基于叶片质外体相关成分和酶活性响应的龙葵耐Cd机理研究

为探讨龙葵(Solanum nigrum L.)耐镉(Cd)机理,采用基质培养法,研究了50 μmol·L^-1 Cd(CdCl₂·2.5H₂O)处理120 d对其生长和叶片细胞壁果胶、木质素含量、多聚半乳糖醛酸酶(PG)、纤维素酶(Cx)、NADH-过氧化物酶(NADH-POD)、愈创木酚过氧化物酶(GPOD)和松柏醇过氧化物酶(CAPX)活性以及质外体汁液中过氧化氢(H₂O₂)含量和GPOD、过氧化氢酶(CAT)、抗坏血酸过氧化物酶(APX)和超氧化物歧化酶(SOD)活性的影响。结果表明,与对照相比,Cd处理显著抑制了龙葵地上和地下部的伸长生长、降低了根的干物质积累(P<0.05),但茎的分枝增多,叶面积、叶绿素含量、净光合速率和胞间CO₂浓度无显著变化(P>0.05);细胞壁结合酶NADH-POD、GPOD和CAPX活性以及果胶含量显著增高(P<0.05),而PG活性显著下降(P<0.05),Cx活性和木质素含量无显著变化(P>0.05);质外体汁液中H₂O₂含量显著增加(P<0.05),GPOD和SOD同工酶谱及活性无显著变化(P>0.05);CAT同工酶谱带增宽,APX同工酶谱带数增加,二者活性均显著升高(P<0.05)。说明50 μmol·L^-1 Cd较长时间处理龙葵植株,其地上部分生长仍处于良好状态,表现出对Cd毒害的耐性,这与Cd处理后叶片细胞壁果胶含量和结合酶GPOD以及质外体汁液中APX、CAT活性升高有关,推测这是龙葵耐Cd的一种机理。     

外质体H2O2和木质素积累在镍诱导的水稻对白叶枯病系统抗性中的作用

[目的]植物外质体是病原入侵的结构和生理性屏障.本文研究外质体反应在镍诱导的、对白叶枯病系统抗性中的作用,以探讨植物对病原和重金属交叉抗性的生理机制. [方法]以0.5、1.0和2.0mol·L~(-1)硝酸镍喷施三叶期水稻幼苗的第二叶及以下部位,3 d后在未处理的第三叶接种稻白叶枯菌(xanthomonas oryzae pv.oryzae,Xoo),接种后12d调查病情.测定了2.0 mol·L~(-1)镍处理或/和Xoo接种的幼苗第三叶外质体中愈创木酚POD(G-POD)、NADH-POD,二胺氧化酶(DAO)活性、H_2O_2和木质素含量.[结果]镍诱导了幼苗对白叶枯病的系统抗性,其中2.0 mmol·L~(-1)镍的诱导效果最佳.镍处理下,第三叶外质体液(AWF)中和细胞壁结合的G-POD、NADH-POD,以及AWF中DAO的活性和H_2O_2水平快速上升,木质素含量显著提高;镍诱导且接种组中,上述指标均明显高于未诱导但接种组.另外,产生H_2O_2的POD的抑制剂NaN_3降低了镍诱导的H_2O_2积累水平和对白叶枯病的系统诱抗效应,而NADPH氧化酶抑制剂(diphenyleneiodonium chloride)预处理则无明显影响.[结论]本研究结果提示,NADH-POD和DAO活性上升是镍诱导外质体H_2O_2产生的原因;外质体H_2O_2和木质素在细胞壁中的积累可能参与了镍诱导的水稻对白叶枯病系统抗性的建立.     

