Drought is a major environmental factor limiting crop productivity. Considering a significant area of crop production under water-limited rained conditions, there is a great need to develop production systems to sustain yield potentials under drought stress. Silicon has recently been recognized as an important element in plant nutrition. In this study, it was shown that supplying soybean with soluble silicon in the soil could improve vegetative growth and drought tolerance under water limiting conditions. In order to understand the molecular mechanism how silicon alleviates drought stress, the effects of silicon application on protein expression and antioxidant enzymes were examined. Soybean plants were grown in sand-containing pots supplied with 4 millimolar solutions of sodium silicate. To cancel the effect of sodium, the same amount of sodium chloride was used along with control plants. Soluble proteins were isolated from the leaves and roots of silicon-treated and control plants subjected to water deficit stress. Two-dimensional gel electrophoresis and mass spectrometry approaches were used to identify differentially expressed leaf and root proteins in response to silicon application under water deficit stress. Proteins that showed differential expression in response to silicon application included metabolic enzymes and proteins involved in the proteasome-dependent degradation pathway. These results indicate that silicon application could affect enzymes important for carbohydrate metabolism and stabilize aldehyde dehydrogenases and malic enzyme under water deficit stress, which may be attributable to drought tolerance.

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京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第18599号 / 農博第2086号 / 新制||農||1027(附属図書館) / 学位論文||H26||N4874(農学部図書室) / 31499 / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 三上 文三, 教授 植田 充美, 教授 栗原 達夫 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Occurrence of organic micropollutants (OMPs) in aquatic environment is a worldwide concern. A long list of anthropogenic substances, including pharmaceuticals, hormones, etc., are frequently detected in natural water sources. Wastewater treatment plants are one main source of OMPs pollution, but also a key step to control OMPs dissemination into the environment. This dissertation focuses on the evaluation of Membrane Aerated Biofilm Reactor (MABR) as a sustainable process to treat wastewater polluted by OMPs. Furthermore, application of high intensity pulsed light is proposed as an innovative tertiary treatment to produce reclaimed water of high quality. In Chapter 1, a literature review was performed to investigate the occurrence and toxicity of 12 selected organic micropollutants (OMPs) in surface and ground water and the limitations of current available biological processes. Among these technologies, systems with enriched nitrifying activity were able to enhance the removal of specific OMPs through cometabolic activities. Thus, I proposed the use of a MABR with enriched nitrifying biomass to treat OMP polluted water. In Chapter 2, I studied the influence of biofilm thickness on the removal of 13 OMPs via MABR. Results demonstrated OMP removal was dependent on biofilm thickness and bacterial cell density. MABR demonstrated important efficiencies in the removal of ammonium, COD, acetaminophen and triclosan at early stages of biofilm thickness. However, the removal of nonpolar, hydrophobic 4 OMPs and anionic, acidic OMPs required thicker biofilms, achieving maximum removal at biofilm with 1.02 mm thickness and 2.2 × 106 cell mL-1. In Chapter 3, the contribution of sorption and biodegradation in the removal of OMPs via MABR was evaluated. At three stages of biofilm thickness studied, biodegradation dominated the removal for most OMPs. Heterotrophs played an important role in OMP biodegradation at all biofilm thickness, while autotrophic nitrifiers enhanced their contribution at thickness beyond 0.58 mm. Increased removal of pollutants like estrone and ethinyl estradiol were linked to the MABR enrichment with nitrifying bacteria. Sorption was essential for the removal of lipophilic and recalcitrant pollutants like triclosan. Finally, to provide high quality treated water for reuse, Chapter 4 explores the use of high-intensity pulsed light (HIPL) as post-treatment. The number of pulses and optical energy dose have a significant impact on the OMPs removal. HIPL demonstrated fast kinetics and efficient photodegradation – with significant OMPs removal within milliseconds. The findings from my Ph.D. dissertation indicate that MABR combined with high-intensity pulse light may be an effective treatment train for the efficient removal OMPs present in municipal wastewaters. This combined treatment process could potentially pave the way to produce high quality reclaimed water for various reuse purposes.

