Nitrogen fixation is synchronized with carbon metabolism in Lotus japonicus and Medicago truncatula nodules under salt stress

Abstract

In the present work, the response to NaCl applied at the vegetative stage to Medicago truncatula and Lotus japonicus has been evaluated in order to ascertain whether the effect of salt stress on nitrogen fixation is due to a limitation on nodular carbon metabolism. Results show maximum sucrose synthase (SS) and alkaline invertase (AI) activities were obtained at the vegetative stage, when maximum nitrogenase activity was detected in both species. SS activity decreased with the salt treatment, providing evidence of the regulatory role of this enzyme for the carbon supply to the bacteroids. Phosphoenolpyruvate carboxylase (PEPC) and malate dehydrogenase (MDH) activities could account for higher nitrogen fixation efficiency detected in L. japonicus nodules and isocitrate dehydrogenase (ICDH) activity compensated for the carbon limitations that occur under salt stress. These results support that nitrogenase inhibition in nodules experiencing salt stress is doubt to a carbon flux shortage, as result of carbon metabolism enzymes activities down-regulation.     

盐胁迫下树种幼苗生长及其耐盐性

采用盆栽方法,以11个树种实生幼苗为材料,用不同浓度(0、3、5、8 g·kg-1和10 g·kg-1)NaCl溶液进行1次性浇灌处理,对盐胁迫下各树种的形态表现、生长及耐盐性进行了研究,结果表明:(1)当盐含量达到8 g·kg-1时,欧洲荚蒾、甜桦和光叶漆植株死亡,当含量增加到10 g·kg-1时,沃氏金链花植株死亡,其它各存活树种也均出现不同程度的盐害症状;(2)盐胁迫后,各树种的苗高生长量下降、生物量累积减少,且随着处理浓度的增加均呈下降趋势,其中榆桔、甜桦和光叶漆的降幅最大;(3)盐处理后,各树种的根冠比值增大,其中盐胁迫对光叶漆、银水牛果和沃氏金链花有显著影响(p<0.05);(4)综合分析各树种的生长和形态表现,认为日本丁香、银水牛果、三裂叶漆和豆梨具有高度耐盐性,沃氏金链花、金雀儿、鹰爪豆和榆桔具有中高度耐盐性,而欧洲荚蒾、甜桦和光叶漆具有中度耐盐性.     

Ion‐dependent metabolic responses of Vicia faba L. to salt stress

Salt‐affected farmlands are increasingly burdened by chlorides, carbonates, and sulfates of sodium, calcium, and magnesium. Intriguingly, the underlying physiological processes are studied almost always under NaCl stress. Two faba bean cultivars were subjected to low‐ and high‐salt treatments of NaCl, Na₂SO₄, and KCl. Assimilation rate and leaf water vapor conductance were reduced to approximately 25–30% without biomass reduction after 7 days salt stress, but this did not cause severe carbon shortage. The equimolar treatments of Na⁺, K⁺, and Cl^? showed comparable accumulation patterns in leaves and roots, except for SO₄^2? which did not accumulate. To gain a detailed understanding of the effects caused by the tested ion combinations, we performed nontargeted gas chromatography–mass spectrometry‐based metabolite profiling. Metabolic responses to various salts were in part highly linearly correlated, but only a few metabolite responses were common to all salts and in both cultivars. At high salt concentrations, only myo‐inositol, allantoin, and glycerophosphoglycerol were highly significantly increased in roots under all tested conditions. We discovered several metabolic responses that were preferentially associated with the presence of Na⁺, K⁺, or Cl^?. For example, increases of leaf proline and decreases of leaf fumaric acid and malic acid were apparently associated with Cl^? accumulation.     

盐胁迫对4树种叶片中K^＋和Na^＋的影响及其耐盐能力的评价

通过盆栽试验,对我国南方银杏（Ginkgo biloba L.)、侧柏[Patycladus orentalis(L.)Franco]、火炬松（Pinus taeda L.)和剌槐(Robinia Pseudoacacia L.)4造林树种苗木叶片中K^ 、N^ 浓度浓度和N^ ／K^ 比进行测定,盐处理水平为0.0％（CK）、0.1%、0.3%和0.5％。随着盐浓度的提高,各树种叶片中Na^ 浓度和N^ ／K^ 有逐渐升高的趋势,但K^ 浓度变化较小;随着盐胁迫时间的推移,各树种叶片中Na^ 、K^ 浓度和Na^ /K^ 比都没有明显的变化规律。方差分析和多层比较表明,侧柏、火炬松和剌槐叶片中Na^ 、K^ 浓度和N^ ／K^ 比在4组处理间的差异均达显著或极显著水平。4树种中剌槐和侧柏的耐盐能力最强,银杏次之,火炬松最弱。     

