Results of two studies are reported here, a greenhouse study and a field study. In the greenhouse study, dry matter yield and nitrogen (total and 15N) uptake of leaves, stems and roots of tomato plants (Lycopersicum esculentum Mill., cv. Columbia) subjected to saline stress by NaCl were studied. The integrated effects of responses of these tissues to salinity on the whole plant basis and levels of Na⁺, Cl⁻ and K⁺ accumulation in these tissues were also studied. The treatments consisted of low (control, 0.3 bar), medium (4.3 bars), and high (8.3 bars) salinity. The saline treatments were prepared by adding NaCl to nutrient solution in sand culture. The plants were 80 days old at the start of the treatments and each was in a pot containing 1.8 kg of quartz sand. The ¹⁵N was provided to plants by adding K¹⁵NO₃ to the pots and the 15N treatment continued with the saline treatments up to 30 days. The plants were harvested at each 5-day interval during the treatment period. Dry matter production and nitrogen (total and ¹⁵N) uptake were significantly lower for saline treatments as compared with the control. Differences in dry matter production and ¹⁵N uptake on whole plant basis appeared in the latter part of the treatment period between the two saline treatments. For most of the parameters studied, the leaves were found to be affected most by salinity, the roots were intermediate in their response and the stems were the least affected by salinity. The effect of salinity on the studied parameters were attributed to osmotic effects and specific ion effects of Na⁺ and/or Cl⁻. A field study with two cultivars--Columbia and Pearson was conducted at the Safford Agricultural Center. Three N treatments were used: 0 kg N/ha, 84 kg N/ha and 168 kg N/ha and two treatments consisting of two water sources--river water with an EC of 1.15 dS/m and more saline well water of EC of 2.21 dS/m. Columbia had a significantly higher yield of tomatoes than Pearson for both water types. The N treatments had no effect on tomato yield apparently due to high residual N remaining in the field from the previous crop. Commercially acceptable fresh market yields were approached with both varieties and waters in spite of moderate salinity and sodium under heavy textural soil conditions, high temperatures and the presence of certain diseases in the area.

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Includes bibliographical references

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Salinity stress predisposed Penncross creeping bentgrass to cottony blight caused by P. aphanidermatum. Studies were conducted on the effects of salinity on the mycelial growth of P. aphanidermatum and on the growth of Penncross bentgrass. Mycelial growth increased significantly up to Ec levels of 7.1 ds/m when compared to mycelial growth at the control Ec levels of 0.5 ds/m. Plant growth was reduced to 50% of the control at Ec levels of 4.3 ds/m. 3-month-old Penncross bentgrass plants were inoculated and incubated at two differing temperatures. At 32 C, all plants died within 3 days regardless of salinity treatment. Rates of plant death were greatest at salinity levels over 2.8 ds/m. At 27 C, plants irrigated with water at Ec levels from 4.3 to 7.1 ds/m showed complete necrosis within 5 days, while treatments irrigated with tap water showed no disease symptoms.

