一个颠覆植物铝敏感性的类受体激酶的功能解析

Al toxicity is one of the key factors limiting crop productivity in acidic soils. Reducing Al accumulation in cell wall, and sequestrating Al inside cell into vacuole for compartmentation, are two major strategies for plants to confer high Al resistance. However, the molecular regulatory mechanisms of how plants coordinate these two processes are very poorly understood. In preliminary study, we isolated an Arabidopsis loss-of-function mutant R78 with extremely high Al resistance that harbors a mutation in a receptor-like kinase gene, and found that low concentration of Al had an obvious stimulation effect on root growth of R78 mutant. Moreover, Al accumulation in cell wall of root tips was greatly reduced, while Al content in cytoplasm was significantly increased in R78 mutant versus WT. Such subcellular distribution of Al in root was totally reversed by overexpression of R78 in WT, indicating that R78 is a key regulator coordinating the subcellular distribution of Al in plants and Al resistance. This project hence will employ techniques of RNA-seq analysis, protein interactions, genetic interactions and etc. to explore the upstream and downstream genes that act as putative Al receptor and regulator of Al distribution between cell wall and symplasm respectively, thus answering why the mutation of this gene reverses its response to Al toxicity; on the other hand, we will investigate the expression of R78’s homologous genes in diverse plant species including Al sensitive crops (barley and rape), Al resistant crop (rice) and acid and Al favorite plant (tea tree), meanwhile using overexpression and CRISPR methods to evaluate the function of these genes, so that we can characterize the role of R78 gene in plant evolutionary acclimating to acidic soils. Through the above studies, we hope to elucidate the fundamental role of R78 in plant resistant to Al toxicity and find a new direction to breed genetically modified crops that have super performance on acidic soils with no negative effects of Al toxicity, which thereby will have better root growth and resistant to other coexisting stresses in these soils.

铝毒是酸性土壤上限制作物生产的主要因素。减少铝在细胞壁中的积累以及将进入细胞内的铝进行区室化是植物高抗铝性的根本策略，但对协调这两种途径的调控机制仍不清楚。我们得到一个抗铝性极强且低浓度铝明显刺激根系生长的拟南芥类受体激酶突变体R78，该突变体根尖细胞壁铝含量明显降低，而共质体铝含量显著提高；超表达该基因后，上述分布发生逆转，说明该基因是通过调控细胞内铝分布而影响植物铝抗性的关键节点。本项目将运用转录组、蛋白互作以及遗传互作筛选等手段，挖掘该类受体激酶的上游受体和调控铝分布的下游功能基因，系统地解析其感受及协同消除铝毒的分子生理机制；此外，系统地分析该基因的同源基因在对铝敏感的大麦和油菜，对铝抗性较强的水稻及喜铝的茶树等植物中的表达情况，并通过基因操作技术，进行该基因的功能分析与验证，希望从进化的角度来阐明喜酸铝植物进化的分子基础，为从根本上解决酸性土壤上的铝毒问题指明新的方向。

铝毒是酸性土壤上限制作物生产的主要因素，也是酸沉降导致森林退化的重要因素。尽管植物的耐铝研究已有几十年之久，但植物如何感知铝离子，其受体是什么，这些最基本的科学问题仍然未知。本项目是在前期工作中得到了一个负调控铝抗性的胞质类受体激酶R78的基础上开展的，旨在解析其调控铝抗性的信号通路。在项目的资助下，我们证明了R78一方面通过潜在直接调控AHA蛋白的活性调节低铝条件下的根系生长，另一方面通过潜在调控MSL10活性以及间接调控细胞壁相关基因的表达来调节高铝胁迫下的抗铝性。同时我们进一步发现了与胞质受体激酶R78互作，且通过磷酸化调控其活性的受体激酶PSKR1（也命名为ALR1, Al Receptor 1），证明了PSKR1是首个铝离子受体，揭示了从铝离子感知到关键蛋白激活，到核心转录因子累积，再到下游抗铝反应启动的完整信号通路。研究发现了PSKR1胞内结构域可以识别铝离子，铝离子的结合促进PSKR1招募共受体蛋白BAK1等SERK激酶，进而增强PSKR1对NADPH氧化酶RbohD的磷酸化，促进活性氧ROS的产生；ROS通过氧化修饰F-box蛋白RAE1抑制后者对核心转录因子STOP1的泛素化降解，从而促进STOP1的蛋白累积，进而激活ALMT1等下游抗铝基因的表达来启动抗铝反应。此外，本项目通过解析苹果酸转运蛋白ALMT1的高分辨率结构，阐明了铝离子激活ALMT1转运苹果酸提高抗铝性的分子机制。因此，本项目揭示了植物从胞内和胞外两种途径感知铝离子并启动抗铝反应的分子机制。该研究成果在领域内具有里程碑的意义，为今后抗酸铝作物的定向育种提供了重要的理论依据。项目已发表标注SCI论文3篇，核心成果的论文Science期刊审稿中；获得发明专利1项，PCT专利已公布。