玉米ZmCLE12多肽调控AM真菌共生信号转导的分子机制

Most plants have the ability to establish a symbiosis with arbuscular mycorrhizal (AM) fungi, which facilitates the acquisition of limited nutrients and improves plant resistance against stress. A perception of signal molecules is essential for the establishment of both AM and root symbioses. Our previous results showed that a ZmCLE12 peptide from Zea mays could be perceived by a leucine-rich repeat receptor-like protein kinase (ZmLRR-RLK1), resulting in enhanced infection efficiency of AM fungi. However, how the ZmLRR-RLK1 receptor recognizes ZmCLE12 and activates the symbiotic signaling pathway are currently unknown. To analyze the interaction pattern between both of them, we will first investigate the homologous or heterologous protein complex of ZmLRR-RLK1 induced by ZmCLE12. The functional ectodomain and key amino-acid sites of ZmLRR-RLK1 binding to ZmCLE12 are also analyzed by the experiments of site directed mutagenesis and ligand interaction. Additionally, we further screen and identify the interaction proteins with the kinase domain of ZmLRR-RLK1, and survey the expression of symbiotic genes regulated by these proteins. Taken together, our results are benefit to uncover the molecular mechanism of AM symbiotic signaling regulated by ZmCLE12. By contrast, the analysis of peptide regulating AM symbiosis is innovative. It not only helps us widely understand the recognition mechanism of plant-microbial interaction, but also provides an important guideline for the development of polypeptide preparations, facilitating the symbiotic establishment between AM fungi and plants.

丛枝菌根（AM）真菌可与植物形成互惠共生体，促进根系对营养元素的吸收，增强植株的抗逆性，二者互作应答是共生关系建立的关键。项目组前期发现玉米ZmCLE12多肽可显著提高AM真菌侵染率，而这一信号的识别依赖于LRR蛋白激酶受体ZmLRR-RLK1。但ZmCLE12多肽如何特异地与受体互作、介导胞内信号转导仍不清楚。本项目通过对多肽诱导受体同源或异源复合体形成的鉴定，确定受体与配体的互作模式；利用定点突变和配体互作实验，明确ZmCLE12多肽特异结合受体胞外结构域及关键氨基酸位点；分析ZmCLE12多肽与ZmLRR-RLK1胞外结构域的互作蛋白对下游共生基因表达的影响，从而初步阐明ZmCLE12多肽信号分子调节玉米-AM真菌共生信号转导的分子机制。本项目探讨多肽调控共生机制具有创新性，其结果对丰富植物-微生物共生互作理论，开发AM真菌与植物共生新型多肽促进剂具有重要意义。

丛枝菌根（AM）真菌可与大部分陆生植物形成互惠共生体，促进根系对营养元素的吸收，增强植株的抗逆性，二者互作应答是共生关系建立的关键，植物特定多肽在菌根共生建立的过程中具有重要的调控作用。本项目研究了ZmCLE12多肽影响AM真菌共生效应,进行了ZmCLE12多肽受体蛋白激酶基因筛选和表达分析，研究了ZmCLE12多肽与受体蛋白胞外结构域的互作,并利用受体蛋白激酶基因ZmLRR-RLK1突变体研究了该基因参与菌根共生的生物学功能。结果表明:ZmCLE12多肽能显著地影响AM真菌与玉米共生率,20μM浓度最显著;通过对12个ZmLRR-RLK基因表达分析获得4个ZmCLE12多肽受体候选基因;利用ZmCLE12多肽与候选受体蛋白激酶胞外结构域进行互作分析以及ZmCLE12多肽与菌根类似物CO4结合实验,发现ZmCLE12多肽与ZmLRR-RLK1蛋白胞外结构域能够结合,通过同源二聚体识别ZmCLE12多肽,是ZmCLE12重要受体蛋白;在此基础上,利用ZmLRR-RLK1基因EMS突变体与AM真菌共生率及其共生信号通路基因表达分析,探明了该基因具有影响AM真菌与玉米共生的生物学功能,从而初步揭示了ZmCLE12多肽与受体ZmLRR-RLK1蛋白胞外结构域互作调控共生信号转导的机制。该研究结果对丰富玉米与AM真菌共生互作理论,开发新型多肽菌剂具有重要意义。