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Molecular composition and ultrastructure of holokinetic chromosomes
Šejgunovová, Nikola ; Král, Jiří (advisor) ; Dalíková, Martina (referee)
Holokinetic chromosomes are a specific type of chromosomes which differentiate from standard (monocentric) chromosomes especially by a diffuse form of domain which binds microtubules (holocentromere). It is related to changes on an ultrastructural and molecular level. These changes are shown in modifications in mitotic and meiotic division and in evolution of karyotypes. Holokinetic chromosomes don't have a primary constriction with a localized centromere and therefore neither an inner centromere domain which would connect sister chromatids. Kinetochore structure of holokinetic chromosomes seems to be simpler than kinetochore structure of monocentric chromosomes. Kinetochore covers most of the surface of mitotic chromosomes. There have been described several variants of meiosis of holokinetic chromosomes which differentiate by position of kinetochore on chromosomes. On a molecular level holokinetic chromosomes differentiate from monocentric chromosomes by a distribution of proteins of a centromere-kinetochore complex, which cover most of the surfaces of mitotic and meiotic chromosomes. This applies, for example, to centromeric histone H3 (CENH3), whose amount and distribution changes during interphase and nuclear division, which is unique in comparison to monocentric chromosomes. The distribution...
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Analysis of dosage effect of speciation gene Prdm9 on fertility of mouse hybrids
Flachs, Petr ; Trachtulec, Zdeněk (advisor) ; Stopka, Pavel (referee) ; Král, Jiří (referee)
(eng) The phenomenon of hybrid sterility represents one of the evolutionary mechanisms that enables speciation. Only a few speciation genes have been uncovered. The only one found in mammals is Prdm9 (PR-domain 9). Data in the literature on the involvement of Prdm9 in decreased fertility of various semifertile hybrid males of house mouse subspecies were scarce before the results of this thesis were completed, despite that such males are much more frequent in nature than the fully sterile ones. Utilizing a panel of genetic tools and a battery of phenotyping tests, this thesis shows a central role of Prdm9 in fecundity of hybrids, including many fertility disorders and age dependency. Both increasing and reducing the Prdm9 gene dosage significantly elevated fertility parameters. Surprisingly, even the allele that in one copy causes full hybrid sterility increased F1 hybrid fertility when present in multiple copies. The PRDM9 protein also plays a role in identifying the sites of meiotic recombination. This study also points out the principles of allelic competition in determination of the sites of preferred recombination (hotspots), which suggests a possible link between both previously described Prdm9 roles. This thesis summarizes a set of three logically interconnected publications with the ambition...
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Genetic interactions of the Prdm9 gene
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
The Prdm9 gene (PR domain containing 9, Meisetz, Hybrid sterility 1) encodes enzyme that trimethylates histone 3 on lysines 4 and 36. These methylation marks determine the positions of DNA double-strand breaks that are repaired by meiotic homologous recombination. In this study, we assayed genetic interactions of Prdm9 with two genes important for spermatogenesis - Mili (Piwil2) involved in piRNA biogenesis and Mybl1 encoding transcription factor that regulates many genes important for prophase I, including piRNA precursors. We crossed laboratory mice carrying mutation in Prdm9 with heterozygotes for mutation in Mybl1 or Mili, and created compound heterozygotes and, in case of Mybl1, also double homozygotes. We assessed body weight and male fertility parameters (weight of testes, sperm count, malformed sperm, percentage of tubules containing spermatocytes and of abnormal nuclei of pachytene spermatocytes) of these mice and compared them to controls. We also investigated the effect of Mybl1 and Mili mutations on fecundity of F1 intersubspecific hybrids. Our results revealed possible interactions of Prdm9 and Mybl1 in the laboratory mouse. Decreased gene dosage of Mybl1 reduced fertility of intersubspecific F1 hybrids. Interaction between Prdm9 and Mili in both laboratory mouse and F1 hybrids remain...
