Transfer of mitochondrial DNA. On the importance of studying 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, seven ATP synthetase subunits, three cytochrome oxidase subunits, and one ubiquinol-cytochrome subunit are encoded under the control of the mitochondrial genome. With-reductase. In this case, all proteins except one, two ribosomal and six tRNAs 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. Detailed Study nucleotide sequence The mitochondrial genome allowed us to establish that deviations from the universal genetic code are not uncommon in the mitochondria of animals and fungi. Thus, in human mitochondria, the TAT codon replaces isoleucine in standard code codes for the amino acid methionine, the codons TCT and TCC, which usually code for arginine, are stop codons, and the codon AST, which is a stop codon in the standard code, codes for 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, the peculiarity 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.

Properties and functions of DNA.

DNA, or deoxyribonucleic acid, is the basic hereditary material present in all cells of the body and primarily mediates cell functions, growth, reproduction and death. The structure of DNA, called the double-stranded helical structure, was first described by Watson and Crick in 1953.

From then on, enormous progress was made in the synthesis, sequencing and manipulation of DNA. DNA these days can be virtualized or analyzed for details and even genes can be inserted to cause changes in DNA function and structure.

The main purpose of hereditary material is to store hereditary information on the basis of which the phenotype is formed. Most of the characteristics and properties of the body are determined by the synthesis of proteins that perform various functions. Thus, the hereditary material must contain information about 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. Therefore, in molecules nucleic acids The amino acid composition of proteins must be encoded.
Back in the early 50s, it was suggested that there was a way to record 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 the DNA molecule. To encrypt more than 20 amino acids required amount combinations are provided only by a triplet code, i.e., a code that includes three adjacent nucleotides. In this case, the number of combinations of four nitrogenous bases in threes is 41 = 64. The assumption about the triplet nature of the genetic code later received experimental confirmation, and during the period from 1961 to 1964, a code was discovered with the help of which the order of amino acids is written in nucleic acid molecules peptide.
From the table 6 shows 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, series, etc.). Several triplets do not code for 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.
Study of the genetic code in living organisms with different levels organization showed the universality of this mechanism for recording information in living nature.
Thus, research from the mid-20th century revealed the mechanism for recording hereditary information in nucleic acid molecules using a biological code, which is characterized by the following properties: a) tripletity - amino acids are encrypted by triplets of nucleotides - codons; b) specificity - each triplet encodes only a specific 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 by more than one triplet; e) non-overlapping - information is read sequentially triplet by triplet: AAGCTTCAGCCAT.

In addition to recording and storing biological information, the function of hereditary material is its reproduction and transmission to a new generation in the process of reproduction of cells and organisms. This function of hereditary material is carried out by DNA molecules in the process of its reduplication, i.e. absolutely accurate 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 various types RNA, which ensures the translation process, i.e., the assembly of a protein molecule, occurring in the cytoplasm based on information received from the nucleus (see 2.4). During the implementation 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 during the synthesis of mRNA - 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 of a peptide chain of amino acids on ribosomes based on the information recorded in the mRNA molecule, with the participation of transport tRNAs - translation (see 2.4).
4. Formation of secondary, tertiary and quaternary protein structures, which corresponds to the formation of a functioning protein (simple sign).
5. Formation of a complex trait as a result of the participation of several gene products (enzyme proteins or other proteins) in biochemical processes.

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

Denaturation changes some of the physical properties of DNA, such as its optical density. Nitrogen 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 caused by the interaction of the 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. the optical density shifts towards the value characteristic of free bases. Thus, DNA denaturation can be observed by changes in its optical density.

When DNA is heated, the average temperature of the range at which DNA strands separate is called the melting point and is designated T pl. In solution T pl usually lies in the range of 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 A-T pairs, having two hydrogen bonds, therefore, with increasing G-C-nap content, the T value pl increases. DNA, 40% consisting 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 base composition) 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 makes it possible to reduce T pl, up to 40 °C.

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

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 regenerated. If together. anneal single-stranded DNA originating from different sources, the formation of a double-stranded DNA structure is called hybridization.

Related information.

