Weed out mutants

Stabilizing selection in mammals copes with mutations in mitochondria in just two generationsElena Naimark, Elementy.ru 

Mitochondria are the energy stations of the cell. They have their own set of genes in which information about the enzymes involved in cellular respiration is recorded. Significant mutations in mitochondrial genes can lead to cell and body death. But the cell is somehow freed from mutant mitochondria. Drawing from the website www.ndpteachers.orgScientists were able to trace the selection process of mitochondria in mouse cells.

It turned out that the organism of a higher animal is able to get rid of mutant mitochondria extremely effectively: they disappear after 2-6 generations. These data suggest that the selection of normal mitochondria does not occur by eliminating whole organisms with reduced fitness, but at the level of oocytes (female germ cells) or at the subcellular level. Most likely, the mechanism of stabilizing selection of mitochondria occurs at the level of interaction of eukaryotic cell components. At the same time, the rates of stabilizing selection of mitochondrial tRNA genes and protein-coding genes turned out to be different. A different mechanism of stabilizing selection for protein and tRNA genes is also assumed.

The hypothesis of the origin of the eukaryotic cell as a result of the symbiosis of prokaryotic cells of several types, which shocked the scientific community Lynn Margulis 40 years ago, has already become generally accepted. Biologists are now asking questions not about the probability of this hypothesis, but about the ways of evolution of individual components of this symbiotic system.

The most likely ancestor of mitochondria today is considered to be free-living proteobacteria. After merging with the host cell, proteobacteria took over the functions of energy supply to the cell, and left other functions to other cellular elements. As a result, mitochondria refused to recombine (exchange of genes with their own kind), leaving themselves with a severely truncated genome. The mitochondrial genome in animals underwent the greatest reduction. It contains only information about some enzymes that serve oxidative phosphorylation (cellular respiration), as well as genes of some functional RNAs (transport, ribosomal).

It is clear that the viability of the cell depends on the proper functioning of the mitochondrial genes. Once one of the enzymes acquires a harmful mutation, the energy supply of the cell will be disrupted. Mitochondria deprived of life-saving recombination have no way to get rid of mutations by exchanging genes with other, "healthy" mitochondria. One could assume that mitochondria are extremely stable systems, and the rate of mutation in them is extremely low. However, surprisingly, it turned out that the rate of mutation in the mitochondrial genome is even higher than in the nuclear one. Theoretically, it is clear that the cell somehow gets rid of mutant mitochondria, somehow stabilizing (cleansing) selection works, screening out harmful mutations. But how does it work?

It should be well understood that selection may not work as straightforwardly as it appears in educational schemes: a harmful mutation has appeared, therefore a poorly viable individual is born, it does not leave offspring, and as a result the mutation is eliminated. In this way, higher organisms would not be able to get rid of all the mutations that occur massively in mitochondrial generations. In the case of mitochondria, selection is carried out according to many hierarchical steps. Let's imagine hierarchical levels of mitochondrial transmission to offspring: a mutation appears in mitochondria, and there are many mitochondria in the cell, and not all of them necessarily carry mutations; a female has many oocytes, and not every one of them has mutant mitochondria; and, finally, there are many females in the population, and not every one of them has oocytes with mutant mitochondria.

At each of these hierarchical levels, normal, viable mitochondria can be selected. James Bruce Stewart and colleagues from the Faculty of Laboratory Medicine at the Royal Institute in Stockholm (Sweden) and the Mitochondrial Research Laboratory at Newcastle University (UK) conducted an experiment proving that the selection of normal mitochondria occurs not at the organismic, but at the cellular or subcellular level.

The experimenters worked with mice that carried a mutation in the mitochondrial DNA polymerase gene, the so-called gamma polymerase. Gamma polymerase is responsible for DNA replication in mitochondria, and if this protein is flawed, then copying DNA in mitochondrial genes will result in a lot of errors. As a result, the function of mitochondria — cellular respiration — will be performed inefficiently. The gamma polymerase gene is located not in the mitochondrial genome, but in the central (nuclear) one.

During the experiment, a line of mice homozygous for a mutation in this gene was bred. These mice had signs of mitochondrial diseases: they used to age. Homozygous females were crossed with normal males and received offspring heterozygous for mutation of the gamma polymerase gene (one copy of the gene is mutant, the other is normal). Mitochondria in these mice contained many mutations inherited from the mother (recall that mitochondria are inherited exclusively through the female line).