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Polyploidy with specific view to paleopolyploidy and its detection
Kotz, Matěj ; Král, Jiří (advisor) ; Rothová, Olga (referee)
Polyploidy is a significant phenomenon in the evolution of eucaryotic organisms. It has been a subject of interest of biologists for decades and it has been studied particularly largely in plants, where it most often occurs. This thesis handles occurrence of poylploidy in organims and a special emphasis is laid on old polyploidization events occurrence of polyploidy in animal taxa. Furthermore, the types of polyploidy and obstacles are discussed here, that a polyploid organism has to tackle to stabilize its genome. That also includes the need to ensure the correct chromosome segregation in a meiotic division. A special case of polyploidy is paleopolyploidy (sometimes refered to as "ancient polyploidy" in English), which is an evolutionary old polyploidization event, that is followed by the process of diploidization. This process is characterized besides others by a large number of structural changes in chromosomes and a loss of some DNA sequences and a gradual transition of the polyploid genome to the cytologically diploid state. Because of these changes, it is difficult to detect a paleopolyploidy and the main part of this thesis is dedicated to this problem, in which the main approaches that might lead to its detection are discussed. Key words: polyploidy, animal, plant, meiosis, paleopolyploidy,...
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The role of H3K36 methylation in DNA repair of germinal and somatic cells
Křivánková, Klára ; Mihola, Ondřej (advisor) ; Schierová, Michaela (referee)
Histone modifications affect many cellular processes including DNA damage repair. This thesis focusses on the methylation of lysine 36 of histone 3 (H3K36). The role of this modification in the localization of DNA double-stranded breaks in germinal cells is described in the first part of this thesis. Double-stranded breaks initiate meiotic recombination, which is essential for successful meiosis. This thesis also describes three histone methyltransferases. The first is PRDM9, an enzyme expressed only in oocytes and spermatocytes during meiotic prophase and responsible for the localization of recombination hotspots in most mammals. The second part of this thesis deals with the role of H3K36 methylation in DNA damage repair in somatic cells using homologous recombination (HR) and nonhomologous DNA ending joining (NHEJ). The proteins SETD2 and SETMAR are described in the second part. SETD2 trimethylates H3K36, and H3K36me3 is recognized by the LEDGF protein. Through LEDGF, other components necessary for HR are recruited to DNA. SETMAR dimethylates H3K36 and together with this histone modification promote DNA break repair with NHEJ. The research of H3K36 methylation is important for a better understanding of each DNA repair mechanisms. The correct repair of DNA breaks is necessary for maintaining...
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Molecular evolution of meiosis in diploids and tetraploids of Arabidopsis arenosa
Holcová, Magdalena ; Schmickl, Roswitha Elisabeth (advisor) ; Mozgová, Iva (referee)
Meiosis is functionally conserved across eukaryotes, thus not expected to vary considerably among different species, and even less so among lineages within a species. However, recent studies showed that this is not necessarily the case in Arabidopsis arenosa. Genome scanning identified an excess differentiation in meiosis genes between A. arenosa diploids and tetraploids, interpreted as meiosis adaptation to the whole genome duplication in tetraploids and differentiation was also found between two diploid lineages. Thus, I present a population-based analysis of positive selection acting on meiosis proteins across multiple lineages of A. arenosa. I showed that meiosis proteins were under positive selection in all diploid lineages, mainly in the Pannonian and South-eastern Carpathian lineage. The evidence for positive selection in diploid lineages suggested differential pathways of meiosis adaptations in the species, probably reflecting the necessity to adapt to local environments, among all to temperature. The highest enrichment of amino acid substitutions (AASs) under positive selection was identified in tetraploids, in consistence with previous genome-scan results. As several interacting meiosis proteins were under positive selection in the same A. arenosa lineage, I hypothesize that the close...
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The role of cohesin genes in the meiosis of male house mouse
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
Cohesin genes play an important role in cell division. They ensure proper chromosome segregation during mitosis and meiosis. This study is focused on the role of cohesin genes during meiosis in male house mouse (Mus musculus). At first, this study introduces key processes of mammalian meiosis. Next, the structure of cohesin complex is described; it consists of a heterodimer SMC proteins - SMC3 and SMC1α or SMC1β, which are enclosed to the ring by cleavable subunit RAD21, RAD21L or REC8. Fourth subunit - a STAG protein (STAG1, STAG2 or STAG3) associates with the cleavable subunit. Meiotic function of specific cohesin proteins (SMC1β, RAD21L, REC8 and STAG3) as deduced from the phenotypes of the deficiencies of their genes in male mouse is depicted. All these four genes are necessary for many processes during meiosis, - e.q. sister chromatid cohesion maintenance, synapsis and recombination. STAG3, SMC1β, and REC8 are necessary for centromeric cohesion. STAG3 and RAD21L are important for the assembly of the remaining cohesin subunits. The most important phenotype of deficiency of all four genes is the complete meiotic arrest in male prophase I. Therefore, cohesin research is important for the investigation of the causes of sterility in mammals. key words: cohesin, meiosis, spermatogenesis, mouse,...
