SgALMT1介导根系分泌苹果酸参与柱花草耐低磷抗铝毒的分子机理

Stylo (Stylosanthes) is a dominant leguminous forage, and has superior capability tolerant to low phosphorus (P) and aluminum (Al) toxicity, which are considered as major constraints for crop growth in acid soils. Malate exudation from roots functions in increasing P availability and Al tolerance, which is mainly through the gene function of Al-activated malate transporter, ALMT. However, the molecular mechanisms of stylo adaptation to low P and Al toxicity through malate exudation, mediating by SgALMTs, are still unknown. In the present study, we plan to confirm malate exudation involved in stylo tolerant to low P and Al toxicity through identification of the correlation between SgALMT single nucleotide polymorphism (SNPs) and phenotypic traits of stylo under low P and Al toxicity, as well as analyzing the correlation between SgALMT1 expression and malate exudation, in population of stylo. Furthermore, we plan to elucidate the biological basis of malate exudation mediated by SgALMT1 through identification of the key amino acid, which probably associated with protein phosphorylation modification, and investigating the regulation characteristics of P and Al on SgALMT1 expression via analysis of P- and Al-regulated Cis-element in the promoter region of SgALMT1. Finally, using the transgenic stylo plants overexpressing SgALMT1 to study the biological function of SgALMT1 involved in increasing the availability of P and Al toxicity tolerance through mediating malate exudation. Our results might be important to further dissect the molecular mechanisms of leguminous forage adaptation to low P and Al toxicity in acid soils.

柱花草是重要的热带豆科牧草，对酸性土壤中限制作物生长的低磷和铝毒胁迫具有较强的忍耐能力和独特的适应性机制。苹果酸转运蛋白基因ALMT已被报道具有介导根系分泌苹果酸的功能。SgALMTs基因介导根系分泌苹果酸参与柱花草耐低磷和抗铝毒的分子机理仍不清楚。本项目首先在柱花草群体材料中，进行SgALMT1单核苷酸多态性与柱花草耐低磷和抗铝毒表型性状的相关性分析，研究SgALMT1基因表达与苹果酸分泌的相关性，明确根系分泌苹果酸参与柱花草耐低磷和抗铝毒的过程；通过鉴定SgALMT1受蛋白磷酸化修饰的关键氨基酸位点和SgALMT1启动子磷铝调控顺式作用元件，揭示磷铝调控SgALMT1介导根系分泌苹果酸的生物学机制；创制超量表达SgALMT1转基因柱花草，探索SgALMT1介导根系分泌苹果酸参与柱花草活化利用土壤磷和抗铝毒的生物学功能。研究结果对深入挖掘豆科牧草耐低磷和抗铝毒机制具有重要的科学意义。

低磷和铝毒酸性土壤中限制作物生长和产量的重要因素。柱花草是重要的热带豆科牧草，对低磷和铝毒胁迫具有较强的适应性，但根系分泌苹果酸缓解柱花草低磷和铝毒胁迫的机制仍不清楚。本研究首先以不同柱花草基因型为材料，评价分析了柱花草耐低磷和抗铝毒能力，结果发现不同柱花草耐低磷和抗铝毒能力具有基因型差异，并从中筛选出了不同抗性的柱花草基因型，这为后续抗逆高产柱花草新品种选育提供了重要的材料支撑。本研究结果发现了SgALMT1基因及其启动子的在不同基因型间存在单核苷酸多态性。其多态性位点及蛋白磷酸化位点与柱花草根系分泌苹果酸耐低磷和抗铝毒的相关性不明显，但是，SgALMT1基因表达、根系分泌苹果酸与柱花草耐低磷和抗铝毒能力具有较强的相关性。低磷和铝胁迫处理均显著增强了SgALMT1基因表达以及促进根系分泌苹果酸。并且，超量表达SgALMT1基因能增加拟南芥活化利用难溶性铝磷、植株磷浓度和耐铝毒能力，表明SgALMT1基因具有介导根系分泌苹果酸参与耐低磷和抗铝毒的生物学功能。本研究还进一步优化了农杆菌介导的柱花草遗传转化方法。因此，本研究结果表明SgALMT1基因可以作为培育耐低磷和抗铝毒柱花草新品种的重要候选基因。本研究为深入揭示热带豆科牧草适应酸性土壤逆境胁迫机理提供了重要的理论基础。