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ARP2/3 complex interaction with plant secretory pathway components
Voloshina, Mariia ; Schwarzerová, Kateřina (advisor) ; Oulehlová, Denisa (referee)
The ARP2/3 complex is an evolutionarily conserved protein complex involved in actin nucleation. In animals, the loss of function of ARP2/3 subunits is usually lethal. In contrast, arp2/3 plants are viable and exhibit certain morphological defects. One of the characteristic traits of arp2/3 plants is impaired cell adhesion. This suggests a role for the ARP2/3 complex in plant cell wall secretion, specifically pectins. However, the exact molecular mechanism of ARP2/3 function in this pathway has not been elucidated yet. Preliminary results suggest a possible interaction of ARP2/3 with the secretory pathway proteins GNL1, COPI, and COG. This work combines two methods - TIRFM/VAEM and co-immunoprecipitation - to study the interaction of ARP2/3 with selected markers. However, neither colocalization nor physical interaction was demonstrated. This work also characterises COPI, COG and GNL1 with TIRFM/VAEM and shows that there is an interaction between GNL1 and endocytic markers. Key words: ARP2/3, COPI, COG, GNL1, secretory pathway
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Thermal imaging as a tool for evaluation of plant's cooling effect in green façades
Nohová, Michaela ; Lhotáková, Zuzana (advisor) ; Schwarzerová, Kateřina (referee)
Urbanisation and global warming are gaining momentum, and with it the need to improve the thermal comfort of urban citizens is growing. Due to the geometry of the city and the thermal properties of the materials from which the city consists, the so-called urban heat island effect occurs, which causes the temperature in city centres to be higher than in rural areas. One possible way to mitigate the urban heat island effect is to incorporate vegetation into the urban infrastructure. Green architecture is designed to do just that, and in this thesis I specifically focus on green facades and their cooling effect. Since green facades are usually large areas of green surface, this thesis focuses on thermal imaging and its use to measure the cooling rate of buildings and ambient air by green facades. It is the use of thermal cameras that is characterized by the ability to image large areas and record their temperature in real time, making thermal cameras an ideal tool for green facade research. As this topic is very specific and not yet so widespread in the literature, I discuss thermography from several perspectives in the context of its use for research on plant physiological parameters, and I also discuss the various methods and pitfalls of working with thermal image data. Key words: abiotic stress,...
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Cell inclusions in microscopic eukaryotes
Pilátová, Jana ; Schwarzerová, Kateřina (advisor) ; Gierlinger, Notburga (referee) ; Prášil, Ondřej (referee)
This thesis reports, for the first time, a systematic study identifying various cell inclusions across the broad diversity of all major eukaryotic supergroups. Raman microspectroscopy technique was employed as a powerful technique, which is becoming the method of the first choice for such studies. In Chapter 1, the spatial reconstruction of chloroplasts of the unicellular zygnematophyte Cylindrocystis sp. (Streptophyta) revealed "empty spaces" inside. Subsequent analysis showed massive accumulations of polyphosphate, which is not located in the cytoplasm as in other species, but inside the chloroplasts. This newly discovered ability of the zygnematophytes, a sister group of terrestrial plants, to accumulate polyphosphate leads us to speculate about a possible adaptation to extreme Arctic conditions or even a preadaptation leading to plant terrestrialization. Chapter 2 summarizes the serendipitous discovery of purine crystalline inclusions in dinoflagellates and other microalgae. Investigated dinoflagellate species comprised zooxanthellae, the endosymbionts of reef-building corals, and the species causing toxic algal blooms. Purine crystals were further shown to act as dynamic high-capacity nitrogen storage. To show how widespread purine inclusions are, a revision of crystalline inclusions in all...
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Lignin and lignification mechanisms
Gargoš, Ondřej ; Tylová, Edita (advisor) ; Schwarzerová, Kateřina (referee)
Lignin is an essential compound of all vascular plants. Recent studies have also shown its presence in non-vascular plants. Lignin is important for plant growth and development, provides a mechanical support to plant tissues and is also a major player in the response of plants to various stresses. Lignin is an aromatic heteropolymer and comprises classical lignin units - guaiacyl (G), syringyl (S) and p-hydroxyfenyl (H). The amount and ratio of these lignin units varies among plant species, organs and cell types. Nontraditional monomer units that can be deposited in lignin also increase the variety of lignin. Lignification, i.e. the formation and deposition of lignin, is a complex and precisely controlled process involving the synthesis of monolignols in the cytoplasm, their transport into the cell wall and subsequent polymerization. A number of enzymes are involved in the lignification process, and recent studies are gradually revealing the mechanisms of lignification regulation in various cells of plant body. This bachelor thesis summarizes the knowledge of the importance, synthesis and deposition of lignin, and also compares the mechanisms regulating lignification in different types of plant cells - tracheary elements, sclerenchyma and endodermal cells. An interesting phenomenon of lignification...