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Maping of genes modifying the subspecies-specific roles of the meiotic gene Prdm9
Škaloudová, Eliška ; Trachtulec, Zdeněk (advisor) ; Schierová, Michaela (referee)
The PRDM9 (PR domain containing 9) protein is an epigenetic factor that trimethylates lysine 4 of histone H3 and thereby determines the future meiotic double-strand breaks - sites important for proper segregation of homologous chromosomes. Males of the Mus musculus domesticus (Mmd) origin with homozygous deletion in Prdm9 (Prdm9-/- ) are sterile with a complete arrest in meiotic prophase I, in contrast to the same mutant males of the M. m. musculus (Mmm) subspecies. The aim of this diploma thesis was to identify the genomic loci responsible for the phenotypic difference of these Prdm9-/- males. The major research tool was a population of 182 Mmm x Mmd Prdm9-/- males. The mapping method of quantitative trait loci (QTLs) was based on relating the genotypes of single-nucleotide and microsatellite polymorphisms to the observed phenotypes. At least two QTLs on Chr X were identified. The Mmm alleles of these QTLs reduced fertility of Prdm9-/- males. Both QTLs were confirmed and narrowed down using two types of subconsomic strains. It was not possible to confirm other QTLs, particularly on autosomes. This QTL mapping is the first step towards the identification of genes that modify the resulting phenotype of Prdm9-/- animals. This identification should help designing studies of human infertility that...
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Genes of early meiotic prophase I of spermatogenesis in house mouse
Škaloudová, Eliška ; Trachtulec, Zdeněk (advisor) ; Forman, Martin (referee)
Meiosis is an essential cellular process that is necessary for gamete formation in all sexually reproducing organisms. This work is focused on the description of the genes of early stages of meiotic division in males of a mammalian model, the house mouse. The first part summarizes meiosis focusing on prophase I, which is longer than prophase II. Prophase I is divided into five stages, namely leptotene, zygotene, pachytene, diplotene, and diakinesis. Mouse spermatogenesis and its differences from oogenesis are also briefly described. The second part provides a list of genes encoding proteins required for initiation of meiotic division, pairing and synapses of chromosomes, and initiation of the catalysis of double-strand breaks. Double-strand breaks are repaired by homologous recombination, which may result in so-called crossing-over, the major source of genetic variability. The work deals with the early stage of homologous recombination and components required for this process. Localization of meiotic double-strand breaks in the genome is not random and is under the control of the Prdm9 gene, which seems to take multiple roles, such as the formation of new subspecies of the house mouse. Knowledge of the genes controlling the early stages of meiotic division is a prerequisite to understanding some of...
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Evolution of selected karyotype characters in tetrapulmonate arachnids
Jílková, Klára ; Král, Jiří (advisor) ; Marec, František (referee)
The class Arachnida is not thoroughly explored from the cytogenetic point of view. Previous studies suggest a high diversity of karyotypes and sex determination in arachnids. This study deals with the evolution of sex chomosomes, nucleolar organizer regions (NOR), and telomeric repeats in the tetrapulmonate clade of arachnids, particularly in groups of ancient origin. Sex chromosomes were detected in two orders. Detection of NORs in a large set of species supports the hypothesis that the ancestral karyotype of arachnids contained NOR on one pair of autosomes only. The number of NORs has increased during the evolution of some groups of Pedipalpi. The NORs are located in terminal or subterminal chromosomal regions in most tetrapulmonates. The occurrence of the "insect" telomeric motif was confirmed in majority of tetrapulmonates. Interstitital telomeric repeats were not detected with the exception of one species. Keywords: arachnids, meiosis, sex chromosomes, telomeres, nucleolar organizer, heterochromatin
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