微透析法获取鹅掌柴嫩茎质外体汁液的研究

该文研究的重点是获取质外体汁液。假设微透析技术可以在植物上得到质外体汁液,并以盆栽鹅掌柴为材料,在活体嫩茎插入微透析探针后的不同时间段内分别收集透析液,测定其中的离子浓度和苹果酸脱氢酶活性加以验证。结果表明:在探针插入120 min后,收集的透析液中Na+、K+和Ca2+浓度逐渐趋于平缓,并且苹果酸脱氢酶的活性消失,从而证明120 min后得到的微透析液,没有受到破损细胞液的污染,是纯净的鹅掌柴嫩茎质外体汁液，从而为活体、方便、快捷地获取植物质外体汁液提供新的方法。      

What proteomic analysis of the apoplast tells us about plant–pathogen interactions

Plant pathogens have developed different strategies during their evolution to infect and colonize their hosts. In the same way, plants have evolved different mechanisms acting against potential pathogens trying to infect and colonize their tissues. Regulation of a wide variety of proteins is required in order to perceive the pathogen and to activate the plant defence mechanisms. The apoplast is the first compartment where these recognition phenomena occur in most plant–pathogen interactions, allowing the exchange of different molecules and facilitating inter‐ and intracellular communication in plant cells. Proteomic analysis of the apoplast in recent years has found the initial biochemical responses involved in pathogen recognition and early defence responses. However, this proteomic approach requires some specific experimental conditions to obtain an extract free of cytoplasmic proteins and nonprotein contaminants that affect the subsequent stages of separation and quantification. Obtaining the highest proportion of proteins from the apoplastic space in infected tissues requires different steps such as extraction of apoplastic washing fluids and preparation of total secreted proteins (protein precipitation, solubilization, separation and digestion). Protein identification using mass spectrometry techniques and bioinformatics tools identifying peptides for the extracellular exportation is required to confirm the apoplastic location. This review compiles the most commonly used techniques for proteomic studies, focusing on the early biochemical changes occurring in the apoplast of plants infected by a wide range of pathogens. The scope of this approach to discover the molecular mechanisms involved in the plant–pathogen interaction is discussed.     

Determination of the different components of cadmium short‐term uptake by roots

As the various components of the cadmium (Cd) root sink have not been clearly described, there is a need to precisely measure the respective contributions of apoplast and symplast to short‐term root Cd uptake and to explain the linear component of the absorption isotherms. A new method of fractionating Cd in roots was applied to two plant species with contrasting abilities to accumulate Cd: maize (Zea mays) and a Cd‐hyperaccumulating ecotype of alpine pennycress (Noccaea caerulescens). Their roots were exposed for 1 h to increasing concentrations of labeled Cd. Series of desorption baths were used to obtain the root apoplastic Cd in combination with a brief freezing step in liquid nitrogen to separate the intracellular metal from the apoplastic one. The apoplastic uptake accounted for 15% to 82% and for 48% to 96% of the total Cd uptake of maize and of alpine pennycress roots, respectively. In the case of maize, the concentration‐dependent symplastic net flux fitted a biphasic Michaelis‐Menten function, while in the case of alpine pennycress, a Michaelis‐Menten‐plus‐linear function proved a better fit. The second component of the symplastic net flux may reflect absorption through a low‐affinity transport system. Short‐term Cd uptake by roots is dominated by the high‐affinity transport system for exposure concentrations below 1 μM for maize and 0.2 μM for alpine pennycress, while cell‐wall binding prevailed for higher exposure concentrations.     

Interactive Effects of High Boron and NaCl Stresses on Subcellular Localization of Chloride and Boron in Wheat Leaves

Salinity and boron (B) toxicity often occur simultaneously and may have interactive effects on plant responses. This study aims at further investigating long‐term effects of salinity, B toxicity and their combination on wheat plants. Plants grown in hydroponics were treated with 2.5 μm H₃BO₃ (control), 75 mm NaCl, 200 μm H₃BO₃ or 75 mm NaCl + 200 μm H₃BO₃ and were analysed on a subcellular level 6 weeks after germination. Shoot fresh and dry weight (DW), water content, transpiration rate and osmolality were reduced, while Na⁺ and Cl^? concentrations increased under salinity stress. However, Cl^? was reduced in all compartments by an additional application of B toxicity. At adequate B supply, NaCl increased apoplastic and symplastic soluble B concentrations, whereas the total B content remained unchanged. At high B level, however, soluble and total B was reduced by additional salt stress. Despite this antagonistic effect of a combined salt and B toxicity stress on Cl^? and B concentrations, an additive effect was observed regarding shoot fresh weight reduction. Our results confirm an alleviating effect of the combined stresses on toxic ion concentrations, which did not prevent additive growth reductions.     

