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Elicitor release upon flagellin glycan modification

鞭毛蛋白聚糖修饰后激发子的释放

基本信息

批准号：
BB/R017913/1
负责人：
Renier Van Der Hoorn
金额：
$ 62.12万
依托单位：
University of Oxford
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2018
资助国家：
英国
起止时间：
2018 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FR017913%2F1
关键词：
Elicitor release upon flagellin glycan

项目摘要

Immunity to pathogens in plants is essential for agriculture and even for life on earth. Pathogen recognition is the first crucial step in plant immunity. Most plants recognise bacterial pathogens by their flagella, which bacteria use to move. Different plant species can recognise different fragments of the flagellin proteins, though most seed plants recognise a conserved, 22 amino acid region, known as flg22, the major recognition element known for bacterial pathogens. Recognition of flg22 at the cell surface by the FLS2 receptor-like kinase is very well studied. Flg22 perception elicits immune responses in most plants, including the model plant Arabidopsis thaliana and tobacco relative Nicotiana benthamiana. These immune responses include an oxidative burst, MAP kinase signalling and transcriptional reprogramming, mounting a defence response that includes cell wall strengthening and the secretion of toxic metabolites and harmful proteins. The relevance of flg22-triggered immunity is stressed by the fact that adapted bacterial pathogens all use effectors to block flg22-induced signalling. Currently, flg22-FLS2 signalling is the best understood and most important recognition system of plant pathogens. A crucial step, however, is still unresolved. Flagellin-derived elicitors like flg22 are embedded deep within the structure of flagellin protein and reside inside the flagellar rod. How can these buried elicitors bind to the FLS2 receptor? It seems obvious that they must be released by extracellular hydrolase activities, but elicitor-releasing hydrolases and even the naturally-released elicitor have not yet been identified. One may expect that bacteria suppress these elicitor-releasing hydrolases during infection to prevent their recognition, so it is very likely that flagellin hydrolysis represents an important battlefield at the plant-pathogen interface. Importantly, we discovered an extracellular galactosidase-like GH35 glycosidase that releases flagellin-derived elicitors from bacteria. The relevance of this GH35 enzyme was indicated by a suspicious cover-up: GH35 is specifically inhibited during bacterial infection by a bacterial metabolite. Importantly, GH35 treatment of the model bacterial pathogen Pseudomonas syringae triggers an oxidative burst in Nicotiana benthamiana and Arabidopsis thaliana. This oxidative burst is dependent on the FLS2 receptor in the plant, and the flagellin-encoding fliC gene in the pathogen. Mutant N. benthamiana lacking the GH35 enzyme are more susceptible for P. syringae, confirming their role in immunity.Flagella are glycosilated with a unique trisaccharide glycan. We HYPOTHESISE that GH35 acts on these glycans and that this modification, in concert with other hydrolases, results in the release of flagellin-derived elicitors that bind to FLS2. Using well described P. syringae mutants with altered flagellin glycosylation and Nicotiana benthamiana as a host, we are in the unique position to test how GH35 contributes to immunity. The AIM of this proposal is to elucidate how flagellin-derived elicitors are released by GH35 and other hydrolases and to investigate how common this mechanism is in plant-bacteria interactions. The OBJECTIVES are to: i) elucidate the role and mechanism of GH35 modification of flagellin; ii) investigate the broader role of GH35-mediated elicitor release from flagellin in crop plants and from other pathogens; and iii) elucidate protein processing of flagellin upon GH35 treatment.This project will lead to the elucidation of an important novel mechanism in bacterial pathogen recognition by plants that is probably universal in the plant kingdom. Similar hydrolase-driven elicitor release is expected for the recognition of filamentous pathogens. These discoveries will inspire new crop protection strategies, including the introduction of inhibitor-resilient hydrolases and agrochemicals blocking flagellin glycosylation or GH35 inhibitor biosynthesis.

植物对病原体的免疫力对农业甚至地球上的生命至关重要。病原体识别是植物免疫的第一个关键步骤。大多数植物通过鞭毛识别细菌病原体，细菌利用鞭毛移动。不同的植物物种可以识别鞭毛蛋白的不同片段，尽管大多数种子植物识别一个保守的22个氨基酸区域，称为flg 22，这是已知的细菌病原体的主要识别元件。FLS 2受体样激酶在细胞表面对flg 22的识别已得到很好的研究。Flg 22感知在大多数植物中激发免疫应答，包括模式植物拟南芥（Arabidopsis thaliana）和烟草近缘种本塞姆氏烟草（Nicotiana benthamiana）。这些免疫反应包括氧化爆发、MAP激酶信号传导和转录重编程，产生防御反应，包括细胞壁强化和分泌有毒代谢物和有害蛋白质。适应性细菌病原体都使用效应子阻断flg 22诱导的信号传导，这一事实强调了flg 22触发的免疫的相关性。目前，flg 22-FLS 2信号转导是植物病原体最好理解和最重要的识别系统。然而，关键的一步仍未解决。鞭毛蛋白衍生的激发子如flg 22深深嵌入鞭毛蛋白的结构中，并驻留在鞭毛杆内。这些隐藏的激发子如何与FLS 2受体结合？很明显，它们必须通过胞外水解酶活性释放，但激发子释放水解酶，甚至天然释放的激发子尚未被确定。人们可以预期，细菌在感染过程中抑制这些激发子释放水解酶，以防止它们的识别，因此鞭毛蛋白水解很可能是植物-病原体界面的一个重要战场。重要的是，我们发现了一种细胞外半乳糖苷酶样GH 35糖苷酶，可从细菌中释放鞭毛蛋白衍生的诱导子。这种GH 35酶的相关性由可疑的掩盖表明：GH 35在细菌感染期间被细菌代谢物特异性抑制。重要的是，GH 35处理的模式细菌病原体假单胞菌（Pseudomonasalumingae）在本氏烟草（Nicotianabenthamiana）和拟南芥（Arabidopsis thaliana）中引发氧化爆发。这种氧化爆发依赖于植物中的FLS 2受体和病原体中的鞭毛蛋白编码fliC基因。突变N.缺乏GH 35酶的本塞姆氏菌更易感染P. lingae，证实了它们在免疫中的作用。我们假设GH 35作用于这些聚糖，这种修饰与其他水解酶一起导致鞭毛蛋白衍生的激发子与FLS 2结合。使用描述良好的具有改变的鞭毛蛋白糖基化的P. lingae突变体和本氏烟草（Nicotiana benthamiana）作为宿主，我们处于独特的位置来测试GH 35如何有助于免疫。本提案的目的是阐明鞭毛蛋白衍生的激发子是如何通过GH 35和其他水解酶释放的，并研究这种机制在植物-细菌相互作用中的普遍性。这些措施是：本项目的主要研究内容包括：（1）阐明GH 35修饰鞭毛蛋白的作用和机制;（2）研究GH 35介导的鞭毛蛋白和其他病原菌诱导子释放的广泛作用;（3）阐明GH 35处理后鞭毛蛋白的蛋白质加工过程。类似的水解酶驱动的激发子释放预期用于丝状病原体的识别。这些发现将激发新的作物保护策略，包括引入具有生物弹性的水解酶和阻止鞭毛蛋白糖基化或GH 35抑制剂生物合成的农用化学品。