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Mechanisms of establishment and maintenance of PIN polarity in Arabidopsis
Glanc, Matouš ; Friml, Jiří (advisor) ; Grebe, Markus (referee) ; Luschnig, Christian (referee)
Cell polarity is a key concept in plant biology. The subcellular localization of Pin- formed (PIN) auxin efflux carriers in the root of "#$%&'()*&* is remarkably asymmetrical, making PINs prominent markers to study cell polarity. In spite of its developmental importance and two decades of research, the molecular basis of PIN polarity remains largely unknown. In this thesis, I employed advanced transgenic and fluorescence microscopy approaches to gain insight into several aspects of PIN polarity regulation. I participated in establishing a novel genetically encoded inhibitor of endocytosis, an invaluable tool for the study of the importance of endocytosis for various cellular processes, including PIN polarity. I demonstrated that apical polarity of PIN2 needs to be re-established after cell division and that this process depends on endocytosis, '+!,(-( protein secretion and the action of WAG1 and related protein kinases, but not transcytosis, cell-cell signaling or intact cytoskeleton. Finally, I identified the previously unknown role of MAB4/MEL proteins in PIN polarity, which lies in the ability of MAB4/MELs to reduce PIN lateral diffusion and thus contribute to PIN polarity maintenance. My results, besides broadening current understanding of PIN polarity regulation, identify mechanisms that...
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The evolution of auxin homeostasis mechanisms
Skokan, Roman ; Petrášek, Jan (advisor) ; Buschmann, Henrik (referee) ; Holzinger, Andreas (referee)
The evolution of auxin homeostasis mechanisms Ph.D. thesis Roman Skokan, 2021 Abstract The streptophyte lineage consists of land plants (embryophytes) and several groups of primarily freshwater green algae called charophytes. While the phytohormone auxin is a conserved regulator of land plant development, little has been known of the possible origins of auxin response mechanisms in charophytes. We found that one of these mechanisms, the cellular auxin efflux via the PIN family of transport proteins, is most likely a deeply conserved feature in streptophytes. Additionally, we investigated the state of conservation in the green lineage (Viridiplantae) of the gene families known to be involved in auxin transport in land plants. We revealed that some families are deeply conserved outside land plants, but not others. We also helped uncover a unique radiation within the PIN family in Charophyceae. Striving to uncover the native significance of auxin transport in charophytes, we discovered a growth response to exogenously-applied auxin in Closterium, though the effort to produce stable mutant lines in the native PIN homolog is still underway. Altogether, we brought important insights into the evolution of auxin transport and response in the streptophyte lineage, though many questions still remain.
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The role of auxin in streptophyte algae
Schmidt, Vojtěch ; Petrášek, Jan (advisor) ; Pleskot, Roman (referee)
The phytohormone auxin is an important morphogen with an essential role in the development of land plants, where mechanisms of its action are well described. However, its role in green algae is poorly understood. Land plants are part of the phylum Streptophyta together with six closely related groups of predominantly freshwater green algae (charophytes). So far, the knowledge about the evolutionary origins of auxin action mechanism is mainly based on genomic information, and much less on experimental findings. In this work, the presence of auxin, its precursor, and catabolism products were shown in representative species of charophytes with varying levels measured compounds both produced endogenously and into the culture media. Thus, we gained a comprehensive insight into the possible strategies of auxin homeostasis across the non-land plant streptophytes. Also, an effect of exogenous auxin on the cell morphology and culture growth of the desmid Closterium was investigated. Image analysis of IAA-treated cells revealed a rather pleiotropic effect on cell morphology. The culture growth was inhibited by IAA. Additionally, IAA induced malformations in cell shape, and the extent of this phenomenon across individual cultures was dependent on the culture growth status. Lastly, we optimized the method of...
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Visualization of root apoplastic pH in plants
Wernerová, Daša ; Fendrych, Matyáš (advisor) ; Paris, Nadine (referee)
Plant oriented movements, or tropisms allow the plant to actively respond to environmental stimuli to get more light, better access to nutrients and to grow roots deeper into the soil. Gravitropism drives the growth of roots along the gravity vector. Perception of gravity is triggered by the sedimentation of statoliths in columella root cap, but the exact signalling pathway behind this process is not known. Perception of gravity results in an unequal redistribution of the phytohormone auxin in the outer cell layers which leads to different rate of growth on the root's upper and lower side and bending of the root. The changes in auxin redistribution are accompanied by changes in apoplastic pH. Knowing an exact pattern of these pH changes could shed light on the mechanisms laying behind the gravitropic response pathway. While microelectrodes can be used to measure pH precisely, they are not suitable for the long-term imaging of growing roots. In the past few years, several pH sensitive dyes and genetically encoded sensors emerged. These can be used for long-term live in vivo imaging of pH changes in growing roots. In this thesis, I analysed the performance of several published pH sensitive genetically encoded sensors and available dyes in the roots of Arabidopsis thaliana. I observed that dyes varied...