Historically, the first study of this kind was conducted using mitochondrial DNA. Scientists took a sample from the natives of Africa, Asia, Europe, and America, and in this initially small sample they compared the mitochondrial DNA of different individuals with each other. They found that mitochondrial DNA diversity 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 could have mutationally descended from which. Of all the people whose DNA was tested, it was Africans who found much greater variability. Mitochondrial DNA types 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 DNA remains are found that are not characteristic of the mutations of European man.

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

Thanks to studies of 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 timing of the appearance of the mitochondrial foremother of humanity was established through a comparative study of the mitochondrial DNA of chimpanzees and modern man. Knowing the rate of mutational divergence - 2-4% per million years - we can determine the time of separation of the two branches, chimpanzees and modern humans. This happened approximately 5 - 7 million years ago. In this case, the rate of mutational divergence is considered constant.

Mitochondrial Eve

When people talk about mitochondrial Eve, they don't mean an individual. They talk about the emergence through evolution of an entire population of individuals with similar characteristics. It is believed that Mitochondrial Eve lived during a period of sharp decline in the number of our ancestors, to approximately ten thousand individuals.

Origin of races

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

Comparison of mitochondrial DNA of Neandarthals and modern humans

Additional information about human origins was obtained by comparing the genetic texts of the mitochondrial DNA of Neanderthals and modern humans. Scientists were able to read the genetic texts of mitochondrial DNA from the bone remains of two Neanderthals. The skeletal remains of the first Neanderthal 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 you take a piece of DNA, then out of 370 nucleotides, 27 differ. And if you compare the genetic texts of a modern person, his mitochondrial DNA, you will find a difference in only eight nucleotides. It is believed that Neanderthal and modern man are completely separate branches, the evolution of each of them proceeded independently of each other.

By studying the differences in the genetic texts of the mitochondrial DNA of Neanderthals and modern humans, the date of separation of these two branches was established. This happened approximately 500 thousand years ago, and approximately 300 thousand years ago their final separation occurred. It is believed that Neanderthals settled throughout Europe and Asia and were displaced by modern humans, who emerged from Africa 200 thousand years later. And finally, approximately 28 - 35 thousand years ago, Neanderthals became extinct. Why this happened, in general, is not yet clear. Maybe they couldn’t stand the competition with a modern type of person, or maybe there were other reasons for this.

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All information about the structure of the human body and its predisposition to diseases is encrypted in the form of DNA molecules. The main information is located in the cell nuclei. However, 5% of DNA is localized in mitochondria.

What are mitochondria called?

Mitochondria are cellular organelles of eukaryotes that are needed in order to convert the energy contained in nutrients into compounds that can be taken up by cells. Therefore, they are often called “energy stations”, because without them the existence of the body is impossible.
These organelles acquired their own genetic information due to the fact that they were previously bacteria. After they entered the cells of the host organism, they were unable to retain their genome, while they transferred part of their own genome to the cell nucleus of the host organism. Therefore, now their DNA (mtDNA) contains only a part, namely 37 genes, of the original amount. Mainly, they encrypt the mechanism of transformation of glucose into compounds - carbon dioxide and water with the production of energy (ATP and NADP), without which the existence of the host organism is impossible.

What is unique about mtDNA?

The main property inherent in mitochondrial DNA is that it can be inherited only through the mother's line. In this case, all children (men or women) can receive mitochondria from the egg. This happens due to the fact that female eggs contain a higher number of these organelles (up to 1000 times) than male sperm. As a result, the daughter organism receives them only from its mother. Therefore, their inheritance from the paternal cell is completely impossible.
It is known that mitochondrial genes were passed on to us from the distant past - from our promother - “mitochondrial Eve”, who is the common ancestor of all people on the planet on the maternal side. Therefore, these molecules are considered the most ideal object for genetic examinations to establish maternal kinship.

How is kinship determined?

Mitochondrial genes have many point mutations, making them highly variable. This allows us to establish kinship. During genetic examination, using special genetic analyzers - sequencers, individual point nucleotide changes in the genotype, their similarity or difference, are determined. In people who are not related on their mother's side, the mitochondrial genomes differ significantly.
Determining kinship is possible thanks to the amazing characteristics of the mitochondrial genotype:
they are not subject to recombination, so molecules change only through the process of mutation, which can occur over a millennium;
possibility of isolation from any biological materials;
if there is a lack of biomaterial or degradation of the nuclear genome, mtDNA can become the only source for analysis due to the huge number of its copies;
due to large quantities mutations compared to the nuclear genes of cells is achieved high accuracy when analyzing genetic material.