Grafting influence on the weight and quality of tomato fruit under salt stress

Two commercial tomato cultivars were used to determine whether grafting could prevent decrease of fruit weight and quality under salt stress conditions. The cultivars Buran F1 and Berberana F1 were grafted onto rootstock ‘Maxifort’ and grown under three levels of elevated soil salinity (EC 3.80 dS m^?1, 6.95 dS m^?1 and 9.12 dS m^?1). Fruit weight reduction of grafted plants was lower (about 20–30%) in comparison with non‐grafted ones. Salt stress at the second salinity level (EC 6.95 dS m^?1) induced the highest alteration of examined growth and quality parameters. The total increase of phenols, flavonoids, ascorbate and lycopene content in the fruits of both grafted and non‐grafted plants for both cultivars had a similar trend and intensity, though some inter‐cultivar variation was observed. The possibility of grafting tomato plants to improve salt tolerance without fruit quality loss is discussed.     

Protein pattern changes in tomato under in vitro salt stress

The investigation of salt-induced changes in the proteome would highlight important genes because of a high resolution of protein separation by two-dimensional gel electrophoresis (2-DE) and protein identification by mass spectrometry and database search. Tomato (Lycopersicon esculentum Mill.) is a model plant for studying the mechanisms of plant salt tolerance. Seeds of tomato cv. Shirazy were germinated on water-agar medium. After germination, seedlings were transferred to Murashige and Skoog nutrient medium supplemented with 0, 40, 80, 120, and 160 mM NaCl. After 24 days, leaf and root samples were collected for protein extraction and shoot dry weight measurement. Alterations induced in leaf and root proteins under salt stress treatments were studied by one-dimensional SDS-PAGE. Leaf proteins were also analyzed by 2-DE. With increasing salt concentration in the medium, shoot dry weight decreased. SDS-PAGE showed induction of at least five proteins with mol wts of 30, 62, and 75 kD in roots and 38 and 46 kD in leaves. On the 2-DE gel, more than 400 protein spots were reproducibly detected. At least 18 spots showed significant changes under salt stress. Three of them corresponded to new proteins, while six proteins were up-regulated and five proteins were down-regulated by salt stress. In addition, salinity inhibited the synthesis of four leaf proteins. Ten spots were analyzed by matrix-assistant laser desorption/ionization-time of flight (MALDI-TOF), which led to the identification of some proteins, which could play a physiological role under salt stress. The expression of new proteins(enoyl-CoA hydratase, EGF receptor-like protein, salt tolerance protein, phosphoglycerate mutase-like protein, and M2D3.3 protein) under salt stress indicates that tomato leaf cells respond to salt stress by changes in different physiological processes. All identified proteins are somehow related to various salt stress responses, such as cell proliferation. Published in Russian in Fiziologiya Rastenii, 2007, Vol. 54, No. 4, pp. 526–533. The text was submitted by the authors in English.     

盐胁迫下白杨无性系苗木体内离子分配及比较

以白杨杂种无性系及其亲本当年生苗木为材料 ,分析了盐胁迫下无性系的离子含量 ,及 Na 、K 、Ca 在植物体内运输和分配特点 ,并对无性系间差异进行了比较。研究表明 ,随盐浓度提高 ,植物体内Na 含量迅速提高 ,K 、Ca 含量降低 ;盐分胁迫下 ,根部 Na 含量较高 ,叶片 Na 中含量最低 ,K 、Ca 含量则相反 ,特别是 Ca ,其分布顺序为叶 >茎 >根。杂种无性系 B430及其亲本新疆杨对 K 和 Ca 运输的选择性比毛白杨高 ,而对 Na 运输的选择性则比毛白杨低 ,从而导致根部存留的 Na 较多 ,叶片分配的 Na 数量较少 ,从而减轻 Na 对叶片的伤害。综合分析表明 B430和新疆杨耐盐能力最强 ,毛新杨其次 ,毛白杨最差。     

Application of NMR-based metabolomics to the investigation of salt stress in maize (Zea mays)

