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Mitophagy in Huntington's Disease
Šonský, Ivan ; Hansíková, Hana (advisor) ; Macůrková, Marie (referee)
Mitochondrial dysfunctions contribute to the progression of many neurodegenerative diseases, including Huntington's disease (HD). In HD, mutation in the huntingtin gene (HTT) results in the expansion of CAG repeats, causing the growth of the polyglutamine tract. This growth is responsible for the gain of toxicity function of the protein. The turnover of dysfunctional and damaged mitochondria is mediated via mitophagy - a selective form of autophagy. Additionally, mitophagy impairments have recently been described to play a key role not only in neurodegenerative diseases. The protrusion of mitophagy results in the clustering of defective mitochondria, organelles which are responsible for fulfilling the energetic demands of neural cells. The most distinctive impact of the impairment is on the striatal medium spiny neurons and results in the development of motor and cognitive dysfunctions. This thesis describes how HD affects mitophagy and reveals the biggest obstacle of mitophagy - disruption of mitochondria targeting into emerging autophagosomes caused by the abnormal interaction of mHTT and p62. Induction of mitophagy at this stage could be crucial for the future therapeutic research of HD. Generally, initiation of mitophagy could become a relevant therapeutic target for many other...
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The role of autophagy in neurodegenerative processes
Marková, Veronika ; Novotný, Jiří (advisor) ; Čermák, Vladimír (referee)
The characteristics of many neurodegenerative diseases including Alzheimer's, Parkinson's and Huntington's disease is the accumulation of proteins and damaged organelles in the cytoplasm. Unfortunately, they are not sufficiently eliminated by autophagy. The basal autophagy, that maintains the cellular homeostasis, is disturbed in neurodegeneration. The process of autophagy becomes saturated and unable to remove all the toxic substances. Therefore, other degradation mechanisms are activated, aiming to restore the homeostasis. However, the neuronal cells are damaged under certain conditions leading to their death. The reduction in the number of neurons in specific brain areas may cause severe ataxias and dementias. Better understanding of autophagocytosis and its role v pathogenesis of neurodegenerative diseases may contribute to more effective treatment of these serious diseases in the future. Key words: autophagy, neurodegeneration, Alzheimer's disease, Parkinson's disease, Huntington's disease
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Role of autophagy in yeast cell adaptation
Brádlerová, Michaela ; Kuthan, Martin (advisor) ; Zikánová, Blanka (referee)
Autophagy is an evolutionarily conserved degradative pathway. Autophagy occurs constitutively at a basal level and it is involved in the recycling and turnover of damaged or superfluous organelles and proteins. It has a critical role in cellular homeostasis. Autophagy can be induced in response to starvation or other types of stress. Induction of autophagy during these conditions has a major role in protection and adaptation of the cell. Autophagy needs to be properly regulated. A wide range of diseases is associated with dysregulation of autophagy. Better understanding of autophagy mechanisms can help to develop strategies designed to modulate autophagic responses occuring in a number of diseases. This work is focused on current knowledge of main types of autophagy and how autophagy helps yeast cells to adapt. Key words: autophagy, yeast, degradative pathway, adaptation, TORC1
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Study of selected plant exocyst subunits and its interactors in autophagy pathway.
Rácová, Denisa ; Žárský, Viktor (advisor) ; Wilhelmová, Naďa (referee)
Exocyst is a binding protein complex, which is evolutionary conserved in yeast, animal and in plant cells. It has crucial role in regulation of cell morfogenesis and cell polarity. The function of the exocyst complex is binding of secretoric vesicle to the proper side on plasma membrane in penultimate step of exocytosis. This process is essecial for function and survival of cell. Another process crucial for the cell is autophagy. In plants autophagy plays important role in the responses to nutrient starvation, senescence, abiotic and biotic stress. RabG3b are small GTPases, which have positive role in autophagy. In this work I described the interaction between RabG3b and some of subunits of exocyst complex: Exo70B1, Exo70B2 and Exo84b. I also studied changes in morfogenesis of tonoplast by induction and inhibition of authophagy and induction of anthocyans synthesis in Arabidopsis thaliana.