What can be determined through genetic testing?

Genetic testing of mtDNA will help in diagnosing the following cases.
1. To establish kinship between people on the mother’s side: between a grandfather (or grandmother) and a grandson, a brother and sister, an uncle (or aunt) and a nephew.
2. When analyzing a small amount of biomaterial. After all, each cell contains mtDNA in significant quantities (100 - 10,000), while nuclear DNA contains only 2 copies for each 23 chromosomes.
3. When identifying ancient biomaterial – a shelf life of more than a thousand years. It is thanks to this property that scientists were able to identify genetic material from the remains of members of the Romanov family.
4. In the absence of other material, even one hair contains a significant amount of mtDNA.
5. When determining the belonging of genes to the genealogical branches of humanity (African, American, Middle Eastern, European haplogroup and others), thanks to which it is possible to determine the origin of a person.

Mitochondrial diseases and their diagnosis

Mitochondrial diseases manifest themselves mainly due to defects in the mtDNA of cells associated with a significant susceptibility of these organelles to mutations. Today there are already about 400 diseases associated with their defects.
Normally, each cell can include both normal mitochondria and those with certain disorders. Often, signs of the disease do not manifest themselves at all. However, when the process of energy synthesis weakens, the manifestation of such diseases is observed in them. These diseases are primarily associated with muscle or nervous systems. As a rule, with such diseases there is a late onset clinical manifestations. The incidence of these diseases is 1:200 people. It is known that the presence of mitochondrial mutations can cause nephrotic syndrome during pregnancy and even sudden death of the infant. Therefore, researchers are making active attempts to solve these problems associated with the treatment and transmission of genetic diseases of this type from mothers to children.

How is aging related to mitochondria?

Reorganization of the genome of these organelles was also discovered when analyzing the mechanism of aging of the body. Researchers at Hopkins University published results from monitoring the blood levels of 16,000 elderly American people, demonstrating that the decrease in the amount of mtDNA was directly related to the age of the patients.

Most of the issues considered today have become the basis of a new science - “mitochondrial medicine”, which was formed as a separate direction in the 20th century. Prediction and treatment of diseases associated with mitochondrial genome disorders, genetic diagnostics are its primary tasks.

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.

What is mitochondrial DNA?

Mitochondrial DNA (mtDNA) is DNA located in mitochondria, cellular organelles inside eukaryotic cells that convert chemical energy from food into a form that cells can use - adenosine triphosphate (ATP). Mitochondrial DNA represents only a small part of the DNA in a eukaryotic cell; Most DNA can be found in the cell nucleus, in plants and algae, and in plastids such as chloroplasts.

In humans, the 16,569 base pairs of mitochondrial DNA encode just 37 genes. Human mitochondrial DNA was the first significant portion of the human genome to be sequenced. In most species, including humans, mtDNA is inherited only from the mother.

Because animal mtDNA evolves faster than nuclear genetic markers, it represents the basis of phylogenetics and evolutionary biology. This has become an important point in anthropology and biogeography, as it allows one to study the interrelationships of populations.

Hypotheses for the origin of mitochondria

Nuclear and mitochondrial DNA are believed to have different evolutionary origins, with mtDNA derived from the circular genomes of bacteria that were absorbed by the early ancestors of modern eukaryotic cells. This theory is called the endosymbiotic theory. It is estimated that each mitochondrion contains copies of 2-10 mtDNA. In cells existing organisms the vast majority of proteins present in mitochondria (numbering about 1,500 different types in mammals) are encoded by nuclear DNA, but the genes for some, if not most, of these are thought to be originally bacterial, having since been transferred to the eukaryotic nucleus during evolution.

The reasons why mitochondria retain certain genes are discussed. The existence of genome-less organelles in some species of mitochondrial origin suggests that complete gene loss is possible, and the transfer of mitochondrial genes to the nucleus has a number of advantages. The difficulty of orienting remotely produced hydrophobic protein products in mitochondria is one hypothesis for why some genes are retained in mtDNA. Co-localization for redox regulation is another theory, citing the desirability of localized control of mitochondrial machinery. Recent analysis of a wide range of mitochondrial genomes suggests that both of these functions may dictate mitochondrial gene retention.

