Interplay of dirgent and jacalin-proteins in broad-spectrum disease resistance of plants against fungal pathogens

dirgent 和 jacalin 蛋白在植物对真菌病原体的广谱抗病性中的相互作用

• 批准号: 369034981
• 负责人: Dr. Thomas Classen
• 金额: --
• 依托单位: Institut für Bio- und Geowissenschaften (IBG)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2017
• 资助国家: 德国
• 起止时间: 2016-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/369034981?language=en
• 关键词: Interplay; dirgent; jacalin; proteins; broad

Fusion proteins consisting of a jacalin-related lectin (JRL) and a dirigent domain occur exclusively within the plant family Poaceae. The group of Ulrich Schaffrath has shown that constitutive over-expression of different members of this family in rice, barley and wheat, respectively, led to quantitative broad-spectrum disease resistance against different pathogens including bacteria, oomycetes and fungi. Recombinant proteins with an artificially separated JRL-domain localizes towards the site of penetration when expressed in barley attacked by the powdery mildew fungus. When co-expressed, also a protein consisting of the separated dirigent domain localizes at the penetration site together with the JRL-protein. The activity of both protein partners is required for enhanced pathogen resistance.Based on these results we hypothesize, that JRL- and dirigent proteins are part of an ancient plant defense system which consists of two components, one acting on pathogen recognition and the other on execution of resistance. Fusion of the respective proteins is an invention solely realized in Poaceae species. In the proposed project we will challenge this hypothesis by answering the following questions: i) which carbohydrate signature is exposed during pathogen attack and binds the JRL-domain; ii) which antimicrobial substance is produced in the presence of the dirigent-domain protein; and iii) are there pairs of JRL and dirigent proteins in other plant species that cooperatively act in the proposed manner. In particular, domain swap experiments will be performed with the domains of different Poaceae fusion proteins to unravel functional similarities. The identification of the JRL-domain binding partner and characterization of the biochemical reaction in which the dirigent-domain is involved will be targeted by the group of Thomas Classen, a biochemist. This will include synthesis of recombinant proteins, crystallography, pull-down assays of JRL-lectins with carbohydrates from pathogens and/or plants as well as HPLC-analyses.Arabidopsis will be used as model plant, to search for pairs of proteins that act in disease resistance in a similar manner as the above mentioned Poaceae-specific fusion proteins. Using bioinformatics, we already identified two promising candidate genes which are located next to each other on Arabidopsis chromosome 1. Both proteins will be expressed in barley to elucidate their involvement in penetration resistance. Promising candidates of JRL- and dirigent proteins will be used to construct an artificial fusion protein and test whether this is capable to provide broad-spectrum disease resistance. This finally will prove if translational approaches, in which artificial fusion proteins are expressed in crop plants, are a promising strategy to improve plant stress resistance.The combined expertise of the Schaffrath and Classen labs in molecular phytopathology and biochemistry will build a solid base for the realization of the project.

由jacalin相关凝集素（JRL）和发散域组成的融合蛋白仅存在于禾本科植物中。Ulrich Schaffrath的研究小组已经表明，该家族的不同成员分别在水稻、大麦和小麦中的组成型过表达导致了对不同病原体（包括细菌、卵菌和真菌）的定量广谱抗病性。具有人工分离的JRL结构域的重组蛋白在被白粉病真菌攻击的大麦中表达时定位于穿透位点。当共表达时，由分离的发散结构域组成的蛋白质也与JRL蛋白一起定位在穿透位点。这两种蛋白质伴侣的活性是增强病原体抗性所必需的。基于这些结果，我们假设，JRL-和dirigent蛋白是一个古老的植物防御系统的一部分，该系统由两个组成部分组成，一个作用于病原体识别，另一个作用于抗性的执行。各蛋白质的融合是仅在禾本科物种中实现的发明。在所提出的项目中，我们将通过回答以下问题来挑战这一假设：i）在病原体攻击期间暴露的碳水化合物特征并结合JRL结构域; ii）在存在差异结构域蛋白的情况下产生的抗菌物质;以及iii）在其他植物物种中是否存在以所提出的方式合作作用的JRL和差异蛋白对。特别地，将用不同禾本科植物融合蛋白的结构域进行结构域交换实验以解开功能相似性。生物化学家托马斯·克拉森（Thomas Classen）的小组将针对JRL结构域结合配偶体的鉴定和其中涉及drigent结构域的生化反应的表征。这将包括重组蛋白的合成、晶体学、JRL-凝集素与来自病原体和/或植物的碳水化合物的下拉测定以及HPLC分析。拟南芥将被用作模式植物，以寻找以与上述Poaceae特异性融合蛋白类似的方式在抗病性中起作用的蛋白质对。利用生物信息学，我们已经确定了两个有希望的候选基因，它们位于拟南芥1号染色体上。这两种蛋白质将在大麦中表达，以阐明它们参与抗渗透性。JRL和dirigent蛋白的有前途的候选者将被用于构建人工融合蛋白，并测试这是否能够提供广谱抗病性。这最终将证明，在作物中表达人工融合蛋白的翻译方法是否是提高植物抗逆性的一种有前途的策略。Schaffrath和Classen实验室在分子植物病理学和生物化学方面的专业知识将为该项目的实现奠定坚实的基础。