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Bioactive molecules involved in blood processing by haematophagous monogeneans of the family Diplozoidae
Jedličková, Lucie
Monogeneans from the family Diplozoidae (subclass Heteronchoinea) are bloodfeeding ectoparasites inhabiting gills of common carp. Digestion of blood in diplozoids is an intracellular process taking place in gut cells within lysosomal cycle in the presence of parasite's peptidases. However, information about the blood digestion comes only from ultrastructural and histochemical analyses. Therefore, I have focused in this work on biochemical and molecular characteristics of bioactive molecules which may participate in blood processing by E. nipponicum adults, especially cysteine peptidases of cathepsin L- and B- types, aspartic peptidases of cathepsin D-type, and Kunitz-type inhibitors of serine peptidases. In homogenates and excretory/secretory (E/S) products of E. nipponicum adults, an activity of cysteine peptidases of cathepsins L-type dominated, followed by an activity of cathepsin D-like aspartic peptidases and a minor cathepsin B-like activity. Inhibitors of the abovementioned peptidase types completely blocked hemoglobinolytic activity in the samples. In the transcriptome of E. nipponicum adults, ten cathepsin L-coding transcripts were found and only one cathepsin B-coding transcript. Primary structures of the encoded enzymes were bioinformatically and phylogenetically compared. Two abundant...
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Evolution of euglenid plastid proteome
Novák Vanclová, Anna ; Hampl, Vladimír (advisor) ; Gruber, Ansgar (referee) ; Maier, Uwe (referee)
Endosymbiotic gain and transfer of plastids is a widespread evolutionary phenomenon and a major driving force of eukaryotic evolution. The integration of a new organelle is accompanied by changes in its structure, gene content, molecular mechanisms for biogenesis and transport, and re-wiring of the host and organelle metabolic pathways. To understand the course and underlying mechanisms of plastid evolution, it is important to study these processes in variety of secondary algae and notice their differences and similarities. Euglenophytes gained their plastids from green eukaryotic algae after a long history of heterotrophic lifestyle. In my thesis, I participated in analyses of newly generated sequence datasets: transcriptomes of Euglena gracilis and Euglena longa and mass spectrometry-determined proteome of E. gracilis plastid with especial regard to the potential novelties associated with plastid gain and incorporation. In the resulting publications we particularly focus on plastid protein import machinery and targeting signals and report extremely reduced TIC and completely absent TOC in euglenophyte plastid. Using the proteomic dataset, we predict potential novel plastid protein translocases recruited from ER/Golgi and re-analyze plastid signal domains, characterizing previously overlooked...
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Bioactive molecules involved in blood processing by haematophagous monogeneans of the family Diplozoidae
Jedličková, Lucie ; Mikeš, Libor (advisor) ; Horn, Martin (referee) ; Sojka, Daniel (referee)
Monogeneans from the family Diplozoidae (subclass Heteronchoinea) are bloodfeeding ectoparasites inhabiting gills of common carp. Digestion of blood in diplozoids is an intracellular process taking place in gut cells within lysosomal cycle in the presence of parasite's peptidases. However, information about the blood digestion comes only from ultrastructural and histochemical analyses. Therefore, I have focused in this work on biochemical and molecular characteristics of bioactive molecules which may participate in blood processing by E. nipponicum adults, especially cysteine peptidases of cathepsin L- and B- types, aspartic peptidases of cathepsin D-type, and Kunitz-type inhibitors of serine peptidases. In homogenates and excretory/secretory (E/S) products of E. nipponicum adults, an activity of cysteine peptidases of cathepsins L-type dominated, followed by an activity of cathepsin D-like aspartic peptidases and a minor cathepsin B-like activity. Inhibitors of the abovementioned peptidase types completely blocked hemoglobinolytic activity in the samples. In the transcriptome of E. nipponicum adults, ten cathepsin L-coding transcripts were found and only one cathepsin B-coding transcript. Primary structures of the encoded enzymes were bioinformatically and phylogenetically compared. Two abundant...
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Variability of the domestic chicken breeds in selected immunological traits of hen and egg
Bílková, Barbora ; Vinkler, Michal (advisor) ; Plachý, Jiří (referee) ; Trefil, Pavel (referee)
The avian immune system is a complex system of defence mechanisms that protect bird hosts against threats from ubiquitous pathogens. According to the co-evolutionary models, variability in immune traits of hosts is the key component providing ability to adapt and enhance their defence mechanisms in presence of constant selective pathogen pressure. Domestic chicken (Gallus gallus f. domestica) is used as a model organism in avian biology and also is one of the most important food-producing animals, not only for their meat but also for the egg production. Unfortunately, in research usually only inbred chicken lines are used and modern poultry husbandry is tight with unilateral breeding towards highly productive breeds. Those approaches decrease intra-population polymorphism in chickens. However, especially in case of farm animals, searching and extending the pool of immune variability and enhancing pathogen resistance is crucial for sustaining healthy and biologically secure populations and their products. Morphologically highly distinct traditional chicken breeds, which have evolved for hundreds years under different selective pressures, may represent this desirable immunological variability. In my thesis I described variability in chosen immunological traits, haematological parameters and proteomic...