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The role of ARPC2 in plant cells
Šlajcherová, Kateřina ; Schwarzerová, Kateřina (advisor) ; Klíma, Petr (referee)
ARPC2 protein localization in a plant cell Kateřina Šlajcherová 1 Abstract Actin cytoskeleton is an ubiquitous structure which plays numerous irreplacable roles. Actin nucleation is, beside formins, performed by ARP2/3 complex (actin-related protein), comprising of seven subunits (ARP2, 3, C1-C5) and activated by protein SCAR/WAVE complex. ARP2/3 complex is attached to the membrane and branches existing microfilaments, apart from nucleating them de novo. ARP2/3 mutants in most organisms show severe defects. However, plant mutants exhibit only mild phenotype, for example, Arabidopsis thaliana ARPC2 mutant (dis2-1) has deformed trichomes and leaf epidermal cells, but its viability is not impaired. The aim of the thesis is to map ARPC2 localization within the cell and broaden our understanding of ARP2/3 complex role in plant cell morphogenesis. Tobacco ARPC2 (NtArpC2) subunit was visualized in Arabidopsis plants, using the GFP fusion protein as well as imunofluorescence and anti-ARPC2 antibody. Experiments were undertaken to collocalize the subunit with actin and microtubular cytoskeleton, with mitochondrions, endosomes and other membrane organelles. The specimens were observed in confocal and TIRF microscope. The GFP-NtARPC2 protein shows as motile dots; their movement, but not their existence, is dependent...
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The effect of Arp2/3 complex on vacuolar structure, cytoplasmic streaming and dictyosome motility.
Semerák, Matěj ; Schwarzerová, Kateřina (advisor) ; Eliášová, Kateřina (referee)
In plant cells, actin filaments are nucleated in two different ways: The growth of single filaments or their bundles is enabled by various types of formins, whereas branched meshworks emerge due to Arp2/3 complex activity. Mutations in genes of these nucleators lead to various phenotypic traits. This thesis deals in the first place with influence of Arp2/3 complex subunits' dysfunction on intracellular motility (cytoplasmic streaming, stop-and-go movements of Golgi apparatus cisternae), since it had not been extensively studied before, and also attempts to quantify the already known impacts of mutations in genes for ARP2 and ARPC5 subunits on the vacuolar morphogenesis. For comparison, a few experiments with plants which carried a mutation in gene for FH1 formin were also realised when measuring the cytoplasmic streaming. The experiments were conducted with a model plant Arabidopsis thaliana. The methods particularly included transformation with fluorescent markers by Agrobacterium tumefaciens (or usage of a fluorescent dye), microscopy (both standard and confocal) and subsequent evaluation of the acquired data using a computer. During the cytoplasmic streaming research, effects of cytoskeletal drug latrunculin B were studied, too. The outputs did not prove that the Arp2/3 complex defects would manifest...
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Aluminum toxicity in plant cell
Schiebertová, Lucie ; Schwarzerová, Kateřina (advisor) ; Tylová, Edita (referee)
Aluminium being the third most abundant metal in the earth's crust is in its toxic form a serious threat for crop productivity in acid soils, which comprise almost half of the arable land. As the most phytotoxic form is considered free ion Al3+ , which affects root growth by acting in the root apical zone, resulting in growth inhibition in a very short time at micromolar concentrations. At cellular and molecular level, many cell components are affected by aluminium toxicity including cell wall, plasma membrane, signal transduction pathways, calcium homeostasis, DNA and numerous cytoplasmic enzymes. Although it is difficult to distinguish the primary targets from the secondary effects so far, understanding of the target sites of aluminium toxicity is helpful to elucidate the mechanisms by which aluminium exerts its deleterious effects in root growth. On the other hand, some plant species have evolved mechanisms to cope with aluminium toxicity. In the future, the attention should be paid to basic mechanisms of aluminium toxicity and our understanding of the current problem should be unified. Key words: Al toxicity, Al resistance/tolerance, phytotoxicity, Al stress, acid soils, root elongation
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Actin nucleation in plant cell
Schiebertová, Petra ; Schwarzerová, Kateřina (advisor) ; Cvrčková, Fatima (referee)
The ability of plant cells to nucleate actin is important especially for dynamic of intracellular movements of organelles and vesicles, cell structure and growth organization and coordination of endo- and exocytosis . Actin nucleation means the development of new actin filaments from G-actin. Actin associated proteins - Arp2/3 complex and formins serve for this purpose. Arp2/3 complex polymerizes new "daughter " filament from the side of the "parent" filament at an angle of 70 degrees and after the nucleation remains at minus end of filaments, thus inducing branching of filaments. Nucleation by Arp2/3 complex further requires nucleation promoting factors - NPFs. Deletion of subunits of Arp2/3 complex is often lethal in animal cells. On the other hand, deletion results in rather mild phenotype in plants. Formins catalyze the formation of direct actin fibers and remain on the plus end of the fiber after nucleation. There are multiple isoforms of formins in plants than in animals, which suggest an important role of formins in plants. New actin nucleators Cobl, Lmod, Spire, JMY and APC containing WH2 (WASP homology 2) domain were identified in non-plant cells. This type of actin nucleation was not described in plants. Kew words: Actin, Arp2/3 complex, formins, Scar/WAVE, Cobl, Lmod, Spire
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Role of cytoskeleton in plant cell morphogenesis
Miklánková, Pavlína ; Schwarzerová, Kateřina (advisor) ; Sekereš, Juraj (referee)
The cells are able to acquire variety of shapes, in which cytoskeleton plays an important role. Cytoskeleton influences deposition of cell wall materials, regulates vesicle movement in cell, participates in exocytosis and endocytosis. Cortical microtubules affect celulose accumulation in cell wall and determine direction of cell expansion, although the exact connection between microtubules and cellulose remains unclear. Actin promotes growth and contributes to its spatial regulation in both tip ang diffuse growing cells. Actin is important for secretion in expanding cells but its exact functions in cell growth regulation are not explained yet. Analysis of mutants, spectroscopic methods, cytoskeletal drugs, fluorescence proteins and other methods are used to better understand how actin and microtubule cytoskeleton are integrated during plant cell morphogenesis. Epidermal and trichome cells of Arabidopsis thaliana are a good model of research and they are used for most studies.
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