal genetic code. Another feature of mitochondria is the peculiarity of tRNA codon recognition, which consists in the fact that one such molecule is capable of recognizing not one, but three or four codons at once. This feature reduces the importance of the third nucleotide in the codon and leads to the fact that mitochondria require less variety of tRNA types. In this case, only 22 different tRNAs turn out to be sufficient.

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

Mitochondrial proteins[edit | edit source text]

The number of proteins translated from mitochondrial mRNA that form the subunits of large enzyme complexes is limited. A significant portion of proteins are encoded in the nucleus and synthesized on cytoplasmic 80S ribosomes. In particular, this is how some proteins are formed - electron carriers, mitochondrial translocases, components of protein transport into mitochondria, as well as factors necessary for transcription, translation and replication of mitochondrial DNA. Moreover, such proteins at their N-terminus have special signal peptides, the size of which varies from 12 to 80 amino acid residues. These areas form amphiphilic curls and provide specific contact of proteins with the binding domains of mitochondrial recognition receptors localized on the outer membrane. These proteins are transported to the outer mitochondrial membrane in a partially unfolded state in association with chaperone proteins (in particular, hsp70). After transfer through the outer and inner membranes at the places of their contacts, the proteins entering the mitochondrion again contact chaperones, but of their own mitochondrial origin, which pick up the membrane-crossing protein, promote its retraction into the mitochondrion, and also control the process of correct folding of the polypeptide chain. Most chaperones have ATPase activity, as a result of which both the transport of proteins into the mitochondrion and the formation of their functionally active forms are energy-dependent processes.