Genetic examination of mtDNA

In most multicellular organisms, mtDNA is inherited from the mother (maternal lineage). Mechanisms for this include simple dilution (an egg contains an average of 200,000 mtDNA molecules, whereas healthy human sperm contains an average of 5 molecules), degradation of sperm mtDNA in the male reproductive tract, in the fertilized egg, and, in at least a few organisms, failure The mtDNA of the sperm penetrates into the egg. Whatever the mechanism, it is unipolar inheritance - inheritance of mtDNA, which occurs in most animals, plants and fungi.

Maternal inheritance

In sexual reproduction, mitochondria are usually inherited exclusively from the mother; mitochondria in mammalian sperm are usually destroyed by the egg after fertilization. Additionally, most mitochondria are present at the base of the sperm tail, which is used for sperm cell movement; sometimes the tail is lost during fertilization. In 1999, it was reported that paternal sperm mitochondria (containing mtDNA) are marked by ubiquitin for subsequent destruction within the embryo. Some in vitro fertilization methods, particularly sperm injection into the oocyte, may interfere with this.

The fact that mitochondrial DNA is inherited through the maternal line allows genealogical researchers to trace the maternal line far back in time. (Y-chromosomal DNA is paternally inherited, used in a similar way to determine patrilineal history.) This is usually done on a person's mitochondrial DNA by sequencing the hypervariable control region (HVR1 or HVR2), and sometimes the entire mitochondrial DNA molecule as a DNA genealogy test. For example, HVR1 consists of approximately 440 base pairs. These 440 pairs are then compared to control areas of other individuals (or specific individuals or subjects in the database) to determine maternal lineage. The most common comparison is with the Revised Cambridge Reference Sequence. Vilà et al. published studies on the matrilineal similarity of domestic dogs and wolves. The concept of Mitochondrial Eve is based on the same type of analysis, attempts to discover the origins of humanity, traces the origin back in time.

mtDNA is highly conserved, and its relatively slow mutation rates (compared to other regions of DNA such as microsatellites) make it useful for studying evolutionary relationships—the phylogeny of organisms. Biologists can determine and then compare mtDNA sequences from different types and use comparisons to construct evolutionary trees for the species studied. However, due to the slow mutation rates it experiences, it is often difficult to distinguish closely related species to any extent, so other methods of analysis must be used.

Mitochondrial DNA mutations

Individuals undergoing unidirectional inheritance and little or no recombination can be expected to undergo Müllerian ratchet, the accumulation of deleterious mutations until functionality is lost. Animal mitochondrial populations avoid this accumulation due to a developmental process known as the mtDNA bottleneck. The bottleneck uses stochastic processes in the cell to increase cell-to-cell variability in mutant load as the organism develops, such that one egg cell with some proportion of mutant mtDNA creates an embryo in which different cells have different mutant loads. The cellular level can then be targeted to remove these cells with more mutant mtDNA, resulting in stabilization or reduction of the mutant load between generations. The mechanism underlying the bottleneck is discussed with recent mathematical and experimental metastasis and provides evidence for a combination of random partitioning of mtDNA into cell divisions and random turnover of mtDNA molecules within the cell.

Paternal inheritance

Double unidirectional inheritance of mtDNA is observed in bivalves. In these species, females have only one type of mtDNA (F), whereas males have type F mtDNA in their somatic cells, but M type mtDNA (which can be up to 30% divergent) in germline cells. Maternally inherited mitochondria have additionally been reported in some insects such as fruit flies, bees, and periodical cicadas.

Male mitochondrial inheritance was recently discovered in Plymouth Rock chickens. Evidence supports rare cases of male mitochondrial inheritance in some mammals. In particular, documented cases exist for mice where male-derived mitochondria were subsequently rejected. In addition, it was found in sheep, as well as in cloned large cattle. It was once discovered in a man's body.

Although many of these cases involve embryo cloning or subsequent rejection of paternal mitochondria, others document inheritance and persistence in vivo in vitro.

