我所研究揭示广谱抗性蛋白RPW8.1增强拟南芥和水稻的PTI抗性

近日，我所王文明团队和美国马里兰大学萧顺元教授团队合作，在Plant biotechnology Journal在线发表了题为“RESISTANCE TO POWDERY MILDEW8.1 boosts pattern-triggered immunity against multiple pathogens in Arabidopsis and rice”的研究论文。该研究通过对拟南芥广谱抗性蛋白RPW8.1转基因株系进行转录组测序分析和系统抗性分析，揭示了RPW8.1通过增强拟南芥PTI（PAMP-Triggered Immunity）来提高对细菌和真菌的抗性；有意思的是，异源表达RPW8.1的转基因水稻对稻瘟病和白叶枯病表现抗性，证明RPW8.1可在水稻上强化PTI，为水稻抗性改良提供了新的选择基因。

RPW8（Resistance to Powdery Mildew Locus 8）是部分生态型拟南芥特有的位点，包含RPW8.1与RPW8.2两个紧密连锁基因。马里兰大学萧顺元教授经过多年研究证明，这两个基因分别或联合对所有能侵染拟南芥的4个白粉菌种表现抗性(Xiao et al., 2001)，也明显抗烟草白粉病、拟南芥霜霉病和烟草花叶病毒病等多种植物寄生性病害(Wang et al., 2007)。不同于其他NBS-LRR抗性基因，RPW8.1和RPW8.2均编码长度为100多个氨基酸的蛋白质，蛋白结构独特，抗性广谱且持久(Orgil et al., 2007)，但与NB-LRR类抗病基因一样，RPW8仍然利用水杨酸（Salicylic acid）信号传导途径表达抗病性(Xiao et al., 2005)。RPW8.2介导的抗性机理比较清楚：RPW8.2被病原菌诱导产生后，精准地定位到寄主-病原菌界面，激发如H2O2和胼胝质等抗病物质在寄主-病原菌界面的高浓度积累，限制病原菌的入侵；当这步防御应答不足以抵御病原菌的入侵时，RPW8.2的表达量便不断增加直至受侵染的寄主细胞死亡，从而达到广谱抗病的效果(Wang et al., 2009)。

RPW8.1定位于叶绿体和质体附近，其作用机理尚不清楚。在拟南芥中，通过转录组测序分析发现，与对照材料相比，在由RPW8.1自己的启动子表达RPW8.1-YFP的转基因株系中,许多PAMP（flg22和chitin）诱导的基因转录水平呈组成型上调；当用PAMP处理这些转基因植株后，RPW8.1蛋白表达显著增高，转基因植株表现更强的胼胝质沉积，活性氧产生，抗性基因转录和产生类似超敏反应的细胞坏死等反应。毒性细菌在RPW8.1转基因植株上的增殖被显著抑制，可能归因于RPW8.1部分抑制毒性菌效应因子（effectors）的分泌和部分抑制效应因子的活性。当PTI信号受体（FLS2或CERK1）缺失或下游关键信号分子（BIK1）缺失时，RPW8.1丧失对PTI的增强作用。

在水稻中，利用水稻自身启动子，异源表达RPW8.1-YFP融合蛋白，可显著增强水稻对稻瘟病菌（Pyricularia oryza，真菌性病原菌）和百叶枯菌（Xanthomonas oryzae pv. oryzae，细菌性病原菌）的抗性。这些结果证明RPW8.1介导的抗性和PTI之间存在紧密联系，RPW8.1作为一个广谱抗性基因，可以用于基因工程改良植物，包括单子叶类作物的抗病性。

该研究由四川农业大学和美国马里兰大学合作完成，李燕副研究员和硕士生张勇、王青霞、王婷婷为论文第一作者。相关工作得到了国家自然科学基金、四川农业大学高端人才项目资助。

作者：Yan Li,Yong Zhang,Qing-Xia Wang,Ting-Ting Wang,Xiao-Long Cao,Zhi-Xue Zhao,Sheng-Li Zhao,Yong-Ju Xu,Zhi-Yuan Xiao,Jin-Lu Li,Jing Fan,Hui Yang,Fu Huang,Shunyuan Xiao,Wen-Ming Wang

摘要：The Arabidopsis gene RESISTANCE TO POWDERY MILDEW8.1 (RPW8.1) confers resistance to virulent fungal and oomycete pathogens that cause powdery mildew and downy mildew, respectively. However, the underlying mechanism remains unclear. Here, we show that ectopic expression of RPW8.1 boosts pattern-triggered immunity (PTI) resulting in enhanced resistance against different pathogens in both Arabidopsis and rice. In Arabidopsis, transcriptome analysis revealed that ectopic expression of RPW8.1-YFP constitutively up-regulates expression of many pathogen-associated molecular pattern-(PAMP-) inducible genes. Consistently, upon PAMP application, the transgenic line expressing RPW8.1-YFP exhibited more pronounced PTI responses such as callose deposition, production of reactive oxygen species, expression of defense-related genes and hypersensitive response-like cell death. Accordingly, the growth of a virulent bacterial pathogen was significantly inhibited in the transgenic lines expressing RPW8.1-YFP. Conversely, impairment of the PTI signaling pathway from PAMP cognition to the immediate downstream relay of phosphorylation abolished or significantly compromised RPW8.1-boosted PTI responses. In rice, heterologous expression of RPW8.1-YFP also led to enhanced resistance to the blast fungus Pyricularia oryzae (syn. Magnaporthe oryzae) and the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo). Taken together, our data suggest a surprising mechanistic connection between RPW8.1 function and PTI, and demonstrate the potential of RPW8.1 as a transgene for engineering disease resistance across wide taxonomic lineages of plants.

出版源：http://onlinelibrary.wiley.com/doi/10.1111/pbi.12782/full