|
|
Functional characterization of plant EXO70 exocyst subunit isoforms and their membrane targeting mechanisms
Sekereš, Juraj ; Potocký, Martin (advisor) ; Jaillais, Yvon (referee) ; Ischebeck, Till (referee)
Vesicle tethering complex exocyst is one of the key regulators of the cell polarity and morphogenesis in eukaryotes. The complex interacts with the secretory vesicle, as well as plasma membrane, and facilitates formation of cis SNARE complex leading into fusion of the vesicle with target destination. Two of the eight exocyst subunits, the SEC3 and EXO70 are known to bind plasma membrane via protein and lipid interactors in Opisthokont model organisms. Genomes of angiosperm plants encode a surprisingly wide repertoire of EXO70 isoforms with over 20 present in both Arabidopsis and diploid tobacco genome. It has been proposed that different EXO70 isoforms would form parts of functionally distinct subtypes of the plant exocyst complex driving membrane trafficking to various membrane domains. Specific interactions of peripheral membrane proteins with particular membrane phospholipids largely contribute to targeting of cellular components to subcellular compartments and membrane domains. This thesis focuses on role of protein-lipid interactions in regulation of plant cell polarity and contributes to functional analysis of the plant EXO70 family diversity. We introduce the topic with the theoretical reviews summarizing role of protein-lipid interactions in establishing plant cell membrane domains at...
|
|
|
Function of phospholipases D and lipid phosphate phosphatases in the regulation of plant cell morphogenesis
Bezvoda, Radek ; Žárský, Viktor (advisor) ; Hašek, Jiří (referee) ; Vaňková, Radomíra (referee)
of the thesis The presented work explores the function and regulation of intracellular signaling that utilizes phospholipase D (PLD) and phosphatidic acid (PA), especially in the context of cellular morphogenesis of plants. PLDs cleave membrane phospholipids to phosphatidic acid, which has important biophysical and signaling role in many contexts, such as stress response, regulation of cytoskeletal dynamics and vesicular transport. Vesicular transport is essential in focused tip growth of plant pollen tubes and root hairs. Part of the work deals with NADPH oxidases, that are an emerging counterpart of PLD/PA signaling. Tobacco pollen tubes served as the main experimental model, as it enables assessing of changes in secretory pathway after pharmacological or genetic treatments. A technique utilizing antisense oligonucleotides was used for selective knock-down of PLD isoforms, NADPH oxidase and newly studied family of lipid phosphate phosphatases (LPPs) in pollen tubes. This enabled to assess functions of individual isoforms. For studying of selected gene families, various bioinformatic tool were utilized, such as dendrogram construction, analysis of available expression data and creating of virtual proteome. These tools together enabled to select potentially important genes for further experimental...
|
|
|
Function of phospholipases D and lipid phosphate phosphatases in the regulation of plant cell morphogenesis
Bezvoda, Radek ; Žárský, Viktor (advisor) ; Hašek, Jiří (referee) ; Vaňková, Radomíra (referee)
of the thesis The presented work explores the function and regulation of intracellular signaling that utilizes phospholipase D (PLD) and phosphatidic acid (PA), especially in the context of cellular morphogenesis of plants. PLDs cleave membrane phospholipids to phosphatidic acid, which has important biophysical and signaling role in many contexts, such as stress response, regulation of cytoskeletal dynamics and vesicular transport. Vesicular transport is essential in focused tip growth of plant pollen tubes and root hairs. Part of the work deals with NADPH oxidases, that are an emerging counterpart of PLD/PA signaling. Tobacco pollen tubes served as the main experimental model, as it enables assessing of changes in secretory pathway after pharmacological or genetic treatments. A technique utilizing antisense oligonucleotides was used for selective knock-down of PLD isoforms, NADPH oxidase and newly studied family of lipid phosphate phosphatases (LPPs) in pollen tubes. This enabled to assess functions of individual isoforms. For studying of selected gene families, various bioinformatic tool were utilized, such as dendrogram construction, analysis of available expression data and creating of virtual proteome. These tools together enabled to select potentially important genes for further experimental...
|
|
|
Subunits of exocyst complex in the development of Arabidopsis epidermis
Vojtíková, Zdeňka ; Žárský, Viktor (advisor) ; Soukup, Aleš (referee)
Exocyst is protein complex evolutionary conserved in yeasts, animals and plants, which plays a role in control of cell morphogenesis and polarity. It is a tethering complex whose function is to attach secretory vesicles to specifi c foci on plasma membrane. Complex exocyst is formed by eight subunits. Subunit EXO70 is encoded by 23 paralogue genes in genome of Arabidopsis thaliana. Mutation in paralogue subunit EX070H4 causes defect in trichome maturation. Mutant trichomes have thin, not reinforced cell wall, making them soft and elastic. Transcription of EXO70H4 gene is induced by UV radiation, therefore observations of plants cultivated on UV-B radiation were done. Analysis of mutants cultivated on UV-B radiation revealed hyperaccumulation of vesicules in cytoplasm, which were visible by light microscope. Hyperaccumulation was not observed in control plants cultivated on UV-B radiation, but thickening of cell wall was induced. Th is reaction to UV in trichomes hasn't been described yet. Analysis of cellular localization made with YFP tagged constructs revealed that EXO70H4 localizes into mobile corpuscules associating with Golgi apparatus. It was found with yeast two hybrid system that EXO70H4 interacts with TRS120, subunit of tethering complex TRAPPII which is active in Golgi apparatus....
|
guest ::