Mitochondrial donation

IVF, known as mitochondrial donation or mitochondrial replacement therapy (MRT), results in offspring containing mtDNA from female donors and nuclear DNA from the mother and father. In the spindle transfer procedure, an egg nucleus is introduced into the cytoplasm of an egg from a female donor that has had the nucleus removed but still contains the mtDNA of the female donor. The composite egg is then fertilized by the man's sperm. This procedure is used when a woman with genetically defective mitochondria wants to produce offspring with healthy mitochondria. The first known child to be born as a result of mitochondrial donation was a boy born to a Jordanian couple in Mexico on April 6, 2016.

Mitochondrial DNA structure

In most multicellular organisms, mtDNA - or the mitogenome - is organized as round, circularly closed, double-stranded DNA. But in many unicellular organisms (for example, tetrahymena or the green alga Chlamydomonas reinhardtii) and in rare cases in multicellular organisms (for example, some species of cnidarians), mtDNA is found as linearly organized DNA. Most of these linear mtDNAs possess telomerase-independent telomeres (i.e., the ends of the linear DNA) with different modes of replication, which have made them interesting subjects of study, since many of these single-celled organisms with linear mtDNA are known pathogens.

For human mitochondrial DNA (and probably for metazoans), 100-10,000 individual copies of mtDNA are typically present in a somatic cell (eggs and sperm are exceptions). In mammals, each double-stranded circular mtDNA molecule consists of 15,000-17,000 base pairs. The two strands of mtDNA differ in their nucleotide content, the guanide-rich strand is called the heavy chain (or H-strand) and the cynosine-rich strand is called the light chain (or L-strand). The heavy chain encodes 28 genes and the light chain encodes 9 genes, for a total of 37 genes. Of the 37 genes, 13 are for proteins (polypeptides), 22 are for transferring RNA (tRNA), and two are for small and large subunits of ribosomal RNA (rRNA). The human mitogenome contains overlapping genes (ATP8 and ATP6, and ND4L and ND4: see Human genome map of mitochondria), which is rare in animal genomes. The 37-gene pattern is also found among most metazoans, although, in some cases, one or more of these genes are missing and the range of mtDNA sizes is greater. Even greater variation in the content and size of mtDNA genes exists among fungi and plants, although there appears to be a core subset of genes that is present in all eukaryotes (except for the few that have no mitochondria at all). Some plant species have huge mtDNA (as much as 2,500,000 base pairs per mtDNA molecule), but surprisingly, even these huge mtDNA contain the same number and types of genes as related plants with much smaller mtDNA.

The cucumber (Cucumis Sativus) mitochondrial genome consists of three circular chromosomes (length 1556, 84 and 45 kb), which are completely or largely autonomous with respect to their replication.

Six major genome types are found in mitochondrial genomes. These types of genomes were classified by "Kolesnikov and Gerasimov (2012)" and differ in various ways, such as circular versus linear genome, genome size, presence of introns or plasmid-like structures, and whether the genetic material is a distinct molecule, a collection of homogeneous or heterogeneous molecules.

Decoding the animal genome

In animal cells, there is only one type of mitochondrial genome. This genome contains a single circular molecule between 11-28 kbp of genetic material (type 1).

Decoding the plant genome

There are three various types genome contained in plants and fungi. The first type is a circular genome that has introns (type 2) ranging from 19 to 1000 kbp in length. The second type of genome is a circular genome (about 20-1000 kbp), which also has a plasmid structure (1kb) (type 3). The final type of genome that can be found in plants and fungi is the linear genome, consisting of homogeneous DNA molecules (type 5).

Decoding the protist genome

Protists contain the most diverse mitochondrial genomes, which include five different types. Type 2, type 3 and type 5, mentioned in plant and fungal genomes, also exist in some protozoa, as well as in two unique genome types. The first of these is a heterogeneous collection of circular DNA molecules (type 4), and the final genome type found in protists is a heterogeneous collection of linear molecules (type 6). Genome types 4 and 6 range from 1 to 200 kb.

Endosymbiotic gene transfer, the process of genes encoded in the mitochondrial genome being carried primarily by the cell's genome, likely explains why more complex organisms, such as humans, have smaller mitochondrial genomes than simpler organisms, such as protozoa.

Mitochondrial DNA replication

Mitochondrial DNA is replicated by the DNA polymerase gamma complex, which consists of a 140 kDa catalytic DNA polymerase encoded by the POLG gene and two 55 kDa accessory subunits encoded by the POLG2 gene. The replication apparatus is formed by DNA polymerase, TWINKLE and mitochondrial SSB proteins. TWINKLE is a helicase that unwinds short stretches of dsDNA in the 5" to 3" direction.