Introduction – High salinity, caused by either natural (e.g. climatic changes) or anthropic factors (e.g. agriculture), is a widespread environmental stressor that can affect development and growth of salt‐sensitive plants, leading to water deficit, the inhibition of intake of essential ions and metabolic disorders. Objective – The application of an NMR‐based metabolic profiling approach to the investigation of saline‐induced stress in Maize plants is presented. Methodology – Zea Maize seedlings were grown in either 0, 50 or 150 mM saline solution. Plants were harvested after 2, 4 and 6 days (n = 5 per class and time point) and ¹H NMR spectroscopy was performed separately on shoot and root extracts. Spectral data were analysed and interpreted using multivariate statistical analyses. Results – A distinct effect of time/growth was observed for the control group with relatively higher concentrations of acetoacetate at day 2 and increased levels of alanine at days 4 and 6 in root extracts, whereas concentration of alanine was positively correlated with the shoot extracts harvested at day 2 and trans‐aconitic acid increased at days 4 and 6. A clear dose‐dependent effect, superimposed on the growth effect, was observed for saline treated shoot and root extracts. This was correlated with increased levels of alanine, glutamate, asparagine, glycine‐betaine and sucrose and decreased levels of malic acid, trans‐aconitic acid and glucose in shoots. Correlation with salt‐load shown in roots included elevated levels of alanine, γ‐amino‐N‐butyric acid, malic acid, succinate and sucrose and depleted levels of acetoacetate and glucose. Conclusions – The metabolic effect of high salinity was predominantly consistent with osmotic stress as reported for other plant species and was found to be stronger in the shoots than the roots. Using multivariate data analysis it is possible to investigate the effects of more than one environmental stressor simultaneously. Copyright © 2010 John Wiley & Sons, Ltd.     

荒漠盐生植物根际土壤盐分和养分特征

中国西北地区是我国干旱、盐碱化土壤分布面积较广、土壤积盐较重的地区,这里发育着丰富的盐生植物。目前对于干旱荒漠区盐生植物根际特征的研究相对较少,而不同盐生植物的根际特征对于研究盐生植物适应盐渍环境的机制有着重要意义。本研究利采用盆栽根袋法对7种不同类型的荒漠盐生植物的根际盐分和养分特征进行了初步探索。结果表明:盐分在盐生植物根际发生富集,稀盐盐生植物和泌盐盐生植物根际土壤中总盐和8种主要盐分离子的含量都有所增加,而在拒盐盐生植物根际中增加不显著,其中Cl-和Na 的富集程度相对其它6种离子的富集程度要高。稀盐盐生植物和泌盐盐生植物根际土中的SO42-/Cl-比土体有显著的降低,表明在稀盐盐生植物和泌盐盐生植物根际土壤中Cl-的富集程度比SO42-高,拒盐盐生植物根际土盐分SO42-/Cl-比略有提高。7种盐生植物根际土中的Na /K ,Na /Ca2 ,Na /Mg2 比均较土体有显著的增加,芦苇根际土中的增加最小。在所有研究植物中,根际土壤中全N含量比土体的含量高,但全P和全K含量却比土体的含量低;根际土壤中有效态养分的变化则与全态相反,根际土壤中的有效N含量比土体中的都显著降低,除芦苇外,其他六种盐生植物根际土壤中有效P和有效K的含量都高于土体,但有效P的富集不及有效K富集的程度高。在研究的七种植物中,钠猪毛菜根际土壤的有效N亏缺量最高,有效P和速效K富集也最少。7种植物,尤其是稀盐盐生植物和泌盐盐生植物的地上部分的主要盐离子含量比地下部分高,如Cl-、Na 、Ca2 和K ,在根际富集程度最高的Cl-和Na ,在植株的地上部分也增加的最多。     

盐碱胁迫下燕麦生长及阳离子吸收特征

在吉林白城地区盐碱土上监测了3个盐分胁迫水平下燕麦的生长动态及对土壤阳离子的吸收特征.结果表明:在低度、中度和高度3个胁迫水平下燕麦地上部生长和产量构成因素没有显著差异,但根部干质量和根冠比随盐碱胁迫程度的加重而显著降低,在成熟期,中度和高度胁迫下的根冠比分别是低度胁迫下的77.2％和64.5％.3个胁迫水平下燕麦的K+/Na+和Ca2+/Na+在三叶期差异最大,而在抽穗期无显著差异.随着盐分胁迫程度的加重,燕麦的离子吸收选择性系数在灌浆期显著降低,而运输选择性系数无显著变化.因此,燕麦能够在一定程度上适应苏打碱土的盐碱胁迫,适应能力随着胁迫水平的增加而降低;燕麦根部生物量的减少和抽穗期较强的离子选择吸收能力可能有利于地上部的生长和产量构成.     

