黑麦品种间耐铝差异性机制研究

采用室内模拟方法,研究了铝诱导黑麦根系分泌有机酸、根尖磷对铝的固定作用。结果表明,在铝胁迫下冬牧品种（Win）相对根伸长率高于King品种。在铝胁迫下,经磷预处理的根尖铝和磷含量增加,且以Win品种根尖中的Al、P含量较高,说明根尖磷对铝的固定是黑麦耐铝机制之一。另一方面,在铝胁迫下,两品种根系均分泌柠檬酸和苹果酸,且Win的分泌速率较高。有机酸的分泌随着铝处理浓度（10、30、50 mol/L）和时间（0.5、3、6、9、12 h）的增加而增加,但在低温（4C）下柠檬酸分泌量显著减少。Al处理0.5 h后苹果酸已明显分泌,而柠檬酸的分泌在铝处理6 h后才明显增加。在铝处理前,进行缺磷预处理（3 d）不能增加有机酸的分泌,10 mol/L的La3+、Cu2+、Ni3+也不能诱导根系分泌有机酸。说明铝诱导根系专一性分泌有机酸也是黑麦品种间耐铝性差异的机制。     

铝诱导黑麦的根尖分泌有机酸

虽然较多的研究已发现黑麦对铝毒害的忍耐能力较强，但是有关黑麦对铝毒害的抵御机理的报告很少。为了阐明黑麦（品种：King)对铝毒害的抵御机理，本研究探讨了黑麦的根系有机酸分泌的特性及其作用，铝能够诱导黑麦的根系分泌柠檬酸和苹果酸。柠檬酸主要从距顶端0－5mm的根尖分泌。有机酸的分泌量随着处理的铝浓度(0,50,100,300μmol/L AlCl3）和处理的时间（0．5，1，3，5h）的增加而增加。0，50，100，300μmol/L的AlCl3处理根尖3h后，从根尖分泌的柠檬酸和残留在根尖中的柠檬酸之和随着处理的铝浓度的增加而增加，而各处理间的苹果酸总量无显著差异。50μmol/L的柠檬酸或者400μmol/L的苹果酸能够消除50μmol/L 的AlCl3铝对小麦（Scout 66)根尖表面细胞的伤害。这些结果表明，铝促进柠檬酸的合成和柠檬酸、苹果酸从根尖的分泌是黑麦抵抗铝毒害的机理。     

在铝胁迫下黑麦根系分泌的柠檬酸和苹果酸的解毒机制的研究

在我国普遍种植的黑麦(冬牧70),在铝胁迫下其根系能分泌柠檬酸-苹果酸复合有机酸。为了揭示在铝胁迫下黑麦根系分泌的复合有机酸(柠檬酸-苹果酸)的解毒机理,研究了上述复合有机酸对铝胁迫下小麦幼根细胞膜透性、过氧化物酶(POD)、过氧化氢酶(CAT)、H -ATPase活性和根伸长的影响。结果显示,复合有机酸使小麦幼根电解质渗漏率下降,根尖过氧化物酶、过氧化氢酶、H -ATPase活性提高,幼根伸长量增加,并且复合有机酸的效果随其浓度(25~200μm ol/L)的增加而增强。在50或100μm ol/L苹果酸的基础上,柠檬酸与苹果酸比值(C¨M)从1¨2增加到1¨1,复合有机酸的解毒效果更显著;而在相同柠檬酸浓度下,苹果酸浓度增加一倍对小麦铝毒害无显著影响。可以推断,在铝胁迫环境下,黑麦根系分泌的柠檬酸-苹果酸复合有机酸可以维持细胞膜结构的完整性,减轻自由基伤害和铝对根生长的毒害,并且复合有机酸中柠檬酸的作用强于苹果酸。铝胁迫下根系分泌复合有机酸是黑麦抵御铝毒害的有效机制。     

