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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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Autophagy as a mechanism of adaptation the yeast
Zieglerová, Leona ; Váchová, Libuše (advisor) ; Zikánová, Blanka (referee)
Autophagy is a degradation pathway, conserved from yeast to mammals. The uniqueness of this pathway lies in its function, it is applied in the cell especially under the adverse conditions. It helps the cell to deliver essential nutrients for life, it removes the damaged or superfluous organelles, protein aggregates and helps with recycling and maintains a constant inner environment. These functions can prolong cell life and the cells survive the adverse conditions. Autophagy may induce the programmed cell death type II. This paper describes the basic of autophagy machinery, regulation and influence of yeast autophagy to adapt to the stressful conditions. Understanding the mechanism and regulation of autophagy in yeast may help with the study of autophagy in mammals. In mammals, this degradation pathway disorders cause many diseases (especially neurodegenerative), autophagy also effects the formation of tumors. Powered by TCPDF (www.tcpdf.org)
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Autophagy in the heart
Šprláková, Katarína ; Hlaváčková, Markéta (advisor) ; Tomšů, Eva (referee)
Currently, it is growing evidence that autophagy is involved in the prevention of various diseases, which of course also includes heart diseases. This thesis is therefore aimed at clarifying the role of autophagy in the heart, especially during ischemia and subsequent reperfusion. Autophagy is a physiological cellular process by which the cell maintains homeostasis by eliminating long-lived proteins and damaged organelles. The role of autophagy during ischemia/reperfusion in the heart is complex. Predominantly it functions as a pro-survival pathway, because it protects the heart from ischemia or hypoxia. However, when triggered over, which happens during reperfusion, it may lead to cell death. In the heart autophagy is activated in response to various stimuli, such as decrease in ATP and subsequent activation of AMPK, protein Bnip3, reactive oxygen and nitrogen species, the opening of mitochondrial permeability transition pore, endoplasmic reticulum stress or unfolded protein response.
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Autophagy in the immune system
Vávra, Dan ; Černý, Jan (advisor) ; Janštová, Vanda (referee)
Autophagy is an essential, homeostatic process - survival mechanism that protects cells by various ways: cells break down their own components to recycle nutrients, remodel and dispose unwanted cytoplasmic constituents. Autophagy is involved in the degradation of long-lived proteins and entire organelles, but paradoxically, considering important prosurvival functions, autophagy may be deleterious. It plays an important role during development, tumor suppression, immunity and is required for the adaptation to environmental stresses such as starvation. Recent studies indicate, that autophagy is a central player in the immunological control of bacterial, parasitic and viral infections. The process of autophagy may degrade intracellulal pathogens. This work describes the mechanism of autophagy and highlights the role of autophagy in innate and adaptive imunity, summarizes some advances in understanding the functions of autophagy and its possible roles in the causation and prevention of human deseases.
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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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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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Conventional and Novel Functions of the Exocyst Complex in Plants
Kulich, Ivan ; Žárský, Viktor (advisor) ; Baluška, František (referee) ; Hašek, Jiří (referee)
Exocyst is an octameric protein complex, conserved across all Eukaryotes. Its role, originally described in yeast, resides in a tethering of the secretory vesicles to the plasma membrane prior to the membrane fusion of the two membranes. Subunits SEC3 and EXO70 are believed to be spatial landmarks for the vesicles delivery. While yeast genome encodes single EXO70, we find dozens of them in land plants (23 in Arabidopsis). This work is focused at a role of the exocyst complex in plant cells. Its first part documents, that exocyst is essential for delivery of the cell wall components, namely pectins, but also for pathogen induced secondary cell wall thickening. Second part reveals an unconventional role of EXO70B1 subunit harboring exocyst subcomplex at an autophagic pathway to the vacuole and raises many questions about plant secretory pathway.
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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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Secretion and autophagy in plant defence against microbes
Dobešová, Karolína ; Žárský, Viktor (advisor) ; Burketová, Lenka (referee)
Plants are sessile organisms and when attacked by microbes, they cannot easily run away. For this reason, they have developed sophisticated defensive mechanisms, that allow them to defend themselves. Since plants, unlike mammals, do not have any special immune cells, their defense takes place in each cell separately. The key moment during a microbial infection is the recognition of the microbe by the plant through its released molecular patterns (mostly proteins) associated with microbes (MAMPs). MAMPs trigger signaling cascades that lead to the secretion of antimicrobial compounds to the site of an attack. The process of autophagy is also important in the defense against microbes, which not only maintains a cellular homeostasis and controls the level of phytohormones and defense proteins in the plant cytoplasm, but also participates in the secretory activity of the cell. Recent analyzes of plant secretome have shown that plants secrete many proteins (including defensive ones) independently of the signal peptide and compartments of a conventional secretion. During exocytosis a vesicle fuses with the cytoplasmic membrane. The octameric protein complex exocyst and SNARE proteins take part in this process. The exocyst complex is highly diversified in plants - especially it's EXO70 subunit, which is...
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Autophagy in plant abiotic stress
Kosťová, Natálie ; Hála, Michal (advisor) ; Martinec, Jan (referee)
Autophagy is a cellular mechanism when cell gets rid of its damaged or unwanted components. These are wrapped into a double membrane and creates so-called autophagosome, which is transported to the vacuole, where its contents are degraded or recycled. Autophagy runs essentially through whole life of the plant, but if it is necessary, for example in response to stressful conditions, it is significantly intensified. It is relatively complexly regulated mechanism, which, especially in plants, still has not been completely described. Autophagy plays an important role under the influence of stress conditions. Especially, the effects of abiotic stress play an important role in plant life. Plants are immobile organisms so they must therefore develop mechanisms to enable them to survive in adverse conditions. In response to different types of stress, we can observe different roles in the involvement of autophagy, whether it is disposal of poorly folded proteins, remobilization of nutrients or antioxidant mechanism. Autophagy is also especially important for the process of stress memory and regulation.
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