During embryogenesis, mtDNA replication is tightly regulated from the fertilized oocyte through the preimplantation embryo. Effectively reducing the number of cells in each cell, mtDNA plays a role in the mitochondrial bottleneck, which exploits cell-to-cell variability to improve the inheritance of damaging mutations. At the blastocyte stage, the onset of mtDNA replication is specific to trophtocoder cells. In contrast, cells of the inner cell mass restrict mtDNA replication until they receive signals to differentiate into specific cell types.

Mitochondrial DNA transcription

In animal mitochondria, each strand of DNA is continuously transcribed and produces a polycistronic RNA molecule. There are tRNAs present between most (but not all) protein-coding regions (see Map of the Human Mitochondria Genome). During transcription, tRNA acquires a characteristic L-form, which is recognized and cleaved by specific enzymes. When mitochondrial RNA is processed, individual fragments of mRNA, rRNA, and tRNA are released from the primary transcript. Thus, folded tRNAs act as minor punctuations.

Mitochondrial diseases

The concept that mtDNA is particularly susceptible to reactive oxygen species generated by the respiratory chain due to its proximity remains controversial. mtDNA does not accumulate more oxidative base than nuclear DNA. It has been reported that at least some types of oxidative DNA damage are repaired more efficiently in mitochondria than in the nucleus. mtDNA is packaged with proteins that appear to be as protective as nuclear chromatin proteins. Moreover, mitochondria have evolved a unique mechanism that maintains mtDNA integrity by degrading excessively damaged genomes followed by replication of intact/repaired mtDNA. This mechanism is absent in the nucleus and is activated by several copies of mtDNA present in mitochondria. The result of a mutation in mtDNA can be a change in the coding instructions for certain proteins, which can affect the metabolism and/or fitness of the organism.

Mitochondrial DNA mutations can lead to a number of diseases, including exercise intolerance and Kearns-Sayre syndrome (KSS), which causes a person to lose full function of heart, eye and muscle movements. Some evidence suggests that they may be a significant contributor to the aging process and age-related pathologies. Specifically, in the context of disease, the proportion of mutant mtDNA molecules in a cell is called heteroplasm. The distributions of heteroplasm within and between cells dictate the onset and severity of disease and are influenced by complex stochastic processes within the cell and during development.

Mutations in mitochondrial tRNAs may be responsible for severe diseases such as MELAS and MERRF syndromes.

Mutations in nuclear genes encoding proteins that use mitochondria may also contribute mitochondrial diseases. These diseases do not follow mitochondrial inheritance patterns, but instead follow Mendelian patterns of inheritance.

IN Lately mutations in mtDNA have been used to help diagnose prostate cancer in biopsy-negative patients.

Mechanism of aging

Although the idea is controversial, some evidence suggests a link between aging and mitochondrial dysfunction in the genome. Essentially, mutations in mtDNA disrupt the careful balance of reactive oxygen production (ROS) and enzymatic ROS production (by enzymes such as superoxide dismutase, catalase, glutathione peroxidase, and others). However, some mutations that increase ROS production (for example, by reducing antioxidant defenses) in worms increase, rather than decrease, their longevity. In addition, nude moth rats, rodents the size of mice, live approximately eight times longer than mice, despite having decreased antioxidant defenses and increased oxidative damage to biomolecules compared to mice.

At one point there was believed to be a virtuous feedback loop at work ("Vicious Cycle"); as mitochondrial DNA accumulates genetic damage caused by free radicals, mitochondria lose function and release free radicals in the cytosol. Decreased mitochondrial function reduces overall metabolic efficiency. However, this concept was finally refuted when it was demonstrated that mice genetically modified to accumulate mtDNA mutations at an increased rate age prematurely, but their tissues do not produce more ROS, as predicted by the "Vicious Cycle" hypothesis. Supporting the link between longevity and mitochondrial DNA, some studies have found correlations between the biochemical properties of mitochondrial DNA and species longevity. Extensive research is being conducted to further explore this connection and anti-aging treatments. Currently gene therapy and nutraceutical supplements are popular areas of current research. Bjelakovic et al. analyzed the results of 78 studies between 1977 and 2012, involving a total of 296,707 participants, and concluded that antioxidant supplements did not reduce mortality from any cause or prolong life expectancy, while some of these, such as beta-carotene, vitamin E and higher doses of vitamin A, may actually increase mortality.