铝胁迫对小麦根尖细胞蛋白质及苹果酸分泌的影响

以一对近等位基因小麦品系(ET8和ES8)为材料,研究了铝胁迫条件下根系生长情况,根尖铝含量,根尖细胞质蛋白组分以及铝胁迫下两个小麦品系苹果酸分泌的差异。结果表明,小麦品系ET8的耐铝性明显高于ES8;而ET8根尖铝含量则明显低于ES8;50mol/L铝处理24h后ES8小麦的细胞质蛋白组成受到明显影响,而ET8则无明显影响。由于铝胁迫条件下ET8分泌的苹果酸的量明显高于ES8,因此可以认为,分泌的苹果酸一方面与铝结合使其无毒化;另一方面将铝排斥于根尖细胞外,从而阻止铝进入细胞质内对细胞产生毒害作用。     

铝胁迫下黑麦根尖分泌有机酸和钾离子的研究

采用药理学研究方法,研究了几种抑制剂和脱落酸对铝诱导黑麦根尖分泌有机酸和钾离子的影响。结果表明,在铝(100、200、300μmol/L AlCl3)胁迫下,根尖分泌柠檬酸、苹果酸和钾离子,且随着铝处理浓度的增加其分泌量显著增加。阴离子通道抑制剂苯甲酰甲醛(0.1、0.2 mmol/L)抑制根尖在铝(200μmol/L AlCl3)胁迫下分泌有机酸的同时,也抑制根尖分泌钾离子;钾离子通道抑制剂四乙基铵(20、40 mmol/L)和铯(10、20 mmol/L CsCl)在抑制根尖分泌钾离子的同时,也抑制铝诱导的有机酸分泌。25、50μmol/L的ABA处理后,铝诱导的有机酸分泌和钾离子的分泌均显著增加。但是,铝诱导的有机酸分泌在受到阴离子通道抑制剂尼氟灭酸(0.4、0.8 mmol/L)处理抑制后,钾离子的分泌并不减少;铝胁迫下根尖分泌的钾离子在ATP酶抑制剂钒酸钠(0.25、0.50、2.00 mmol/L)处理后受阻的同时,有机酸的分泌却显著增加。这些结果说明,钾离子是铝诱导黑麦根尖分泌有机酸的陪伴离子,而并非有机酸分泌的调控因子。     

几种禾本科作物对铝的敏感性或耐性

通过调查几种禾本科作物根的伸长，根尖的铝含量、铝诱导其根系有机酸的分泌，探讨作物耐铝（铝敏感）性及作物对铝毒害的抵御机理。结果表明，水稻、黑麦是耐铝的作物，而大麦、小麦（Scout 66)是对铝敏感的作物，玉米和高梁对铝敏感性较小麦弱。小麦和玉米品种间对铝的敏感性差异显著，而高梁对铝的敏感性品种间差异性较小。铝能够诱导水稻、黑麦和耐铝的玉米及小麦品种的根系分泌柠檬酸和/或苹果酸。这表明铝诱导有机酸的分泌是它们抵御铝毒害的机理。     

铝胁迫下大豆根系脱落酸累积与柠檬酸分泌的关系研究

为探讨铝(Al)胁迫条件下脱落酸(ABA)调控植物根系有机酸分泌的机制,进行了ABA与Al诱导大豆根系柠檬酸分泌的关系试验。结果表明:1)外源ABA和ABA合成抑制剂fluridone分别提高和降低了Al诱导的大豆根尖ABA含量的增加,但对根系柠檬酸分泌量均无影响,ABA对根系内源柠檬酸含量和柠檬酸合成酶的活性也没有影响;2)分根试验表明,与Al直接接触的根部(Part A)内源ABA含量发生变化,且有柠檬酸的分泌,而不与Al直接接触的根部(Part B)内源ABA含量也发生变化,但没有柠檬酸分泌;3)Al胁迫下,大豆耐Al基因型柠檬酸分泌量远高于敏感基因型,但二者的内源ABA含量却没有差异;4)30μmol AlCl3处理,在0~12 h柠檬酸分泌速率和内源ABA含量随Al处理时间增加而增加,去除Al胁迫时(12~18 h),柠檬酸分泌速率继续增加,但内源ABA含量则迅速下降。综合以上结果,推测ABA不是通过提高Al诱导柠檬酸分泌来调控大豆耐Al性。     