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Dynamics and role of the Arabidopsis thaliana IAA17/AXR3 protein in regulation of root growth by auxin
Kubalová, Monika ; Fendrych, Matyáš (advisor) ; Glanc, Matouš (referee)
Auxin is phytohormone that regulates several developmental processes and environmental responses. One of the most well-described outcome of the auxin signalling pathway is regulation of gene transcription. Aux/IAA proteins play an important role in this process, acting as transcriptional repressors. Recent studies revealed that several root growth responses are too rapid to be explained by changes in the level of transcription. The correlation between the amount of Aux/IAAs and the root growth rate suggests that these proteins might be involved in root growth regulation, especially during rapid growth responses that are not associated with transcriptional reprogramming. This work is focused on one of the 29 Arabidopsis Aux/IAA proteins - the IAA17/AXR3 protein. First, we produced stable transgenic lines of Arabidopsis thaliana expressing different combinations of fluorescently labelled AXR3-1 proteins and/or fused to subcellular localization tags under the control of different tissue-specific promoters, in order to characterize the subcellular localization of the studied protein. Subsequent visualization by confocal microscopy methods confirmed information about the role of IAA17/AXR3 protein in root growth responses, its involvement in auxin signalling, and gravitropism. Next, we showed that the...
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Řízkování vybraných kultivarů konopí v řízených podmínkách
Jakešová, Kristýna
The work subject is a general description of hemp (Cannabis sativa) and research on the practice of cutting, which greatly affects the production of hemp. The object of the research was to determine the effect of auxins in certain proportions and concentrations on rooting of cuttings of two varieties (Bedrocan and Bedrolite), which were imported from the Dutch company Bedrocan. We used auxins as indole-3-acetic acid (IAA), indole-3-butyric acid (IBA) and 1-naphthaleneacetic acid (NAA).
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Expresní profily vybraných genů při vyrůstání kotylárních pupenů hrachu
Psotová, Martina
The diploma thesis was aimed to monitor the expression profiles of selected genes in the growing of cotyledonary buds in pea (Pisum sativum L.). The apical stem was decapitated and 0.5% auxin paste was applied to the stump. From the treated plants cotyledonary buds were collected in intervals 0, 1, 3, 6, 12, 24, 48 and 120 hours. The biological material was analyzed for expression of genes PsDRM1, PsBRC1, PsAUX1, PsPIN1, PsYUC1. A gene expression normalization factor was calculated based on constitutive genes PsEF1-alpha, PsActin and Ps beta-tubulin. Application of auxin to the stem stump prevented the growth of cotyledonary buds. This inhibition of cotyledonary buds was accompanied by increased expression of dormancy markers PsDRM1, PsBRC1 and auxin biosynthesis gene PsYUC1 and decreased expression of the auxin transporter genes PsPIN1 and PsAUX1.
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Regulácia vyrastania kotylárnych pupeňov hrachu
Kucsera, Attila
This work was aimed to achieve more details in regulation of cotyledonary bud outgrowth of pea plants (Pisum sativum L.). Plants were studied after decapitation of the apex and removal of cotyledons and their subsequent substitution with agar gels containing different susbstances. Experiments were perform in light and dark conditions. The obtained results proved that auxin could regulate outgrowth of pea cotyledonary buds. Furthermore, sucrose showed partly inhibitory effect on bud outgrowth. This inhibitory effect was pronounced if plants were cultivated in dark. Next experiments showed that cotyledons could regulate apical dominance depending on light or dark conditions.
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Růstové korelace u rostlin konopí setého
Michálková, Klára
The teoretical part of my bachelor thesis Growth correlations in cannabis plant is mainly focused on the issue of growth correlations. In the bachelor thesis, concrete examples of growth correlations are characterized including more details on problematics of apical dominance of the stem. The basic botanical, morphological and genetic characteristics of cannabis are also described, including important content substances. In the experimental part, the intensity of shoot branching of intact plants of selected technical and medical cannabis genotypes is investigated. The increase in length of axillary buds of first, second and third node was measured. Within the period of a few days, plant response to decapitation and exogenous auxin application was tested. In technical cannabis there was a strong response to exogenous application of auxin. On the other hand in the most cases of medical cannabis there was a reduced sensitivity to exogenous application of auxin. On the topic of shoot branching it was observed that the branching of medical hemp genotypes is significantly more intensive compared to the varieties od technical cannabis. The results of the grafting experiment then imply that the rootstock of technical cannabis did not influence the intensity of shoot branching of the medical cannabis scion.
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Vliv vybraných stanovištních faktorů na proteolytickou aktivitu lesních půd s důrazem na organické a anorganické látky
Holík, Ladislav
PhD thesis focuses on soil biochemistry, namely the effects of organic and inorganic substances on proteolytic ktivity in forest soils. The thesis is composed of several impacted journal articles and two manuscripts. Selected organic compounds feature L- and D-tartaric acid, phenolics, proteinogenic and non-proteinogenic amino acids, auxins and cytokinins, whilst inorganic substances are represented by salts. Furthermore, an article dealing with the effects of different forest management practises on soil enzymatic ktivity (including proteases) is adjoined. Its aim is to monitor the effect of forest regenerative elements (forest stand gaps) on enzymatic ktivity, whereas the intervention into forest ktivity changes the moisture and thermal conditions of the site. The results show that out of the organic substances tested L- and D-tartaric acid, proteinogenic and non-proteinogenic amino acids and auxins have a positive effect on the proteolytic ktivity. Conversely, phenols and cytokinins acted as inhibitors. Soil salinization also had a negative effect on proteolytic ktivity of forest soils. The Influence of forest management through forest regenerative elements on soil protease ktivity was not detected. Potential urease ktivity was the only enzyme that reacted to the change of climatic conditions of forest habitat; however, ktiv not a principal topic of this work.
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