By crossing heterozygotes with each other, the researchers obtained a second generation with a classic 1:2:1 cleavage by mutation of gamma polymerase (25% of mice with two normal copies of the gene, 50% of heterozygotes and 25% of mice with two mutant copies of the gene). From this generation, the experimenters selected females that did not carry the mutant gamma polymerase gene, but inherited mitochondrial DNA with harmful mutations from the mutant grandmother.

These females were then crossed with normal males: they received the next generation, then another and another, and so they received 6 consecutive generations. All these mice carried a normal nuclear gene, but inherited damaged mitochondrial DNA from their mother. mtDNA was sequenced in each generation and the number of nucleotide substitutions was calculated. It was important for scientists to assess the rate at which the number of mutant mitochondrial genes decreases in a number of generations. To do this, we used a standard indicator of the ratio of significant and insignificant nucleotide substitutions and found out how much this ratio differs from the random one. (Here I will clarify that those nucleotide substitutions that lead to the replacement of an amino acid in the encoded protein are considered significant. By the ratio of significant and insignificant substitutions, one can judge the effectiveness of the cleansing selection, which should reject significant substitutions and ignore insignificant ones.)

It turned out that by the sixth generation of mice, there were almost no mutant mitochondria with significant replacements. In other words, the selection of mitochondria for compliance with a high energy standard occurs very quickly. And it is conducted, judging by the rate of disappearance of harmful mutations, not on the basis of the fitness of the whole organism, but at lower levels of organization — at the subcellular level or at the level of oocytes. That is, the body somehow copes very quickly with errors in the reproduction of mitochondria, eventually giving out generations of mitochondria freed from mutations.

The paper does not show the mechanism of this purification, but the phenomenon of hierarchical selection is clearly demonstrated. This phenomenon is important from both theoretical and practical positions. The development of a hierarchical selection model is important for understanding the evolution of symbiotic organisms, and in the world, as it is now becoming clear, there are very few organisms that do not have symbionts. Meanwhile, classical selection models exploit the fitness characteristics of the whole organism, that is, they take into account only one hierarchical level.

From a practical point of view, understanding how to get rid of mutant mitochondrial genes should help in finding ways to treat mitochondrial diseases. In humans, as well as in mice, about 58% of mitochondrial diseases are caused by mutations in mitochondrial genes encoding transport RNAs (tRNAs). At the same time, in order for the disease to manifest, the level of mutant mitochondria must become quite high. The experiment showed that the mechanism of purifying selection seems to work differently for protein and tRNA genes. It should be emphasized that in the experiment, the level of mutations in the tRNA genes remained high, that is, fast and effective purification selection worked only for genes encoding proteins. What is the difference here? Why does selection stop working when it comes to tRNA?

I would like to note that extremely interesting projects are being conducted on this topic in Moscow, at the Institute of Information Transmission Problems of the Russian Academy of Sciences and Moscow State University. In particular, Moscow biologists led by M. S. Gelfand managed to compare the effectiveness of getting rid of mutations in mitochondria and proteobacteria, whose ancestors once became symbionts-mitochondria. They came to the unexpected conclusion that in mitochondria, despite the complete absence of recombination and the relatively low number of "populations", the efficiency of mutation elimination is extremely high, much higher than in free-living analogues or in obligate parasitic proteobacteria. Apparently, it's about some interactions within the eukaryotic cell. In general, researchers still have a lot of work and new discoveries to make.

Sources:

1) James Bruce Stewart, Christoph Freyer, Joanna L. Elson, Anna Wredenberg, Zekiye Cansu, Aleksandra Trifunovic, Nils-Göran Larsson. Strong Purifying Selection in Transmission of Mammalian Mitochondrial DNA // PLoS Biology 6(1): e10 doi:10.1371/journal.pbio.0060010.

2) David M. Rand. Mitigating Mutational Meltdown in Mammalian Mitochondria // PLoS Biology 6(2): e35 doi:10.1371/journal.pbio.0060035.

Portal "Eternal youth" www.vechnayamolodost.ru
27.02.2008