【作者】 尤江峰；

【导师】 郑绍建；

【作者基本信息】 浙江大学 ， 农业资源利用， 2006， 博士

【摘要】 铝(Al)毒是酸性土壤上影响作物产量和植物生长的主要限制因子之一。筛选得到高抗Al的植物种类或品种,并进一步阐明植物的抗Al机制,是选育和通过生物技术手段创建适应酸性Al毒土壤上生长的植物的经济有效的手段。Al诱导植物根系分泌有机酸是迄今为止公认的一种最为重要的植物抗Al机制,但Al调控有机酸分泌的过程还没有完全明确。为深入理解Al调控有机酸分泌的过程,本文研究了三种蓼属植物(Polygonum)的抗Al程度与其生长地域土壤性质之间的关系,并从酸性土壤地区筛选得到一中度抗Al的作物一饭豆(Vigna umbellate [Thunb.]Ohwi&Ohashi),继而以这几种植物为材料,从有机酸分泌在植物抗Al中的作用,有机酸分泌时间模式比较,共质体和质外体Al在诱导有机酸分泌中的作用以及金属元素镁(Mg)提高Al诱导有机酸分泌的机制等多方面研究了Al胁迫下蓼属植物和饭豆根系有机酸的分泌特性,结果概述如下: 1.蓼属植物的抗Al性比较及其抗Al机制 源自强酸性土壤上的萹蓄(Polygonum aviculare L)和酸模叶蓼(Polygonum lapathifolium L)具有特殊高抗Al性,而中性土壤上的本氏蓼(Polygonum bungeanum Turcz)对Al极敏感。三种野生蓼属植物的抗Al程度与其生长地域土壤的pH值及活性Al含量密切相关,因而得出土壤环境是寻找发现高抗Al植物的良好指示。Al胁迫下三种蓼属植物根系的草酸分泌量与其抗Al程度正相关,与根尖的Al含量负相关,当草酸的分泌受抑时,加重了根系受Al抑制的程度,所以Al诱导草酸的分泌是蓼属植物的重要抗Al机制。与同属蓼科的荞麦相比,抗Al的萹蓄和酸模叶蓼都不是Al累积植物,尽管它们的抗Al性分别高于及等同于荞麦,而它们的有机酸分泌量远远低于荞麦,可知,除Al诱导有机酸的分泌外,萹蓄和酸模叶蓼还拥有其它有待于进一步探明的抗Al机制。 2.萹蓄和饭豆在Al胁迫条件下有机酸分泌特性的比较更多还原

