|
|
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...
|
|
|
Specificity of selected exocyst subunits in trichome development
Glanc, Matouš ; Žárský, Viktor (advisor) ; Binarová, Pavla (referee)
Trichomes are fine epidermal outgrowths covering aerial organs of most land plants. Although unicellular trichomes of Arabidopsis thaliana have long been used as a model system in plant cell and developmental biology, surprisingly little is known about the processes involved in cell wall biogenesis during the last stage of trichome maturation. A role of EXO70H4, a putative subunit of the vesicle tethering complex exocyst, in trichome maturation has recently been identified in our laboratory. Image analysis, histochemical detection and FT-IR spectroscopy methods were used in this study to analyze cell wall defects of the exo70H4 LOF mutant, revealing the mutation causes altered deposition of pectins and possibly also lignins and hemicelluloses. Transgenic lines with EXO70 paralogues driven by the EXO70H4 promoter were prepared and their analysis revealed that the closest paralogue EXO70H3, unlike EXO70A1 and EXO70B1, can complement the exo70H4 mutation. Based on the results, questions concerning trichome cell wall composition, the role of EXO70H4 in trichome maturation and functions of the plant exocyst complex are discussed. Keywords: Arabidopsis, trichome, cell wall, secretory pathway, exocyst complex, EXO70H4, FT-IR spectroscopy
|
|
|
Secretory pathway in plant cell wall polysaccharide biosynthesis and FT-IR spectroscopy methods in plant cell wall composition analysis
Glanc, Matouš ; Žárský, Viktor (advisor) ; Votrubová, Olga (referee)
All plant cells are encapsulated in a cell wall that determines the cells' shape and size and is essential to many of their vital processes. The cell wall of streptophyte plants is composed mainly of polysaccharides of high molecular weight. Cellulose, the main constituent of the plant cell wall, is synthesized by protein complexes bound to the PM, while hemicelluloses and pectins are synthesized in the Golgi apparatus. Several proteins that participate in the synthesis of cell wall polysaccharides have been identified, but hundreds of them remain to be discovered. Secretory pathway plays an important role in plant cell wall biogenesis as it transports cellulose synthase complexes and noncellulosic polysaccharide molecules to the cell surface. Some regulatory mechanisms that might be involved in cell wall material secretion, such as actin cytoskeleton, Ca2+ gradient or PIP kinases have been proposed; however, the regulation of this process is very complex and far from being understood. FT-IR spectroscopy is a method that can detect molecular vibrations and provide information about chemical composition of virtually all substances. It has been used successfully in screens for cell wall mutants, the study of interactions between cell wall polymers, as well as other areas of cell wall biology. FT-IR...
|
|
|
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...
|
|
|
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...
|
|
|
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...
|
|
|
Specificity of selected exocyst subunits in trichome development
Glanc, Matouš ; Žárský, Viktor (advisor) ; Binarová, Pavla (referee)
Trichomes are fine epidermal outgrowths covering aerial organs of most land plants. Although unicellular trichomes of Arabidopsis thaliana have long been used as a model system in plant cell and developmental biology, surprisingly little is known about the processes involved in cell wall biogenesis during the last stage of trichome maturation. A role of EXO70H4, a putative subunit of the vesicle tethering complex exocyst, in trichome maturation has recently been identified in our laboratory. Image analysis, histochemical detection and FT-IR spectroscopy methods were used in this study to analyze cell wall defects of the exo70H4 LOF mutant, revealing the mutation causes altered deposition of pectins and possibly also lignins and hemicelluloses. Transgenic lines with EXO70 paralogues driven by the EXO70H4 promoter were prepared and their analysis revealed that the closest paralogue EXO70H3, unlike EXO70A1 and EXO70B1, can complement the exo70H4 mutation. Based on the results, questions concerning trichome cell wall composition, the role of EXO70H4 in trichome maturation and functions of the plant exocyst complex are discussed. Keywords: Arabidopsis, trichome, cell wall, secretory pathway, exocyst complex, EXO70H4, FT-IR spectroscopy
|
|
|
Secretory pathway in plant cell wall polysaccharide biosynthesis and FT-IR spectroscopy methods in plant cell wall composition analysis
Glanc, Matouš ; Žárský, Viktor (advisor) ; Votrubová, Olga (referee)
All plant cells are encapsulated in a cell wall that determines the cells' shape and size and is essential to many of their vital processes. The cell wall of streptophyte plants is composed mainly of polysaccharides of high molecular weight. Cellulose, the main constituent of the plant cell wall, is synthesized by protein complexes bound to the PM, while hemicelluloses and pectins are synthesized in the Golgi apparatus. Several proteins that participate in the synthesis of cell wall polysaccharides have been identified, but hundreds of them remain to be discovered. Secretory pathway plays an important role in plant cell wall biogenesis as it transports cellulose synthase complexes and noncellulosic polysaccharide molecules to the cell surface. Some regulatory mechanisms that might be involved in cell wall material secretion, such as actin cytoskeleton, Ca2+ gradient or PIP kinases have been proposed; however, the regulation of this process is very complex and far from being understood. FT-IR spectroscopy is a method that can detect molecular vibrations and provide information about chemical composition of virtually all substances. It has been used successfully in screens for cell wall mutants, the study of interactions between cell wall polymers, as well as other areas of cell wall biology. FT-IR...
|
guest ::