Growth and nitrogen fixation in Lotus japonicus and Medicago truncatula under NaCl stress: nodule carbon metabolism

Lotus japonicus and Medicago truncatula model legumes, which form determined and indeterminate nodules, respectively, provide a convenient system to study plant-Rhizobium interaction and to establish differences between the two types of nodules under salt stress conditions. We examined the effects of 25 and 50mM NaCl doses on growth and nitrogen fixation parameters, as well as carbohydrate content and carbon metabolism of M. truncatula and L. japonicus nodules. The leghemoglobin (Lb) content and nitrogen fixation rate (NFR) were approximately 10.0 and 2.0 times higher, respectively, in nodules of L. japonicus when compared with M. truncatula. Plant growth parameters and nitrogenase activity decreased with NaCl treatments in both legumes. Sucrose was the predominant sugar quantified in nodules of both legumes, showing a decrease in concentration in response to salt stress. The content of trehalose was low (less than 2.5% of total soluble sugars (TSS)) to act as an osmolyte in nodules, despite its concentration being increased under saline conditions. Nodule enzyme activities of trehalose-6-phosphate synthase (TPS) and trehalase (TRE) decreased with salinity. L. japonicus nodule carbon metabolism proved to be less sensitive to salinity than in M. truncatula, as enzymatic activities responsible for the carbon supply to the bacteroids to fuel nitrogen fixation, such as sucrose synthase (SS), alkaline invertase (AI), malate dehydrogenase (MDH) and phosphoenolpyruvate carboxylase (PEPC), were less affected by salt than the corresponding activities in barrel medics. However, nitrogenase activity was only inhibited by salinity in L. japonicus nodules.     

盐雾胁迫对舟山海岛7个造林树种存活和生长的影响

以普陀樟﹝Cinnamomum japonicum var. chenii ( Nakai) G. F. Tao〕、舟山新木姜子﹝Neolitsea sericea ( Bl.) Koidz.〕、红楠( Machilus thunbergii Sieb. et Zucc.)、全缘冬青( Ilex integra Thunb.)、石栎﹝Lithocarpus glaber ( Thunb.) Nakai〕、厚叶石斑木﹝Raphiolepis umbellata ( Thunb.) Makino〕和滨柃﹝Eurya emarginata ( Thunb.) Makino〕7个舟山海岛造林树种为研究对象,用浓度18 mg·g-1盐水通过盐雾发生装置每日喷雾0.0(对照)、0.5、1.0和2.0 h,并连续喷雾35 d,对翌年生长季各树种的形态特征、存活率和不同部位的干质量进行测定,并据此确定各树种的耐盐雾性。结果表明：随盐雾日处理时间及胁迫天数的延长,7个树种幼苗均表现出不同的受害症状并逐渐加重,但各树种的受害程度有差异。盐雾日处理0.5、1.0和2.0 h石栎幼苗的存活率,以及盐雾日处理2.0 h红楠、舟山新木姜子和滨柃幼苗的存活率均显著低于对照,但不同盐雾日处理时间对普陀樟、全缘冬青和厚叶石斑木幼苗的存活率均无显著影响。与对照相比,每日短时间的盐雾处理对普陀樟和全缘冬青幼苗的苗高和地径无显著影响,但随盐雾日处理时间延长二者的苗高和地径显著下降;盐雾日处理0.5、1.0和2.0 h,红楠、舟山新木姜子和滨柃幼苗的苗高总体上显著下降,厚叶石斑木幼苗的苗高则显著高于对照,但它们的地径总体上与对照无显著差异。每日短时间盐雾处理对普陀樟和滨柃幼苗单株的叶、根、茎干质量及总干质量均无显著影响,但随盐雾日处理时间的延长,二者单株的不同部位干质量及总干质量均逐渐下降;盐雾处理可导致舟山新木姜子、红楠、全缘冬青和石栎幼苗单株的不同部位干质量及总干质量显著或极显著下降,且降幅总体上随盐雾日处理时间的延长而增大,其中石栎幼苗单株的不同部位干质量及总干质量均最低;盐雾处理对厚叶石斑木幼苗生长则有一定的促进作用,使其单株的不同部位干质量及总干质量不同程度增加,但随盐雾日处理时间的延长增幅减小。盐雾处理对供试各树种叶、茎和根的质量比以及根冠比有不同的影响效应,其中盐雾胁迫造成各树种不同程度落叶,使多数树种根冠比增大,但全缘冬青和厚叶石斑木的根冠比则有所下降。根据实验结果,结合供试树种在舟山海岛的生长环境,初步确定厚叶石斑木的耐盐雾性最强,全缘冬青、普陀樟和滨柃的耐盐雾性次之,红楠和舟山新木姜子的耐盐雾性较弱,石栎的耐盐雾性在7个树种中最弱。     