冬小麦冠层-大气氨交换的季节性特征及其影响因素

为探究冠层氨交换对麦田氨挥发的贡献,从农学和生理角度揭示冠层氨挥发的机理,本文以冬小麦‘小偃22’和‘郑麦366’为材料,研究了不同生育期冬小麦冠层NH_3交换动态;枯、黄叶和根茬对冠层NH_3挥发的影响;监测了不同生育期的叶片硝酸盐含量、硝酸还原酶活性、谷氨酰胺合成酶活性、质外体铵浓度及可溶性蛋白含量等氮代谢指标,考察了其与冠层氨交换的关系。结果表明:冬小麦孕穗～扬花之前冠层吸收氨,为氨汇,其对农田氨挥发的消减率可达23%～40%;孕穗～扬花之后以冠层释放氨,为氨源,其对农田氨挥发的贡献率达17%～27%。小偃22冠层氨的吸收与挥发分别较郑麦366高28%～47%和19%～68%。氮肥施用下的土壤和冠层氨交换比不施氮处理高2.82～3.19kg·hm~(-2)和0.972～1.463 kg·hm~(-2),高肥力土壤和冠层氨交换比低肥力土壤高15.18～15.55 kg·hm~(-2)和0.947～1.438 kg·hm~(-2)。枯、黄叶对冠层氨挥发的作用受品种、生育期及土壤肥力等的影响。在高肥力和施氮(N180)条件下,收获后的麦茬也是氨释放源。冠层氨挥发与土壤温度、质外体铵浓度和可溶性蛋白呈显著线性正相关,与谷氨酰胺合成酶活性和硝酸盐含量呈显著线性负相关。结论:冬小麦营养生长期冠层与大气的氨交换以吸收为主,进入灌浆期后,以释放为主,氨交换方向受到生育期及氮素代谢过程的影响,同时也受到小麦品种和土壤肥力的影响。     

Aluminum inhibits apoplastic flow of high–molecular weight solutes in root apices of Zea mays L.

Using an aluminum (Al)‐sensitive maize cultivar, we investigated the influence of Al on the apoplastic solute bypass flow and its relationship with Al‐induced (1 h, 50 μM) callose formation and root growth. We selected the fluorescent probes 8‐hydroxypyrene‐1,3,6‐trisulfonic acid, trisodium salt (MW 524) (HPTS) and dextran‐Texas Red (TR) conjugates (MW 3,000, 10,000, and 40,000) to monitor their apoplastic transport. Confocal laser–scanning microscopy (CLSM) analysis and spectrofluorometric quantification showed Al‐induced callose formation in peripheral root cells within 1 h. Pretreatment of plants with the callose synthesis inhibitor 2‐deoxy‐D‐glucose (DDG) reduced the callose formation by half. Uptake experiments with both HPTS and dextrans showed uniform dye distribution in control root apices. After Al treatment for 1 or 2 h, which inhibited root growth by 32% or 50%, respectively, the dyes accumulated in the epidermal and outer cortical cell layers, especially in the 1–2 mm apical root zone. Al treatment reduced the export of the dyes out of the apical 1 cm treatment zone. This was due to strong sorption of HPTS but not of dextrans by Al‐loaded cell walls. Aluminum treatment reduced loading into the xylem sap particularly of higher–molecular weight dextrans. Pretreatment of roots with DDG and presence of 50 mM mannitol during the Al treatment partially forestalled the inhibitory effect of Al on the dye transport, but only slightly reduced the Al‐induced growth inhibition. Exudation experiments revealed that xylem water flow remained unaffected by the Al treatment of the root tips. The results with dextran suggest that Al binding in cell walls of the root apex inhibits apoplastic bypass flow of higher–molecular weight solutes, which might contribute to Al‐induced inhibition of root growth.     

