茉莉酸调控番茄丛枝菌根形成的机理研究

Currently, the interaction between plants and soil microorganisms is one of the most interesting topics in the interdisciplinary research amongst ecology, plant science, microbiology, soil science and agricultural science, wherein the arbuscular mycorrhizal (AM) is one of the most classic experiment model. AM is the most widespread terrestrial symbiosis, and at least 80% of land plant species can form AM with Glomeromycota fungi. The AM hyphal network supply the host plant with nutrients (predominantly phosphate) and water, and obtain in turn carbohydrates from the host. AM symbiosis contributes significantly to global phosphate and carbon cycling and influences primary productivity in terrestrial ecosystems. In addition, AM can improve efficiently its host resistance to pathogens and various abiotic stresses. The study on AM formation mechanism is not only necessary to improve AM formation efficiency, also provides scientific reference for realizing the interaction mechanisms between host plants and rhizobia or pathogens. Recent years, increasing evidence has demonstrated that jasmonic acid (JA) plays a crucial role in the several key stages during the symbiosis between plants and AM fungi, but the molecular mechanisms remain to be elucidated. In planta, a complex interaction, either synergistically or antagonistically, is existed between JA and other signal molecules such as salicylic acid or ethylene. Additionally, a pleiotropic effect and a time effect are the distinctive special feature of JA. Therefore, we speculate that the regulatory role of JA in AM formation may be involved in complex biological processes, wherein a cross talk may be existed. In addition, the biological functions of JA may be dynamically changed depending on the AM developmental stages. In order to reveal these, we will use proteomic and real-time PCR techniques to dissect the action mechanism of JA in the symbiosis between tomato and Glomus mosseae or G. intraradices. JA-altering tomato mutants are employed including the deficient mutation in JA biosynthetic pathway named as spr2 and def1, and JA signaling blockage mutant jar1. In addition, grafting, root-splitting and application of exogenous JA on the aerial parts are also performed. More specifically, the total proteins are separated by 2-D electrophoresis, and the differential expression proteins are analyzed by MALDI-TOF/TOF mass spectrometer to obtain peptide mass fingerprints, and then matched automatically to the known proteins in database. Furthermore, the expression pattern of scored proteins with important functions in AM formation are investaged by real-time PCR. We believe that the study results are helpful to reveal the biological processes responsible for the JA-mediated AM formation and their cross talk mechanism, and to elucidate the time-course of JA action and the systemic action manner of aerial JA on AM formation.

植物与微生物之间的相互作用是当前多学科交叉研究的重要课题，其中对丛枝菌根形成机理的研究不仅是提高其形成效率进而改善生态系统的前提条件，也为认识植物与其它有益菌或病原菌的互作机理提供借鉴。近年来，增加的证据表明茉莉酸在菌根形成的多个阶段起关键调节作用，但其分子机理尚不清楚。基于茉莉酸生物学作用的多效性、时效性以及与其它信号分子之间的协同或拮抗等特性,我们推测茉莉酸对菌根形成的调节涉及复杂的生物学过程及其相互间的交叉对话（cross talk），且在菌根形成过程中具有动态调节机制。为了揭示之，本项目拟以番茄野生型及其茉莉酸生物合成途径不同位点的缺失突变体spr2和def1、信号转导阻断突变体jai1为材料，采用嫁接、分根和外施茉莉酸等手段，通过蛋白组学研究和基因表达分析，摸清茉莉酸在菌根形成中的调节机理。研究结果有助于揭示茉莉酸调节作用的生物学过程及其相互间的交叉对话、作用时序和系统作用方式。

丛枝菌根是自然界广泛存在的植物与真菌互利互惠共生体，80%以上的陆生植物可借助其吸收营养和水分、提高对逆境的耐受性以及加强物质和能量循环。由于菌根形成是一个十分复杂的生物学过程，对其形成机理仍缺乏足够的认识。基于此，本研究重点探讨了茉莉酸在菌根形成过程中可能的调节机制。主要研究内容为：茉莉酸对菌根形成的影响、菌根形成过程中茉莉酸对蛋白质表达水平的调节、差异表达蛋白质的生物学功能。研究目标概括为：阐明茉莉酸调控菌根形成的生物学过程及其交叉对话机制、茉莉酸在菌根形成的几个关键阶段的特异性作用方式、地上部分的茉莉酸对菌根形成的系统作用方式。实验采用了番茄茉莉酸生物合成和信号转导相关的缺失突变体，并设计了嫁接、叶面喷施茉莉酸等实验，运用蛋白组学、基因表达分析和生理生化分析等技术，揭示了茉莉酸在菌根形成中可能的调节机制。研究发现，接种10天时，寄主的蛋白组学变化幅度远大于菌根定殖阶段的水平，显示菌根形成初期是更关键的阶段。因此，后续工作主要围绕这一阶段进行。主要（或重要）发现有：（1）根激素水平及信号转导参与菌根形成调节，其中茉莉酸、乙烯和脱落酸起正作用，而水杨酸可能起负作用；（2）根角质/木栓质起正调节作用，这在茉莉酸调节菌根形成中还未见报道；（3）氨基酸对菌根形成可能起正调节作用；（4）类黄酮生物合成与基因表达调控起重要作用；（5）细胞壁甲酯化起重要调节作用，这在茉莉酸调节菌根形成中尚未见报道；（6）中性蔗糖酶起正调节作用，这在茉莉酸调节菌根形成中尚未见报道；（7）硝酸盐代谢起正调节作用；（8）转录因子和激酶参与菌根形成调节；（9）蛋白质泛素化参与菌根形成的调节，这在茉莉酸调节菌根形成中尚未见报道；（10）光合作用过程参与菌根形成；（11）抗氧化防卫反应在菌根形成中起调节作用。上述发现有助于对菌根形成机制的认识，并为进一步深入研究积累了资料、指明了方向，因此具有重要理论意义和学术价值。