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The role of protein kinase C and its targets in cardioprotection
Holzerová, Kristýna ; Hlaváčková, Markéta (advisor) ; Alán, Lukáš (referee) ; Vízek, Martin (referee)
The mortality of cardiovascular diseases remains high and it likely tends to increase in the future. Although many ways how to increase the resistance against myocardial ischemia- reperfusion damage have been described, few of them were transferred into clinical practice. Cardioprotective effect of chronic hypoxia has been described during 60s of the last century. Its detailed mechanism has not been elucidated, but a number of components has been identified. One of these components presents protein kinase C (PKC). The role of PKC was described in detail in the mechanism of ischemic preconditioning, but its involvement in the mechanism of cardioprotection induced by chronic hypoxia remains unclear. One reason is the amount of PKC isoforms, which have often contradictory effects, and the diversity of hypoxic models used. The most frequently mentioned isoforms in connection with cardioprotection are PKCδ and PKCε. The aim of my thesis was to analyze changes in these PKC isoforms at two different cardioprotective models of hypoxia - intermittent hypobaric (IHH) and continuous normobaric hypoxia (CNH). We also examined the target proteins of PKCδ and PKCε after the adaptation to IHH, which could be involved in the mechanism of cardioprotection. These included proteins associated with apoptosis and...
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The role of autophagy and selected beta-tubulin isotypes in taxane resistance in breast cancer cells
Kábelová, Adéla ; Jelínek, Michael (advisor) ; Truksa, Jaroslav (referee)
Drug resistance in cancer cells is a frequent cause of breast cancer therapy failure. The aim of this thesis was to elucidate mechanisms of resistance to taxanes, that are used in therapy of various types of cancer, including breast cancer. We particularly assessed the role of autophagy and changes in βII- and βIII isotype gene expression in development of taxane resistance. As model of breast cancer we used human sensitive cell lines SK-BR-3, MCF-7 a T47-D and resistant sublines SK-BR-3-PAC/REZ a MCF-7- PAC/REZ which grow in paclitaxel concentration lethal for sensitive sublines. In cell lines SK-BR-3 and MCF-7, taxane application decreased the level of autophagy, however in cell line T47-D led to its activation. We detected no difference between basal levels of autophagy in sensitive subline SK-BR-3 compared to resistant subline SK-BR-3-PAC/REZ, but we observed increased basal level of autophagy in sensitive subline MCF-7 compared to the resistant subline. Increase or decrease level of autophagy did not affect taxane resistance, except activation of autophagy in resistant subline SK-BR-3-PAC/REZ, that further increased the resistance to paclitaxel. Taxane application in cell line T47-D increased the levels of βII- and βIII-tubuline expression, however we did not find any similar effect in other tested...
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Vazba paralogů EXO70 na ATG8 a funkční rozdělení rodiny EXO70 dle účasti v autofagii (Arabidopsis thaliana).
Semerádová, Hana ; Kulich, Ivan (advisor) ; Motyka, Václav (referee)
The exocyst, an octameric protein complex conserved among all eukaryotes, mediates tethering of the vesicle prior to its fusion with the target membrane. Apart from the function of exocyst in exocytosis, new studies from both mammalian and plant fields report its involvement in the cellular self-eating process called autophagy. In land plants the number of paralogs of some exocyst subunits is extraordinarily large. There are 23 paralogs of Exo70 subunit in Arabidopsis thaliana. It is supposed that these paralogs have acquired functional specialization during the evolution - including involvement in autophagy. Using yeast two- hybrid assay it is shown here that Exo70B1 and Exo70B2, but not other Arabidopsis Exo70 paralogs interact with Atg8, an autophagosomal marker. The proximity of these two paralogs and Atg8 in vivo was confirmed by independent Förster resonance energy transfer (FRET) method. Interestingly, interaction of Atg8f with Exo70B2 paralog appears to be stronger than with Exo70B1. Exo70B1-mRUBY expressed under the natural promoter shows punctate membrane structures that are mostly static. That changes after the tunicamycin treatment - movement of some of these dots was induced. Homology modeling of Exo70B1 and Exo70B2 proteins tertiary structure in combination with bioinformatic prediction based...
