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Characterization of promoter regions of HGSNAT and GBA genes, and a contribution to the study of pathogenesis of MPS IIIC and Gaucher disease
Richtrová, Eva ; Hřebíček, Martin (advisor) ; Macek, Milan (referee) ; Adam, Tomáš (referee)
Pathogenesis of mucopolysaccharidosis type IIIC (MPS IIIC) and Gaucher disease has not been yet fully clarified, and the causes of phenotypical variability between the patients with the same genotype in Gaucher disease remain obscure. Because the variants in the regulatory regions of genes can cause phenotypical differences mentioned above, I have studied promoter regions of HGSNAT and GBA genes mutated in these lysosomal disorders. I have shown that there is an alternative promoter of GBA (P2). Additional studies were aimed to elucidate possible physiological functions of P2, and its possible role in the pathogenesis of Gaucher disease. I have found that P2 is not tissue specific, and that its variants do not influence the variability of phenotype in Gaucher patients with the same genotype. P2 is used differentially neither during the differentiation of monocytes to macrophages nor in macrophages from controls and Gaucher patients, in whom there is a prominent storage only in cells of macrophage origin. We have thus not found any changes that would suggest a role for P2 in the pathogenesis of Gaucher disease. I have characterized the promoter region of HGSNAT and shown that the binding of Sp1 transcription factor is important for its expression. Sequence variants found in HGSNAT promoter in...
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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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Vesicular trafficking into the plant vacuole
Semerádová, Hana ; Kulich, Ivan (advisor) ; Vosolsobě, Stanislav (referee)
Vacuole is very important plant cell organelle which can occupy 90% of cell volume. It provides wide range of functions. Considering enormous size of the vacuole, vesicle trafficking into the plant vacuole is major vesicle movement inside the cell. Transport pathway into the vacuole is very dynamic field of plant cell biology. This sorting machinery shares similarities within all eukaryotes, but plants also have their own specificities. Soluble cargo is targeted through secretory pathways by vacuolar sorting receptors (VSRs). These proteins due to its transmembrane localization can interact with sorted cargo and take it to the right organelle within the cell. Membrane fusion is facilitated with Rab-GTPase and SNARE protein complexes. A special kind of vesicle traffic is autophagy, the self consuming process that protects the cell from various type of stress or enables apoptosis.
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The significance of autophagy and its communication with the apoptotic machinery for cellular survival or cell death
Pazour, Vítězslav ; Anděra, Ladislav (advisor) ; Černý, Jan (referee)
Autophagy is a cellular proces, taht allows degradation of a portion of cytoplasm, protein aggregates or entire organelles. Major function of autophagy is the maintainance of cellular homeostasis, the protection against stress and mobilization of internal resources. However, autophagy also has a role in imunity, development and differentiation. Autophagic signaling can interact with apoptotic machinery at several levels via regulatory proteins of both pathways or via mutual degradation or cleavage of the components of both pathways. Autophagy can communicate with both extrinsic and intrinsic pathways of apoptosis. Under certain circumstances can autophagy by itself also induce cell death. Autophagic cell death called also programmed cell death of type II is accompanied by massive vacuolization and lysosomal autodestruction of the affected cell. Autophagic cell death was documented during Drosophila development but also in mammalian cells. Autophagy also play importamt role in tumorogenesis, where it can either protect tumor cells against various stresses or it can contribute to their death. Further research of autophagic signaling and mechanisms of communication between autophagy and apoptosis may ont only extend our knowledge on these essential processes but can also contribute to cancer therapy. Powered...
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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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The role of melatonin in SIRT1 and p-AMPK regulation in HT-29 cell line
Shkut, Anastasiya ; Ramos Mandíková, Jana (advisor) ; Svobodová, Hana (referee)
Charles University in Prague Pharmaceutical Faculty in Hradec Králové Department of Pharmacology and Toxicology Student: Anastasiya Shkut Supervisors: Mgr. Jana Mandíková, Virginia Motilva Ph.D. Title of diploma thesis: The role of melatonin in SIRT1 and p-AMPK regulation in HT-29 cell line. Sirituin 1 (SIRT1) is NAD+ dependent deacetylase present in wide range of organisms including mammals. It was found to extend life span in yeasts during calorie restriction (CR) conditions. SIRT1 deacetylates many regulator proteins and thus control metabolic status of cell as well as AMP-activated kinase (AMPK), which is also energy regulator enzyme depending on NAD+ levels in cell. Both of them play roles in cancer and could influence autophagy, although the exact mechanism remains unclear. We focused our study on hormone melatonin, which has anti-inflammatory and anti-cancer effects, to study its influence on human colon cancer cell line HT-29. If it has impact on SIRT1 and AMPK and what is hierarchic relationship between SIRT1 and AMPK. Also we observed its possible influence on autophagy. We used Western blotting (WB) technique and from our results it seems that melatonin has significant effect on SIRT1, which might activate AMPK as well as autophagy. Nevertheless some of results did not have sufficient...
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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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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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Mitochondrial production of reactive oxygen species and its role in physiological regulations
Holzerová, Eliška ; Mráček, Tomáš (advisor) ; Kalous, Martin (referee)
The production of mitochondrial reactive oxygen species and the resulting oxidative stress is an important phenomenon driving long-lasting research and intense discussions. Knowledge of exact mechanisms of reactive oxygen species production and pathways leading to their formation could help us to directly affect their production, a task with potential terapeutic implications. The molecular nature of the production of reactive oxygen species by some enzymes has already been well documented, but others still remain controversial and current theories are obviously far from the truth. Much more interesting is the question of physiological importace of this production. The reactive oxygen species were considered harmful factors clearly distorting the integrity of the organism for a long time. However, recent research suggest that their existence can also be beneficial and effective. Evidently they can serve as a signaling molecules in several metabolic and regulatory pathways occurring in the organism. This bachelor thesis offers insight into the current state of knowledge. It focuses on the most detailed description of the reactive oxygen species production by mitochondrial respiratory chain enzymes. Furthermore, it deals with some signaling cascades, where involvement of mitochondrially generated...
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