低分子量有机酸对石灰性土壤磷吸附动力学的影响

根据植物在缺磷胁迫下,根系分泌的有机酸种类的数量,用流动法研究了柠檬酸、苹果酸、草酸和酒石酸对石灰性土壤磷吸附动力学的影响。结果表明：有机酸均能明显降低土壤对磷的吸附,不同有机酸降低磷吸附的能力大小的次序为草酸≥柠檬酸〉苹果酸≥酒石酸。     

铝胁迫下萹蓄与饭豆根系有机酸分泌特性比较

以蓄和饭豆为试验材料,从对Al胁迫的响应时间、蛋白质合成抑制剂的影响、体内有机酸含量变化及阴离子通道抑制剂的影响等方面,比较研究了Al诱导根系分泌有机酸的差异,以进一步明确铝(Al)诱导植物根系有机酸分泌的过程。结果表明,萹蓄根系在Al胁迫后30.min内分泌出草酸,而Al胁迫至少4.h后饭豆根系才开始分泌柠檬酸;蛋白质合成抑制剂环己亚酰胺(Cycloheximide,CHM)不影响萹蓄根系草酸的分泌,但抑制了84%Al诱导的饭豆根系柠檬酸的分泌,表明前者不需要新蛋白质的诱导合成,却是后者所必需的。Al处理不改变萹蓄根尖草酸的含量,但明显提高了饭豆根尖柠檬酸的含量;阴离子通道抑制剂苯甲酰甲醛(Phenylglyoxal,PG)和蒽-9-羧酸(Anthrancene-9-carboxylic.acid,A-9-C)分别有效抑制Al诱导萹蓄根系草酸的分泌及饭豆根系柠檬酸的分泌,再次证明两者有机酸的分泌是通过某种被Al所诱导或激活的阴离子通道所实现的。     

铝胁迫下水稻幼苗根尖表面pH和有机酸的动态变化

采用添加pH指示剂溴甲酚紫的琼脂糖凝胶平板技术, 检测了铝胁迫下水稻幼苗根尖表面pH和根尖细胞有机酸含量的动态变化。铝胁迫下, 幼苗根尖表面颜色在处理第1 h、3 h、6 h无明显变化; 而对照处理第1 h时水稻幼苗根尖表面颜色已发生明显变化, 呈浅橙色, 处理第3~6 h幼苗根尖表面变为橙红色。表明在正常条件(pH 4.4)下水稻幼苗根尖表面pH有碱化趋势, 铝胁迫条件下(pH 4.4)水稻幼苗根尖表面pH碱化的趋势被抑制。在测定的铝胁迫水稻幼苗根尖细胞细胞质10种有机酸中, 发现3种有机酸含量的变化趋势不同。草酸含量无明显变化; 柠檬酸含量在铝处理0~3 h内变化不明显, 处理3 h后迅速提高, 处理12 h达最大值后转而下降, 铝胁迫24 h时柠檬酸含量比胁迫12 h时降低16.27%, 差异达显著水平, 胁迫24 h后的柠檬酸含量变化不大但仍高于胁迫6 h的柠檬酸含量; 铝处理3 h后苹果酸含量较胁迫前下降幅度不大, 差异不显著(P>0.05), 3 h后显著下降, 6 h后下降趋势不明显。     

小麦的铝毒及耐性

为探明Al的毒性和忍耐机理 ,比较了Scout 66和Atlas 66Al敏感和抗性的 2个小麦品种的根对Al的积累模式、根细胞壁对Al的吸附以及Al诱导的有机酸的分泌。结果表明 ,Al对Scout 6 6根伸长的抑制作用较Atlas 66明显。根系吸收的Al主要积累于 0至 5mm根尖处。Scout 6 6的根尖及Al处理后分离的根尖细胞壁对Al的积累量大于Atlas 6 6。但是 ,Al处理前分离根尖细胞壁 ,Al处理后细胞壁对Al的吸附量两品种间无显著差异。Al可诱导Atlas 6 6的根系分泌苹果酸 ,而Scout 6 6的分泌物中未发现Al诱导的有机酸。这些结果表明 ,Atlas 6 6的根尖及其细胞壁较Scout 66积累较少的Al,这种差异与Al诱导的有机酸分泌有关 ,而与根尖细胞壁固有的吸附Al的能力无关     