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Toll like receptors and myeloid cells in development and disease
Balounová, Jana ; Filipp, Dominik (advisor) ; Špíšek, Radek (referee) ; Vannucci, Luca Ernesto (referee)
Toll like receptors (TLRs) are germline-encoded pattern recognition receptors (PRRs) that play a central role in host cell recognition and responses to pathogens. Primarily they are responsible for induction and regulation of the innate and adaptive immune responses whereby the effector function is executed chiefly by differentiated myeloid cells. Somewhat unexpectedly, TLRs have been also shown to be involved in direct pathogen sensing by bone marrow-derived hematopoietic stem cells (HSCs) and hematopoietic progenitors when, under inflammatory conditions, the rapid generation of innate immune effector cells that effectively combat the infection is of utmost priority. While it has been recognized that the release of inflammatory cytokines from inflamed tissues along with the changes in proportions of differentiating cells in the bone marrow (BM) as well as the BM niche can nudge the differentiation of adult BM-derived cells towards myeloid cells and granulocytes, a direct role of TLRs expressed by HSCs in this process has been demonstrated only recently. However, whether a similar mechanism operates also during embryonic hematopoiesis is unknown. Here we show that TLRs and their adaptor proteins are functionally expressed during early stages of embryogenesis by short-lived maternally-transferred...
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Autophagy and other processes in colonies of natural yeast strains
Novosadová, Zuzana ; Palková, Zdena (advisor) ; Groušl, Tomáš (referee)
Abstract The yeast Saccharomyces cerevisiae on solid media forms multicellular colonies. Cells within colonies undergo differentiation and metabolic diversification, including formation of two layers of cells called Upper and Lower cells. The metabolic activity of U and L cells is different. For instance a higher level of autophagy was observed in U cells. This thesis includes a literature review of molecular mechanisms of autophagy in yeasts. Yeast colonies, under starvation conditions, produce volatile ammonia signal. This signal allows them communicate over a distance. Studies, revealing cell differentiation within colonies and ammonia signalling among colonies, were performed in colonies of laboratory strains. Strains isolated from nature, so-called wild strains, form distinct structured colonies, termed fluffy. Yeast within fluffy colonies also form different cell types. However the situation seems to be more complex that within smooth colonies of laboratory strains. Strains were constructed during this study, which express marker proteins Icl2p, Pox1p, Mae1p, Pma1p, Pma2p, Ino1p, Met17p and Atg8p fused with fluorescent labels in order to study cell differentiation and other processes within fluffy colonies. Furthermore, a new system for ...
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Intracellular life of pathogenic bacterium Francisella tularensis in the host.
Rädisch, Robert ; Konopásek, Ivo (advisor) ; Vopálenská, Irena (referee)
Francisella tularensis is a facultative intracellular pathogenic bacterium, which causes disease named tularemia. For the entrance to the host cells Francisella uses host's cell mechanisms by which it is incorporated into cell phagosome. Subsequently, it escapes from phagosome to cytosole where bacterial growth takes place. Some of bacteria are cleared from cytosol by autophagy, from another ones dsDNA is released. This DNA is recognized by cytosolic receptors, which form inflammasome complex. Inflammasome sets off pathway leading to the death of infected cell. Since the penetration to the cell Francisella modulates cell signallization in its own benefit to ensure enough time and nutrients for its growth. Francisella do not act only in the infected cells, where it reduces recognition of itself and clearance from cytosol, but it also induces secretion of factors, which moderate activation of adaptive immunity of the host. Key words: Francisella, tularemia, fagosome, inflammasome, autophagy, adaptive immunity
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