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Identification of key regulators of gene expression in mammalian oocyte and embryo
Jansová, Denisa ; Šušor, Andrej (advisor) ; Gahurová, Lenka (referee) ; Lin, Chih-Jen Lance (referee)
Mammalian oocyte is a highly differentiated cell which gives rise to an embryo after fertilization. Importantly, fully-grown oocytes become transcriptionally inactive at the end of the growth phase. During following stages of development, i. e. meiotic maturation of the oocyte and early embryonic development, only transcripts previously synthesized and stored are used. The tight correlation between mRNA distribution and subsequent protein localization and function provides a mechanism of spatial and temporal regulation of gene expression used by various cell types. However, not much is known about mRNA localization and translation in the mammalian oocyte and early embryo. The aim of my thesis was to determine the localization of transcripts and components of translational machinery in the mammalian oocyte and embryo and to uncover the mechanisms of spatiotemporal regulation of translation as a prerequisite for correct oocyte and embryo development. We have shown that nuclei of both mouse and human oocytes contain RNA molecules and RNA binding proteins. Following the nuclear envelope breakdown (NEBD), translational hot-spots occur in the area surrounding the nuclear region. We suppose that mRNAs previously retained in the nucleus are released to the cytoplasm during NEBD and their subsequent...
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Plant sugar metabolism and arbuscular mycorrhizal symbiosis
Konečný, Jan ; Jansa, Jan (advisor) ; Gryndler, Milan (referee)
The study of arbuscular mycorrhizal symbiosis (AMS) - the mutualist relationship between the most of land plants and evolutionary old fungal group Glomeromycota - is becoming a prestigious topic. The prevalence of and extent of physiological action of AMS on plants is very interesting for the plant biology itself, but its importance grows, notably in time of global climate change, frequent soil degradation and ascending exhaustion of mineral fertilizer reserves. Although the flows in AMS of some minerals, like of phosphorus was enlightened, carbon exchange between the symbionts is still poorly understood. In this experimental work, I utilized the boom of molecular and bioinformatic methods in the quest for completely unexplained carbon flows. The organisms used include barrel medic (Medicago truncatula), the model legume for symbiotic relationships, biotic, and abiotic stresses; Rhizophagus irregularis, the widely used fungus for such experimental studies of AMS; and Sinorhizobium meliloti, the nodulating nitrogen-fixing bacterium compatible with the barrel medic. Two variants - mycorrhizal (M+) and non-mycorrhizal (NM) plants were subjected to several levels of analysis. I have checked the variants, did the measurements of phosphorus and nitrogen contents, as well as I probed the plants with...
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Mitogenomic phylogeography and adaptive evolution of the bank vole Clethrionomys glareolus
Filipi, Karolína ; Kotlík, Petr (advisor) ; Munclinger, Pavel (referee)
This thesis is a part of the project aimed at sequencing the genome and transcriptome of the bank vole (Clethrionomys glareolus). The role of natural selection in the evolution of mitochondrial DNA (mtDNA) has been subject to much discussion; while some studies did not provide evidence that selection affected the phylogeography of the studied species, other considered adaptive evolution important. The bank vole is the key model we use to study the adaptation to climate change. As with other species, the phylogeography of the bank vole has been based on the variation of a small part of mtDNA. The goal of the thesis was to sequence the entire mitochondrial genome for representatives of all main mtDNA lineages of the bank vole using the Sanger and Illumina technologies, and to assess the role of selection and adaptation in the evolution and phylogeography of this species. The adaptive evolution in mtDNA probably was not the main driving force during the postlacial colonization of Europe. However, signatures of adaptive evolution have been found - an amino acid change with possible functional consequences in one gene and an excess of radical changes in physical- chemical properties of amino acids in populations at the latitudinal (northern and southern) extremes of the bank vole distribution. Key...
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Search for the remnant of plastid in the cell of Rhabdomonas sp.
Soukal, Petr ; Hampl, Vladimír (advisor) ; Oborník, Miroslav (referee)
Phylum Euglenozoa contsists of four groups - Euglenoidea, Kinetoplastea, Symbiontida and Diplonemea. Phototrophic euglenids, which constitute a clade, possess green plastid acquired via secondary endosymbiosis from green alga related to recent genus Pyramimonas. According to generally accepted plastid-late hypothesis, the endo- symbiosis took place after split between phototrophic euglenids and all other euglenozo- ans. Endosymbiotic event is always associated with gene transfer from endosymbiont to nuclei of host. Even if the endosymbiont is completely lost we should be able to observe enrichment of the host genome with the genes derived from endosymbiont. Some recent phylogenetic analyses uncovered genes related to green algae in trypanosomas (Kineto- plastea: Trypanosomatida). Based on this observation, authors postulated a hypothesis that the plastid was present already in common ancestor of kinetoplastids and euglenids and was lost in kinetoplastids and some euglenids including osmotrophic Rhabdomonas costata. During analysis of transcriptome of R. costata we found 63 genes, which could originated from green (24 genes) or other (49 genes) algae. In phylogenetic trees only one was robustly related to green and four were robustly related to other algae. Since the number of genes related to...
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High-throughput analysis of uridine insertion and deletion RNA editing in \kur{Perkinsela}
DAVID, Vojtěch
This thesis is a follow-up of my Bachelor thesis about the mitochondrial genome of kinetoplastid protist Perkinsela sp. This work introduces a novel approach in high-throughput analysis method of uridine insertion and deletion RNA editing, describes its background and proposes its further development. Its effect on the interpretation of U-indel editing, both in Perkinsela and in general, is demonstrated via attached manuscript which also introduces other biologically relevant aspects of Perkinsela mitochondrion.
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Transcriptomic and functional analysis of salivary proteins from the tick \kur{Ixodes ricinus}
CHMELAŘ, Jindřich
This thesis was focused on the identification and characterization of the salivary proteins from Ixodes ricinus, the European vector of Lyme disease and tick-borne encephalitis causative agents. In the first part of this work, the the transcriptomic approach was used in order to identify and describe I. ricinus salivary proteins. The second part is dealing with functional and structural characterization of the salivary protein named IRS-2 (I. ricinus serpin-2).
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