Ammonia exchange between rice leaf blades and the atmosphere: Effect of broadcast urea and changes in xylem sap and leaf apoplastic ammonium concentrations

Abstract

To elucidate the effects of broadcast urea on ammonia (NH₃) exchange between the atmosphere and rice, we investigated the NH₃ exchange flux between rice leaf blades and the atmosphere, xylem sap ammonium (NH⁺ ₄) concentration, leaf apoplastic NH⁺ ₄ concentration and pH, and determined the stomatal NH₃ compensation point. Paddy rice (Oryza sativa L. cv. Nipponbare) cultivation using experimental pots was conducted in the open air. Three treatments, no nitrogen (NN), standard nitrogen (SN) and high nitrogen (HN), were prepared for two supplemental fertilizations. Urea with 0, 30 and 60 kg N ha^?1 for the NN, SN and HN treatments, respectively, was broadcast at panicle initiation, and urea with 0, 20 and 40 kg N ha^?1 for the NN, SN and HN treatments, respectively, was broadcast at heading. The NH₃ exchange fluxes between the rice leaf blades and the atmosphere (SN treatment) measured using a dynamic chamber technique showed net deposition in general; however, net emission from the old leaves occurred 1 day after the application at heading. In contrast, the xylem sap NH⁺ ₄ concentrations increased markedly 1 day after both applications, which suggests direct transportation of NH⁺ ₄ from the rice roots to the above-ground parts. The applications resulted in no obvious increase in the leaf apoplastic NH⁺ ₄ concentrations. The relationship between the NH⁺ ₄ concentration in the xylem sap and that in the leaf apoplast was uncertain, although the NH⁺ ₄ in the xylem sap came from the roots and the NH⁺ ₄ in the apoplast might be affected by the stomatal deposition of NH₃. The stomatal NH₃ compensation point of rice was estimated to be 0.1–4.1 nmol mol^?1 air (20°C). The direction and intensity of the exchange flux through the stomata, interpreted on the basis of the temperature-corrected NH₃ compensation point, agreed with the observed exchange flux between the rice leaf blades and the atmosphere.     

盐胁迫下杨树质外体离子对气体交换参数的影响

利用微透析技术,研究盐胁迫下吴屯杨和小胡杨嫩茎质外体中离子的动态变化对气体交换参数的影响.杨树嫩茎质外体离子采用微透析与原子吸收联用进行监测,同时采用Li-6400便携式光合仪进行气体交换相关参数的测定.盐胁迫下,2个杨树品种嫩茎质外体中Na+积累的程度不同,吴屯杨经过一段时间后能够自我调节,减少№+含量,而小胡杨质外体一直积累Na+,从而造成细胞的渗透胁迫,抑制植物生长;随着盐处理对间的增加,2种杨树K+浓度基本呈现下降的趋势.吴屯杨在100 mmol·L-1NaCl胁迫早期光合受抑制的主要原因是非气孔限制,后期是气孔限制;而小胡杨光合受抑制的主要原因是气孔限制.对Na+浓度、K+浓度、Ca2浓度、Na+/K+、Pn进行相关性分析发现,2个杨树品种Pn均与Na+浓度和Na+/K+比值呈显著的负相关,与K+浓度呈极显著的正相关.以Pn为因变量,以Na+,K+,Ca2+浓度和Na+/K+为自变量,进行多元线性逐步回归分析得统计方程,其中只有Na+/K+对Pn的相关系数有显著意义,方程中只有Na+/K+进入方程,其他3个离子都被剔除.盐胁迫下,吴屯杨耐盐性强于小胡杨,统计方程说明盐胁迫下杨树嫩茎质外体离子变化和光合作用存在共线性关系,且Na+/K+对Pn有显著的线性关系.