西藏野生大麦铝毒诱导根系分泌磷酸的特异耐铝机制研究

Aluminum (Al) toxicity is a major constraint for plant growth and production on acid soils. It is imperative to improve the Al tolerance of plant for the promotion of the land resource utilization. Tibetan wild barley (Hordeum spontaneum), an elite and unique germplasm in China, gradually develops abiotic stress tolerance due to its adaption to harsh environments. In our previous studies, we identified Al-tolerant genotypes in Tibetan wild barley. However, their Al tolerance could not be contributed by Al-induced citrate secretion, the main Al-tolerant mechanism in cultivated barley. Instead, we found that Al-tolerant wild genotype secreted more phosphate than Al-sensitive one. Phosphate can bind to Al to form non-toxic complex, thereby reducing the active Al3+ in rhizosphere, which could be a novel mechanism of Al tolerance in Tibetan wild barley. This study aims to verify the specific Al-tolerant mechanism at cellular and molecular levels by following experiments: (1) using microelectrode ion flux estimation (MIFE) technique to investigate the ion flux under Al stress; (2) using confocal microscopy and secondary ion mass spectrometry (SIMS) to study the subcellular distribution of Al and P; (3) using RNA-Seq and in situ PCR techniques to study the molecular mechanism on the transport of Al and P under Al stress. The objectives of this work are to reveal the physiological and molecular mechanisms of Al tolerance in Tibetan wild barley, which will be helpful for plant breeding on developing barley cultivars with Al tolerance.

铝毒是酸性土壤上限制作物生长的主要毒害因子，提高作物耐酸铝能力对促进土壤资源利用具有重大意义。西藏野生大麦是我国独有的珍稀种质，由于长期生长在恶劣的生态环境下，逐渐形成相适应的抗逆机制。在前期工作中，我们鉴定到野生大麦耐铝种质，而现有的栽培大麦耐铝理论不能解释其耐铝机制；发现野生耐铝种质受铝毒胁迫诱导分泌更多的磷酸，磷酸可以螯合根际铝离子，从而缓解铝毒，可能是西藏野生大麦重要耐铝机制。本项目将从细胞和分子水平验证野生大麦特异耐铝机制，研究内容有：（1）采用微电极离子流测定技术，测定铝毒胁迫下根系不同部位的Al3+和HPO42-的离子流；（2）采用激光共聚焦显微技术和二次离子质谱技术，测定根系横切面的铝和磷元素的分布图像；（3）采用转录组测序技术和原位PCR技术，分析铝和磷元素转运和代谢相关基因及其代谢通路。本项目可望阐明野生大麦特异耐铝生理分子机制，为大麦耐铝育种实践提供理论指导。

铝毒是酸性土壤上影响作物产量的主要胁迫因子，一些物种已经进化出通过从根组织中排铝来降低铝毒害的机制。已有报道表明，栽培大麦（Hordeum vulgare L.）的耐铝性（包括Dayton）主要依赖于柠檬酸转运蛋白（HvAACT1）介导的根系柠檬酸分泌螯合铝离子的外部脱毒机制。然而，我们发现野生大麦（Hordeum spontaneum L.）XZ29具有较强的耐铝性，但它不表现出典型的铝毒诱导的柠檬酸外流或HvAACT1高表达。XZ29源于西藏东南部，那里的土壤呈酸性，磷含量高。我们利用MIFE和SIMS技术，检测到根尖的磷酸盐外排，以及根组织中铝元素和磷元素的共定位。此外，铝毒胁迫下，XZ29根尖组织可维持高含量的总磷、无机磷和含磷的糖酵解中间产物，表现出较高的磷利用率和磷的再活化能力。综上，XZ29的耐铝性依赖于铝毒诱导根尖分泌磷酸盐，螯合根际铝离子，在表皮细胞层形成Al-P沉淀，减少了铝离子在皮层组织中的积累。西藏野生大麦为适应西藏东部和东南部高磷酸性土壤的恶劣条件，进化出独特的磷元素转运代谢的耐铝机制。