Der Abbau des Chlorophyll (Chl) ist ein Prozess, der typischerweise während der Blattseneszenz und der Reifung von Früchten und Samen stattfindet. Eine Störung dieses koordinierten Prozesses unter Frostbedingungen verzögert den Chl-Abbau und ist ein grosses Hindernis bei der Herstellung von hochwertigem Rapsöl. Der Abbau von Chl zu farblosen Kataboliten erfolgt in einer Serie von enzymatischen Schritten und wird durch die Chlorophyllase begonnen (Chlase). Es wurde vorgeschlagen, dass ein wasserlösliches Chl Protein (WSCP) den Transport des Chl von der Thylakoidmembran zum Wirkort der Chlase übernimmt. Weiterhin wurde angenommen, dass die Steigerungen der Genexpressionen dieser frühen Schritte den Prozess des Chl-Abbaus beschleunigen. In der vorliegenden Arbeit werden die Auswirkungen der Überexpression der Chlase aus Citrus clementii (CcCHLASE) und von WSCP aus Blumenkohl (Cau-WSCP) in transgenen Tabakpflanzen analysiert. Dazu wurde die cDNA Sequenz der CcCHLASE in E. coli exprimiert und mittels in vitro Experimenten die Hydrolysierung von Chl durch die Chlase bestätigt. Anschließend wurden CcCHLASE exprimierende Tabakmutanten generiert und drei T1-Linien wurden unter verschiedenen Stress- und Seneszenzbedingungen untersucht. Die Chlase überexprimierenden Linien zeigten unter allen getesteten Bedingungen einen im Vergleich zum Wildtyp erhöhten Chlide a Gehalt. Trotzdem unterschied sich die Menge an Endkataboliten in diesen Mutanten nicht vom Wildtyp. Andererseits zeigten WSCP überexprimierende Linien zwar keine erhöhten Chlide a Gehalte jedoch erhöhte Protochlorophyllid-(Pchlide)-Level. Das deutet auf eine Rolle des WSCP als Speichermolekül für Chlorophyllvorstufen hin. Die photoprotektive Funktion des WSCP wurde zusätzlich in WSCP überexprimierenden Linien bestätigt. Diese zeigen im Vergleich zu Wildtyp-Tabakpflanzen auch bei hohen Lichtintensitäten von 700 – 900 µmol Photonen m-2 s-1 verringerte Gehalte an Zeaxanthin und reduzierte Peroxidaseaktivitäten. / Chlorophyll (Chl) catabolism is characteristically seen during leaf senescence, fruit ripening and seed maturation. Disruption of this coordinated process under frost conditions delays Chl breakdown and is a great concern in rapeseed oil production. The present work addresses this problem by studying the effect of enhanced Chl catabolism in genetically modified tobacco plants. Chl is catabolised to colourless catabolites through a series of enzymatic reactions initiated by Chlorophyllase (Chlase). A water soluble chlorophyll protein (WSCP) has been proposed to transport Chl from thylakoid membranes to the site of action of Chlase. It was assumed that enhancing the gene expression of these early events in Chl catabolism would increase the Chl breakdown process. The present work analysed the overexpression of Chlase from Citrus clementii (CcCHLASE) and WSCP gene from cauliflower (Cau-WSCP) in modified tobacco plants. Initially, the cDNA sequence of CcCHLASE was expressed in E. coli and in vitro tests confirmed the hydrolytic activity of Chlase on Chl. Subsequently, tobacco plants overexpressing CcCHLASE were generated and three T1 lines were analysed at various stress and senescence conditions. The in vivo production of Chlorophyllide (Chlide) indicated the extent of increased Chl breakdown. The Chlase overexpressor lines showed higher Chlide a steady state levels under all tested conditions in comparison to the WT tobacco plants. However, the end catabolites did not show much difference from WT plants. On the other hand, WSCP overexpressor lines did not show any increase in Chlide a levels, but demonstrated an increased protochlorophyllide (Pchlide) levels. This suggested the role of WSCP as a storage molecule of Chl precursors. Additionally, photoprotective function of WSCP was confirmed in WSCP overexpressors, by lower zeaxanthin levels and peroxidase activity even at high light intensities of 700 – 900 µmol photons m-2 s-1 in comparison to the WT tobacco plants.