Antioxidant enzymes and physiological characteristics in two Jerusalem artichoke cultivars under salt stress

The effects of NaCl stress on the activity of antioxidant enzymes, lipid peroxidation, cell membrane stability, net photosynthetic rate, gas-exchange, and chlorophyll content were investigated in two Jerusalem artichoke cultivars, Dafeng (salt-tolerant) and Wuxi (salt-sensitive), grown under control (nutrient solution) or salt stress (nutrient solution containing 75, 150, and 225 mM NaCl) conditions for 7 days. In leaves of salt-tolerant cv. Dafeng, superoxide dismutase (EC 1.15.1.1), peroxidase (EC 1.11.1.7), and catalase (EC 1.11.1.6) activities significantly increased as compared to the controls, whereas no significant change was observed in cv. Wuxi. Lipid peroxidation and cell membrane injury were enhanced in both cultivars. Net photosynthesis and stomatal conductance decreased in response to salt stress, but cv. Dafeng showed a smaller reduction in photosynthesis than cv. Wuxi. The results indicated that stomatal aperture limited leaf photosynthetic capacity in the NaCl-treated plants of both cultivars. However, significant reduction in the leaf chlorophyll content due to NaCl stress was observed only in cv. Wuxi. These results suggested that salt-tolerant Jerusalem artichoke varieties may have a better protection against reactive oxygen species, at least in part, by increasing the activity of antioxidant enzymes under salt stress.     

根环境供氧状况对盐胁迫下棉花幼苗光合及离子吸收的影响

以溶液培养的棉花(Gossypium hirsutum L.)幼苗为材料,测定了不同盐胁迫程度和不同根环境供氧状况条件下棉花幼苗的叶片气体交换参数、叶绿素荧光参数和植株的Na~+、K~+离子含量等的变化,以探索根环境供氧状况对盐胁迫下棉花光合作用和离子吸收的影响。结果表明,盐胁迫和根环境供氧不足均导致净光合速率下降。在处理后的前期,盐胁迫对棉花叶片光合作用的不利影响大于供氧不足(不通气)的影响,而后期根环境供氧不足的不利影响快速增大,并逐渐超过盐胁迫的影响。在低浓度盐胁迫和根环境不通气处理的初期,棉花叶片光合速率下降的主要原因是气孔因素(气孔关闭或部分关闭引起的CO_2供应不足);随着盐胁迫程度的增大和胁迫持续时间的延长,光合速率下降的原因逐渐转变为非气孔因素(光合系统损伤引起的光合能力下降)。相同程度盐胁迫下,根环境通气处理的棉花叶片的净光合速率和PSⅡ最大光化学效率等均显著高于根环境不通气处理的,说明根环境供氧不足加重了盐胁迫对光合作用的不利影响。对棉花植株各器官离子积累量的测定、分析发现,盐胁迫导致了棉花根系拒Na~+、吸K~+的能力和选择性运输K~+的能力降低,使棉花根系和叶片的Na~+含量增多、K~+含量减少、[Na~+]/[K~+]比值升高;而根环境通气则可显著提高盐胁迫下根系的拒Na~+、吸K~+能力和根系向叶片选择性运输K~+的能力,降低根系和叶片的[Na~+]/[K~+]比值。试验还发现,根系K~+、Na~+含量受盐胁迫的影响较大,而叶片K~+、Na~+含量受根环境通气状况的影响更大一些。综合分析可见,盐胁迫和根环境供氧不足均可导致棉花叶片光合速率下降、光合机构损伤以及离子平衡失调,而根环境通气可以缓解盐胁迫对棉花叶片光合作用的不利影响、增加根系和叶片对K~+的选择吸收和积累、降低[Na~+]/[K~+]比值,从而增强棉花植株对盐胁迫的适应性和抵抗力。     

Research on the adaptive mechanism of photosynthetic apparatus under salt stress: New directions to increase crop yield in saline soils

Photosynthesis is the largest organic synthesis on Earth, salinity limits crop yield and quality worldwide directly or indirectly related to the decrease in photosynthetic efficiency. The mechanism by which photosynthetic apparatus responds to salt stress is extremely complex and varies with plant genotype, developmental stage, the history of the plant cell and duration of stress imposed. Recent studies have partially revealed the mechanisms from different levels: molecular, physiological and biochemical, morphological; but there is currently no unified mechanism to explain the effect of stress on photosynthesis. This study comprehensively reviews the adaptive mechanism of photosynthetic apparatus under salt stress, summarises methods for increasing the resistance and provides a practical way to increase grain yield in saline soils.