【Abstract】 Aluminum (Al) toxicity is one of the most significant constaints limiting plant growth and crop production in acid soils, which comprise approximately 30% of arable land worldwide. Al at micromolar concentrations rapidly inhibits root elongation and the uptake of water and nutrients, leading to reduced plant growth and increased susceptibility to environmental stresses. However, some plant species or genotypes, especially native plant species, grow vigorously in acid soils, indicating these species or genotypes have adapted well to high levels of Al in the growth medium. The mechanisms of Al tolerance are not fully understood despite extensive research efforts have contributed to our understanding of Al resistance in plant. For example, the role of organic acids in detoxifying Al both externally and internally in some species has been well documented in the past decade, but it cannot explain high Al resistance in others. To date, most studies of Al-resistance mechanisms have focued on Al-resistant genotypes of crops that have a long breeding history. Therefore, it is very likely that some Al resistance genes have been lost when selection for other agronomic features. In the present study, three native Polygonum species were selected to study Al resistant mechanisms and the relationship between their Al resistance and the soil characteristics of their orgins. On the other hand, although Al-induced organic acid anions secretion has been proven as one of the most important Al resistance mechanisms in many plant species and cultivars, the process of how Al regulated the organic acid anions secretion has not been completely understood. In the present study, Polygonums and rice bean {Vigna umbellate [Thunb.] Ohwi & Ohashi) were selected as the plant materials to study the physiological basis of Al-inuduced oranic acid anions secretion. Because we have proven that Polygonum aviculare L exude oxalate rapidly under Al stress and fitted into Pattern Ⅰ type, but rice bean exude citrate delayed by several hours in response to Al stress and fitted into Pattern Ⅱ type. Thus, the two species must differ greatly in the mechanisms of Al-induced organic acid anions efflux. The results were as follows:1. A comparison of aluminum resistance among Polygonum species originating onstrongly acidic and neutral soilsPolygonum aviculare L and Polygonum lapathifolium L grown in tea garden soils at pH around 3.5 showed high Al resistance, but Polygonum bungeanum Turcz grown in neutral soils was extremely sensitive to Al. Hydrophonic experiments showed that the root elongation was only inhibited by 15% in P. aviculare and 35% in P. lapathifolium after exposure to 50 uM Al for 24 h. Their Al resistance was respectively higher than and similar to that in an Al resistant buckwheat {Fagopyrum esculentum Moench) cultivar. In contrast, the root elongation of P. bungeanum was inhibited by 80% under the same condition. The differences in Al resistance among Polygonum species were confirmed in a 10-d intermittent Al treatment experiment, the root biomass of the former two species were unaltered but decreased by 50% in the latter species. However, high Al accumulation was not found in the leaves, indicating these species were not Al accumulators. Oxalate efflux was detected in root exudates of both Al resistant species. No organic acid anions were detected in the root exudates of the Al sensitive species. The anion channel inhibitor phenylglyoxal (PG) inhibited the oxalate efflux greatly. Inhibition of root elongation was greater in the presence of PG, confirming that oxalate efflux was associated with the Al resistance. However, since the efflux rate was much lower than their related species buckwheat, other mechanisms must be involved in Al resistance and these need to be studied further.2. A comparative study on Al induced oxalate secretion in P. aviculare and citrate secretion in rice beanIn order to clarify the processes of Al induced organic acid anions secretion in different plants with different patterns, the characteristics of Al induced organic acid anions efflux in the roots of rice bean and P. aviculare was investigated. Oxalate secretion in P. aviculare roots occurred within 30 min of Al exposure, while citrate secretion in rice bean roots was induced by at least 4 h Al exposure. Therefore, the former was classified to pattern I, and the latter pattern II. A broad protein-synthesis inhibitor cyclohexide had no obvious effect on the Al-induced oxalate secretion in P.aviculare, but inhibited the Al-induced citrate secretion in rice bean roots by about 84%. This suggested that novel protein synthesis didn’t involved in the Al-induced oxalate secretion in P. aviuculare, but was essential in regulating the Al-induced citrate secretion in rice bean. Al exposure did not affect the oxalate content in P. aviculare root apices, but stimulate citrate synthesis in rice bean. The oxalate secretion in P. aviculare was sensitive to anion channel blocker PG and the citrate secretion in rice bean was sensitive to anthrancene-9-carboxylic acid, indicating anion channels involved in organic acid anions secretion in both plants but their pharmacological properties were different from each other.3. Al in symplast played very important role in regulating oxalate secretion in P. aviculare rootsIn order to determine whether Al in the apoplast or symplast activated the organic acid anion channels in plasma membrane, P. aviculare was grown in nutrient solution with or without NaCl to modify the root cell wall properties. Results showed that neither root plasma membrane intergrity nor oxalate content in root tips was affected by NaCl pretreatment. However, NaCl pretreatment resulted in about 36 % increase in total pectin content of P. aviculare root apices, and enhanced the Al adsorption rate and total Al accumulation in the extracted root cell walls. The oxalate efflux occurred after 3 h of Al exposure in the NaCl pretreatment P. aviculare roots although that occurred within 30 mins Al exposure in the control roots. This was consistent with the delay of Al entry into root symplasm. Therefore, it was suggested that Al must enter into symplast to induce oxalate secretion from P. aviculare roots. An alternative explanation for the present result is that there is a critical Al content in root symplsam to active oxalate efflux. However, the possibility remains that NaCl pretreatment delayed the oxalate secretion by affecting the activity of enzymes or signal substance involved in the opening of anion channels. This needs further investigation in the future.4. The physiological mechanism of Mg alleviating Al toxicityMagnesium (Mg) can alleviate Al toxicity, but the physiological mechanisms arestill not clear. Here we found that micromolar level of Mg obviously alleviated Al-induced but not lanthanum-induced root elongation inhibition in rice bean. The increase of Al-induced citrate efflux paralleled with the decrease of root elongation inhibition with the addition of Mg at either dose (10-50 uM) or time-course (0-4, 4-8, 8-12 and 12-24 h) manner, suggesting that the ameliorative effect by Mg might be closely associated with Mg-enhanced citrate efflux from rice bean roots. Vanadate (an effective H+-ATPase inhibitor) decreased the Al-induced citrate efflux, while addition of Mg partly restored the efflux;Mg addition obviously increased the activity of Al-reduced plasma membrane H+-ATPase, and Mg and calcium contents in root apices significantly. Taken together, we proposed the following scheme: Mg addition compensated the Al-induced Mg decrease, the increased intracellular Mg content up-regulated the activity of plasma membrane H+-ATPase, and finally resulted in the increase of Al-induced citrate efflux.更多还原