Deletion breakpoints often occur within or adjacent to regions exhibiting non-canonical (non-B) conformations, namely hairpin, cross, and clover-like elements. In addition, there is evidence that helical distortion curvilinear regions and long G-tetrads are involved in detecting instability events. In addition, higher density points were consistently observed in regions with GC skew and in close proximity to the degenerate sequence fragment YMMYMNNMMHM.

How is mitochondrial DNA different from nuclear DNA?

Unlike nuclear DNA, which is inherited from both parents and in which genes are rearranged through the process of recombination, there is usually no change in mtDNA from parent to offspring. Although mtDNA also recombines, it does so with copies of itself within the same mitochondrion. Because of this, the mutation rate of animal mtDNA is higher than that of nuclear DNA. mtDNA is a powerful tool for tracing matrilineage and has been used in this role to trace the ancestry of many species hundreds of generations ago.

The rapid rate of mutation (in animals) makes mtDNA useful for assessing the genetic relationships of individuals or groups within a species, and for identifying and quantifying phylogenies (evolutionary relationships) among different species. To do this, biologists determine and then compare the mtDNA sequence from different individuals or species. Data from the comparisons are used to construct a network of relationships between sequences that provide an estimate of the relationships between the individuals or species from which the mtDNA was taken. mtDNA can be used to assess relationships between closely related and distant species. Due to the high frequency of mtDNA mutations in animals, 3rd position codons change relatively quickly, and thus provide information about genetic distances between closely related individuals or species. On the other hand, the substitution rate of mt proteins is very low, so amino acid changes accumulate slowly (with corresponding slow changes in 1st and 2nd codon positions) and thus they provide information about the genetic distances of distant relatives. Statistical models that consider substitution rates among codon positions separately can therefore be used to simultaneously estimate phylogenies that contain both closely related and distant species.

History of the discovery of mtDNA

Mitochondrial DNA was discovered in the 1960s by Margit M. K. Nas and Silvan Nas using electron microscopy as DNase-sensitive strands within mitochondria, and by Ellen Hasbrunner, Hans Tappi and Gottfried Schatz from biochemical analyzes on highly purified mitochondrial fractions.

Mitochondrial DNA was first recognized in 1996 during Tennessee v. Paul Ware. In 1998, in the court case Commonwealth of Pennsylvania v. Patricia Lynn Rorrer, mitochondrial DNA was admitted into evidence for the first time in the State of Pennsylvania. The case was featured in Episode 55 of Season 5 of the True Drama Forensic Court Case Series (Season 5).

Mitochondrial DNA was first recognized in California during the successful prosecution of David Westerfield for the 2002 kidnapping and murder of 7-year-old Danielle van Dam in San Diego, and has been used to identify both humans and dogs. This was the first test in the US to resolve canine DNA.

mtDNA databases

Several specialized databases have been created to collect mitochondrial genome sequences and other information. Although most of them focus on sequence data, some include phylogenetic or functional information.

• MitoSatPlant: microsatellite database of mitochondrial viridiplants.

• MitoBreak: Mitochondrial DNA Breakpoint Database.

• MitoFish and MitoAnnotator: fish mitochondrial genome database. See also Cawthorn et al.

• MitoZoa 2.0: database for comparative and evolutionary analysis of mitochondrial genomes (no longer available)

• InterMitoBase: an annotated database and protein-protein interaction analysis platform for human mitochondria (last updated in 2010, but still not available)

• Mitome: database for comparative mitochondrial genomics in metazoans (no longer available)

• MitoRes: a resource for nuclear-encoded mitochondrial genes and their products in metazoans (no longer updated)

There are several specialized databases that report polymorphisms and mutations in human mitochondrial DNA along with assessments of their pathogenicity.

• MITOMAP: a compendium of polymorphisms and mutations in human mitochondrial DNA.

• MitImpact: Collection of predicted pathogenicity predictions for all nucleotide changes that cause nonsynonymous substitutions in human mitochondrial protein-coding genes.