鹽鹼化是世界上最主要非生物脅迫之一，它主要是由於土壤中的鹽（氯化鈉）過量積累所導致的, 不僅影響植物的生長而且也影響農作物的產量。大豆是世界上最重要的經濟類豆科植物之一，由於其種子內含有豐富的營養物質例如蛋白質，油，糖分和纖維，所以它為我們提供了極為重要和大量的油脂和蛋白。在鹽脅迫下大豆的產量會有明顯的降低。由於以上這些情況，我們希望搞清楚大豆對於鹽脅迫的反應機制。首先是通過蛋白質組學弄清楚鹽脅迫的生理過程和大豆如何耐受鹽脅迫的。一般來說，蛋白質組學包括了鑑定蛋白類的化合物和測量在生物系統中的蛋白含量的學科。近期，質譜的發展提供了一個去研究細胞內蛋白質的動態變化十分有用的平台。定量蛋白質組學的發展對於我們系統性的了解蛋白質的功能的分子是十分重要的並且預期會提供給我們各種生物過程和系統的分子機制的見解.其次，表觀遺傳性特別是組蛋白修飾。因為組蛋白修飾通過重新編排染色體的結構和組成參與了許多重要的生物過程並且這些翻譯後修飾對於植物的耐受鹽脅迫也發揮著十分重要的作用。因此我們希望了解組蛋白修飾是如何參與這一過程的。 / Salinity stress, which is caused by the accumulation of excessive amount of salts in the soil, is one of the most severe abiotic stresses that constraint not only crop plant growth but also crop productivity. Soybean (Glycine max) is one of the most important economical legume crops in the world because of its richness of nutritional compositions including protein, oil, sugar and fiber in the seed. Soybean yield of sensitive cultivars is decreased dramatically under salt stress. Because of this, we tried to figure out the mechanism of how soybean response to salinity stress. Firstly, Proteomics--to elucidate the affected physiological process in the salinity stress and the way to tolerant the stress. In general, proteomics involves the identification of protein components and the measurement of protein abundance in biological systems. Recent mass spectrometry (MS)-based technology developments have provided useful platforms for the study of quantitative changes in protein components within the cell. Quantitative proteomics is important for the system-based understanding of the molecular function of each protein component and expected to provide insights into molecular mechanisms of various biological processes and systems. Secondly, Epigenetics--particularly histone modifications. Because histone modifications play important roles in many fundamental biological processes by rearranging the structure and composition of chromatin and PTMs have more roles in response salinity stress. So we want to understand how PTMs involve in this process from epigenetics. / Detailed summary in vernacular field only. / Peng, Xu. / "October 2012." / Thesis (M.Phil.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 44-54). / Abstracts also in Chinese. / Thesis committee --- p.i / Declaration --- p.ii / Abstract (in English) --- p.iii / Abstract (in Chinese) --- p.iv / Acknowledgements --- p.v / Table of contents --- p.vi / General abbreviations --- p.vii / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1 --- Soybean --- p.1 / Chapter 1.2 --- Salinity stress and plants’ response to salinity stress --- p.1 / Chapter 1.3 --- Mass Spectrometry base Proteomics --- p.3 / Chapter 1.3.1 --- Introduction to proteomics --- p.3 / Chapter 1.3.2 --- Proteomic studies in plants --- p.5 / Chapter 1.4 --- Introduction to Epigenetics --- p.6 / Chapter 1.5 --- Present studies --- p.8 / Chapter Chapter 2 --- Proteomic studies in soybean --- p.9 / Chapter 2.1 --- Materials and methods --- p.9 / Chapter 2.1.1 --- Plant materials and stress treatment --- p.9 / Chapter 2.1.2 --- Protein extraction and nuclei extraction and histone isolation --- p.10 / Chapter 2.1.3 --- Protein Preparation for Mass Spectrometry --- p.11 / Chapter 2.1.4 --- Analysis of Protein using nanoLC-MS/MS and Data analysis --- p.12 / Chapter 2.2 --- Results and discussions --- p.13 / Chapter 2.2.1 --- One thousand two hundred seventeen proteins were identified by LC MS/MS-based proteomics technique --- p.13 / Chapter 2.2.2 --- Functional analyses of identified proteins --- p.22 / Chapter 2.2.3 --- One hundred sixty-three proteins are changed under salinity stress as identified by LC MS/MS-based proteomics technique --- p.26 / Chapter 2.2.4 --- Important stress relate proteins were identified by LC MS/MS-based proteomics technique --- p.30 / Chapter 2.2.5 --- Histone PTMs in soybean under salinity stress --- p.34 / Chapter Chapter 3 --- Conclusions and perspectives --- p.35