Aluminum tolerance of two wheat cultivars (Brevor and Atlas66) in relation to their rhizosphere pH and organic acids exuded from roots

Phytotoxicity of aluminum (Al) has become a serious problem in inhibiting plant growth on acid soils. Under Al stress, the changes of rhizosphere pH, root elongation, absorption of Al by wheat roots, organic acids exuded from roots, and some main factors related to Al-tolerant mechanisms have been studied using hydroponics, fluorescence spectrophotometry, and high performance liquid chromatography (HPLC). Two wheat cultivars, Brevor and Atlas66, differing in Al tolerance are chosen in the study. Accordingly, the rhizosphere pH has a positive effect on Al tolerance. Atlas66 (Al-tolerant) has higher capability to maintain high rhizosphere pH than Brevor (Al-sensitive) does. High pH can reduce Al3+ activity and toxicity, and increase the efficiency of exuding organic acids from the roots. More inhibition of root elongation has been found in Brevor because of the exposure of roots to Al3+ solution at low pH. Brevor accumulate more Al in roots than Atlas66 even at higher pH. Al-induced exudation of malic and citric acids has been found in Atlas66 roots, while no Al-induced organic acids have been found in Brevor. These results indicate that the Al-induced secretion of organic acids from Atlas66 roots has a positive correlation with Al tolerance. Comprehensive treatment of Al3+ and H+ indicates that wheat is adversely influenced by excess Al3+, rather than low pH.     

铝和镉胁迫对两个大麦品种矿质营养和根系分泌物的影响

A hydroponic experiment was carried out to study the effect of aluminum （Al） and cadmium （Cd） on Al and mineral nutrient contents in plants and Al-induced organic acid exudation in two barley varieties with different Al tolerance. Al- sensitive cv. Shang 70-119 had significantly higher Al content and accumulation in plants than Al-tolerant cv. Gebeina, especially in roots, when subjected to low pH （4.0） and Al treatments （100 μmol L^-1 Al and 100 μmol L^-1 Al ＋1.0 μmol L^-1 Cd）. Cd addition increased Al content in plants exposed to Al stress. Both low pH and Al treatments caused marked reduction in Ca and Mg contents in all plant parts, P and K contents in the shoots and leaves, Fe, Zn and Mo contents in the leaves, Zn and B contents in the shoots, and Mn contents both in the roots and leaves. Moreover, changes in nutrient concentrations were greater in the plants exposed to both Al and Cd than in those exposed only to Al treatment. A dramatic enhancement of malate, citrate, and succinate was found in the plants exposed to 100 μmol L^-1 Al relative to the control, and the Al-tolerant cultivar had a considerable higher exudation of these organic acids than the Al-sensitive one, indicating that Al-induced enhancement of these organic acids is very likely to be associated with Al tolerance.     

Citrate Secretion Induced by Aluminum Stress May Not be a Key Mechanism Responsible for Differential Aluminum Tolerance of Some Soybean Genotypes

Abstract

Eighteen soybean genotypes differing in aluminum (Al) tolerance were used to investigate genotypic differences in Al-induced citrate exudation and its role in Al tolerance. Aluminum accumulation and localization in soybean roots were examined by analysis of total Al and hematoxylin staining. Soybean genotypes exhibited a wide range of Al tolerance. Based on relative root elongation, several Al-tolerant genotypes from Brazil such as B1, B10, and B15 were more tolerant than the Al-tolerant PI 416937 (PI) and Perry. All soybeans exuded citrate in response to Al stress, and some Al-sensitive genotypes secreted more citrate than tolerant ones, showing no correlation between the Al tolerance and Al-induced citrate exudation. Further study found that both copper (Cu) and cadmium (Cd) stimulated citrate and malate exudation in soybean, indicating that organic acid secretion is not specifically induced by Al. Aluminum concentrations were significantly higher in 2–3 and 3–4 cm of segments than that in 0–1 and 1–2 cm segments under 15 μM AlCl₃. Both the root mature zone and apex were heavily stained by hematoxylin after exposure to 10, 15, or 20 μM AlCl₃ (24 h), whereas root elongation zone was not stained. After exposure to 50 μM AlCl₃ for 20 min, the Al-tolerant PI was less stained by hematoxylin than the Al-sensitive Young, suggesting that Al accumulation in root apices seem to be an immediate response to Al stress, and related to differential Al sensitivity. Present results suggest that citrate secretion induced by Al stress may not be a key mechanism responsible for the differential Al tolerance of some soybean genotypes and other mechanism(s) conferring Al exclusion should exist and operate immediately after exposure to Al stress.     

Combined Effect of Short-Term Water Deficit Stress and Aluminum Toxicity on Citrate Secretion from Soybean Roots

Abstract

In order to investigate the combined effect of drought stress and aluminum (Al) toxicity on citrate exudation in soybean, hydroponic cultivation with addition of Al and PEG-6000 was conducted to simulate Al-toxic dry soil. Results showed that 2-day exposure of soybeans to 5.5% (w/v) PEG-6000 or 100 µM AlCl₃ mainly hindered root growth, while combined exposure to PEG and Al (PEG/Al) reduced both root and shoot growth. Photosynthetic rate of first trifoliolate for the Al-tolerant genotype PI 416937 (PI) was not affected by imposition of 5.5% PEG/100 µM AlCl₃ (38–40 h), whereas photosynthetic rate for the Al-sensitive genotype YC was significantly reduced. Based on root fresh weight, Al-induced citrate exudation in the roots of soybean genotypes recovered from pre-treatment with 5.5 or 9% PEG was not altered, but was increased in the plants pretreated with 7% PEG without recovery. After 2 days of recovery from 2-day combined exposure to 5.5% PEG/100 µM AlCl₃, the Al-tolerant PI exuded more citrate than its control, but the Al-sensitive YC exuded significantly less citrate than its corresponding control. Split root experiment revealed that Al-induced citrate exudation in one half of the root system was significantly reduced by exposing the other half of the root system to 0.5 mM CaCl₂ solution containing 9% PEG or 9% PEG/50 µM AlCl₃. However, organic acid secretion was not observed in the half of the root system in the Ca solution when the other part of the root system was exposed to the Ca solution containing 50 µM AlCl₃, 9% PEG or 9% PEG/50 µM AlCl₃. This suggests that no Al- or drought-induced signals such as ABA are involved in the citrate secretion in soybean.     

Aluminum tolerance associated with enhancement of plasma membrane H+-ATPase in the root apex of soybean

Abstract

Seventeen soybean cultivars were screened to discern differences in aluminum (Al) sensitivity. The Sowon (Al-tolerant) and Poongsan (Al-sensitive) cultivars were selected for further study by simple growth measurement. Aluminum-induced root growth inhibition was significantly higher in the Poongsan cultivar than in the Sowon cultivar, although the differences depended on the Al concentration (0, 25, 50, 75 or 100?μmol?L^–1) and the amount of exposure (0, 3, 6, 12 or 24?h). Damage occurred preferentially in the root apex. High-sensitivity growth measurements using India ink implicated the central elongation zone located 2–3?mm from the root apex. The Al content was lower 0–5?mm from the root apices in the Sowon cultivar than in the apices of the Poongsan cultivar when exposed to 50?μmol?L^–1 Al for 12?h. Furthermore, the citric acid exudation rate was more than twofold higher in the Sowon cultivar. Protein production of plasma membrane (PM) H⁺-ATPase from the root apices (0–5?mm) was upregulated in the presence of Al for 24?h in both cultivars. This activity, however, decreased in both cultivars treated with Al and the Poongsan cultivar was more severely affected. We propose that Al-induced growth inhibition is correlated with changes in PM H⁺-ATPase activity, which is linked to the exudation of citric acid in the root apex.