大豆的豐富營養和經濟價值使它成為重要的農產品。但是，土壤鹽漬化影響著大豆的產量。這個問題在沿岸地方特別嚴重。若要改善大豆的耐鹽能力，必先增加對大豆耐鹽機理的了解。 / 本實驗室從大豆中發現了一個受鹽脅迫誘導表達的基因GmSAL1。以往透過體外酶反應分析法， GmSAL1蛋白被介定為一個能作用於1,4,5-三磷酸肌醇 (IP₃) 的肌醇磷酸-5-磷酸酶。這有別於在擬南芥中的SAL1同源蛋白AtSAL1, AtSAL1是一個肌醇磷酸-1-磷酸酶。由於IP₃是信號傳導途徑中的重要分子，本課題對與IP₃信號及耐鹽性相關的GmSAL1蛋白功能進行研究。 / 本課題旨在：（一）　研究GmSAL1對於細胞質內IP₃的累積的體內作用； （二） 研究 GmSAL1 在脫落酸 (ABA)　的信號傳導中的可能角色； （三） 研究 GmSAL1 在鹽脅迫下的保護作用。 / 本研究利用體內報告系統證明了 GmSAL1 對於減少細胞質內IP₃的累積的作用。這種影響IP₃水平的功能減弱了由 ABA 信號所引起的氣孔關閉和種子萌發抑制。利用 GmSAL1 轉基因煙草細胞 (BY-2)　和擬南芥，證明 GmSAL1 在鹽脅迫下起著短暫的保護作用。GmSAL1在鹽脅迫下的保護功能可能是由於蒸騰作用的局部恢復和細胞鈉離子區室化的作用。 / 本研究展示了肌醇信號，ABA信號和鹽脅迫反應三者之間的關係。這是在以前的研究中未被清楚闡釋的。 / Soybean is nutritionally and economically important. However, high soil salinity, particularly in coastal regions, impedes the production of soybean. Understanding the salt tolerance mechanism is the first step towards the enhancement of salt tolerance of soybean. / Our laboratory identified a salt-responsive gene from soybean namely GmSAL1. Previous in vitro enzyme assay suggested that the GmSAL1 protein is an inositol 5’-phopsphatase acting on inositol 1,4,5-trisphosphate (IP₃), which is different from the enzymatic activity reported for the SAL1 homologue (AtSAL1) in Arabidopsis thaliana (A. thaliana) which is an inositol 1-phopsphatase. Since IP₃ is an important molecule involved in signal transduction pathways, this project is to explore the in vivo functions of GmSAL1 in relation to IP3 signaling and salinity tolerance. / The specific objectives of this research are (1) to study the in vivo role of GmSAL1 on cytosolic IP₃ accumulation; (2) to study the possible involvement of GmSAL1 in ABA signaling; (3) to study the protective roles of GmSAL1 under salt stress. / In this project, the function of GmSAL1 to reduce cytosolic IP₃ was demonstrated using an in vivo reporter system. This activity on IP₃ levels reduced the sensitivity of stomatal closure and seed germination inhibition mediated by ABA signals. A transient protection effect against the ionic effect under salt stress by GmSAL1 was shown by gain-of-function tests in transgenic BY-2 cells and transgenic A. thaliana. The protective effect conferred by GmSAL1 may be due to a partial resuming of transpiration through reduction of ABA signals and compartmentalization of Na⁺ into vacuoles. / The study of GmSAL1 in this research demonstrated the link among inositol signaling, ABA signaling, and salinity response which was not well addressed in previous reports. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Ku, Yee Shan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 97-104). / Abstracts also in Chinese. / Statement --- p.i / Abstract --- p.ii / 摘要 --- p.iv / Acknowledgements --- p.v / General Abbreviations --- p.vii / Abbreviations of Chemicals --- p.ix / Table of Contents --- p.xi / List of Figures --- p.xviii / List of Tables --- p.xxi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- General introduction to salinity and agriculture --- p.1 / Chapter 1.1.1 --- Adverse effects of salinity on plants --- p.2 / Chapter 1.1.1.1 --- Osmotic stress --- p.2 / Chapter 1.1.1.2 --- Ionic stress --- p.3 / Chapter 1.1.1.3 --- Separation of ionic effect from osmotic effect --- p.3 / Chapter 1.1.1.4 --- Oxidative stress --- p.4 / Chapter 1.1.2 --- Major physiological responses of plants to achieve salt tolerance --- p.5 / Chapter 1.1.2.1 --- Maintenance of cellular ion homeostasis --- p.5 / Chapter 1.1.2.2 --- Balance between Na⁺ and K⁺ influx --- p.5 / Chapter 1.1.2.3 --- Efflux of Na⁺ from cell --- p.9 / Chapter 1.1.2.4 --- Enhanced compartmentalization of Na⁺ and Cl⁻ in vacuole --- p.11 / Chapter 1.1.2.5 --- Enhanced vacuolar ion compartmentalization --- p.13 / Chapter 1.1.2.6 --- Biosynthesis of osmolytes --- p.13 / Chapter 1.2 --- Signal transduction under salt stress --- p.14 / Chapter 1.2.1 --- General introduction to signal transduction under salt stress --- p.14 / Chapter 1.2.2 --- ABA signaling under salinity --- p.15 / Chapter 1.2.2.1 --- General introduction to ABA signaling --- p.15 / Chapter 1.2.2.2 --- IP₃ and ABA signaling --- p.16 / Chapter 1.2.2.3 --- Introduction to inositol phosphate --- p.16 / Chapter 1.2.2.4 --- Phosphatidylinositol-3-monophosphate --- p.18 / Chapter 1.2.2.5 --- Phosphatidylinositol-4-monophosphate --- p.18 / Chapter 1.2.2.6 --- Phosphatidylinositol-5-monophosphate --- p.18 / Chapter 1.2.2.7 --- Phosphatidylinositol (3,5) bisphosphate --- p.19 / Chapter 1.2.2.8 --- Phosphatidylinositol (4,5) bisphosphate --- p.19 / Chapter 1.2.2.9 --- Inositol (1,4,5) trisphosphate (IP₃) --- p.19 / Chapter 1.2.2.10 --- Inositol metabolism under salt stress --- p.19 / Chapter 1.2.2.11 --- The involvement of IP₃ in ABA signaling --- p.20 / Chapter 1.2.3 --- General introduction to Ca²⁺ signaling --- p.22 / Chapter 1.2.4 --- Ca²⁺ channels --- p.23 / Chapter 1.2.4.1 --- Ligand-gated Ca²⁺ channels --- p.23 / Chapter 1.2.4.1.1 --- IP₃ gated Ca²⁺ channels --- p.23 / Chapter 1.2.4.1.2 --- Cyclic nucleotide gated channels (CNGCs) --- p.24 / Chapter 1.2.4.1.3 --- Glutamate receptor homologs (GLRs) --- p.24 / Chapter 1.2.4.2 --- Voltage-gated Ca2⁺ channels --- p.25 / Chapter 1.2.4.2.1 --- Two-pore channels (TPCs) --- p.25 / Chapter 1.2.4.2.2 --- Mechanosensitive Ca2²⁺permeable channels (MSCCs) --- p.25 / Chapter 1.2.4.2.3 --- Ca²⁺ and ABA signaling --- p.26 / Chapter 1.2.5 --- ABA, IP₃ and Ca²⁺ signaling --- p.26 / Chapter 1.2.5.1 --- Ca²⁺ signaling under salt stress --- p.30 / Chapter 1.2.5.2 --- Ca²⁺ signal mediated cellular responses --- p.30 / Chapter 1.3 --- Introduction to inositol phosphatases --- p.30 / Chapter 1.3.1 --- Previous studies on inositol phosphatases in plant --- p.33 / Chapter 1.3.1.1 --- Inositol polyphosphate 1-phosphatase --- p.33 / Chapter 1.3.1.2 --- Inositol polyphosphate 5-phosphatase --- p.34 / Chapter 1.4 --- Previous research on GmSAL1 in Prof. Hon-Ming Lam’s lab --- p.37 / Chapter 1.5 --- Objective and Significance of this project --- p.38 / Chapter Chapter 2 --- Materials and methods --- p.39 / Chapter 2.1 --- Materials --- p.39 / Chapter 2.1.1 --- Plants, bacterial strains and vectors --- p.39 / Chapter 2.1.2 --- Chemicals and enzymes --- p.40 / Chapter 2.1.3 --- Buffer, medium and solution --- p.41 / Chapter 2.1.4 --- Primers --- p.41 / Chapter 2.1.5 --- Equipments and facilities --- p.44 / Chapter 2.1.6 --- Software --- p.44 / Chapter 2.2 --- Methods --- p.44 / Chapter 2.2.1 --- Measurement of osmolarity --- p.44 / Chapter 2.2.2 --- Plant growth and treatment conditions --- p.45 / Chapter 2.2.2.1 --- NaCl, PEG and ABA treatment on soybean plant --- p.45 / Chapter 2.2.3 --- Artificial crossing of A. thaliana --- p.46 / Chapter 2.2.3.1 --- Screening of double homozygous transgenic A. thaliana lines --- p.46 / Chapter 2.2.4 --- Transformation of tobacco BY-2 cells --- p.47 / Chapter 2.3 --- Molecular techniques --- p.48 / Chapter 2.3.1 --- DNA extraction --- p.48 / Chapter 2.3.2 --- PCR --- p.48 / Chapter 2.3.2.1 --- Screening of transgenes --- p.48 / Chapter 2.3.2.2 --- Synthesis of DIG-labeled DNA probe for northern blot analysis --- p.49 / Chapter 2.3.3 --- DNA gel electrophoresis --- p.49 / Chapter 2.3.4 --- RNA extraction --- p.50 / Chapter 2.3.5 --- Northern blot analysis --- p.51 / Chapter 2.3.5.1 --- RNA treatment --- p.51 / Chapter 2.3.5.2 --- Electrophoresis --- p.51 / Chapter 2.3.5.3 --- RNA blotting --- p.51 / Chapter 2.3.5.4 --- GmSAL1 mRNA detection --- p.52 / Chapter 2.4 --- Cell viability assay --- p.52 / Chapter 2.5 --- Na⁺ compartmentalization assay --- p.53 / Chapter 2.6 --- ABA sensitivity assays --- p.53 / Chapter 2.6.1 --- Seed germination assay --- p.53 / Chapter 2.6.2 --- Stomatal opening assay --- p.54 / Chapter Chapter 3 --- Results --- p.55 / Chapter 3.1 --- Differential response of GmSAL1 expression level to NaCl and PEG treatment --- p.55 / Chapter 3.2 --- The expression of GmSAL1 in host plant is responsive to ABA --- p.59 / Chapter 3.3 --- Effect of GmSAL1 on cytosolic IP₃ level in vivo --- p.62 / Chapter 3.4 --- Overexpression of GmSAL1 down-regulates in planta IP₃ level in guard cell --- p.62 / Chapter 3.5 --- Ectopic expression of GmSAL1 in A. thaliana alters stomatal aperture in the presence of ABA in a Ca²⁺ dependent manner --- p.67 / Chapter 3.6 --- Ectopic expression of GmSAL1 in A. thaliana reduces the ABA inhibitory effect on seed germination --- p.71 / Chapter 3.7 --- Overexpression of GmSAL1 transiently protects A. thaliana against ionic effect under salinity --- p.76 / Chapter 3.8 --- Overexpression of GmSAL1 enhances the survival of tobacco BY-2 cells under salt treatment but not near iso-osmotic PEG treatment --- p.80 / Chapter 3.9 --- Overexpression of GmSAL1 confers enhanced vacuolar compartmentalization of Na⁺ in NaCl treated BY-2 cells --- p.85 / Chapter Chapter 4 --- Discussion --- p.90 / Chapter 4.1 --- GmSAL1 as a novel inositol 5-phosphatase --- p.90 / Chapter 4.2 --- The effect of GmSAL1 expression on ABA signaling --- p.91 / Chapter 4.3 --- Involvement of GmSAL1 in tolerance toward ionic effect under salt stress --- p.92 / Chapter 4.4 --- The protective function of GmSAL1 under salinity --- p.93 / Chapter Chapter 5 --- Conclusion --- p.96 / References --- p.97 / Chapter Appendix I --- Chemicals --- p.105 / Chapter Appendix II --- Formulations of buffer, medium and solution --- p.107 / Chapter Appendix III --- Equipments and facilities --- p.110 / Chapter Appendix IV --- Osmolarity of solutions --- p.111 / Chapter Appendix V --- Result of biological repeat of northern blot analysis of GmSAL1 in soybean leaf under NaCl --- p.113 / Chapter Appendix VI --- Result of biological repeat of northern blot analysis of GmSAL1 in soybean root under NaCl --- p.114 / Chapter Appendix VII --- Result of biological repeat of northern blot analysis of GmSAL1 in soybean leaves under 100μM ABA treatment for 0hr, 0.5hr, 1hr, 2hr and 4hr --- p.115 / Chapter Appendix VIII --- Result of biological repeat experiment on the survival rate of tobacco BY-2 cell in 150mM NaCl supplemented MS medium --- p.116 / Chapter Appendix IX --- Result of biological repeat experiment on the survival rate of tobacco BY-2 cell in 13.3% PEG-6000 supplemented MS medium --- p.117

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