DELT与RBOHD协同调控活性氧迸发的机制研究

ROS (Reactive Oxygen Species) are very important signal molecules in plant innate immunity. It has been known for a long time that ROS are interconnected between different organs, however how the apoplastic ROS regulate the intracellular ROS is still unclear. NADPH oxidases are evolutionally conserved enzymes that mediate the ROS burst, and also called RBOHs (Respiratory Burst Oxidase Homologs) in plants. Within this family, RBOHD plays the most important role in regulating ROS burst against pathogen attack. Lipopolysaccharide (LPS) is a component of the outer membrane of Gram-negative bacteria. Lipid A, the active moiety of LPS, induces the burst of apoplastic and intracellular ROS in Arabidopsis. We have isolated four putative mutants defective in LPS-triggered ROS burst (named delt). Genetic analysis suggested that these four delt mutants were allelic, and the ROS defection was caused by point mutations in a gene encoding a receptor-like cytoplasmic kinase. Our preliminary study indicated that DELT associated with RBOHD. Interestingly, deltΔC mutants, carrying a truncated DELT which is caused by the T-DNA insertion in the C-termini of DELT gene, showed normal apoplastic ROS burst, but defective in intracellular ROS burst after LPS treatment. These results implied the different mechanisms of DELT in regulation of RBOHD-mediated apoplastic and intracellular ROS burst. Therefore, we propose to exploit the molecular mechanism of how DELT and RBOHD synergistically regulate the ROS burst between apoplasts and intracellular organs, which will provide theoretical guidance for developing broad-spectrum resistant varieties.

活性氧（ROS）迸发是植物响应外界刺激的重要免疫信号。根据产生部位，植物ROS包括质外体ROS和胞内ROS，但两者之间的交流机制尚不清楚。植物中NADPH氧化酶RBOHD在质外体ROS和胞内ROS迸发中都起作用。脂多糖是革兰氏阴性细菌细胞壁的重要组分，其活性成分lipid A能够诱导植物产生质外体和胞内ROS。根据这一表型，我们筛选了对lipid A诱导的ROS不敏感突变体delt。克隆后发现DELT编码一个胞内类受体激酶，而且与RBOHD存在互作。有意思的是，delt 羧基端缺失（deltΔC）的突变体在lipid A处理后叶绿体ROS异常，而质外体ROS正常，暗示了DELT通过不同机制特异调控RBOHD介导的质外体ROS和胞内ROS的迸发。因此，本项目将探究DELT与RBOHD协同调控质外体ROS与胞内ROS产生的分子机制，以期为开发植物广谱抗病品种提供理论指导。

植物通过多层次的免疫系统抵抗病原菌的入侵，而活性氧迸发在多层次免疫系统中都起着重要的信号作用。植物抗病过程中活性氧迸发主要是由呼吸爆发氧化物同系酶RBOHD产生，一般认为在不同的免疫层次中，RBOHD被不同的胞内受体激酶RLCKs激活。但我们发现一个胞内受体激酶RIPK在所有的免疫系统中都起作用。病原菌激发子处理后，RIPK可以直接磷酸化RBOHD的N端S343/S347。RIPK自身的磷酸化也是RBOHD介导的ROS迸发所必须的。RIPK和RBOHD突变后，气孔免疫、系统获得抗性、以及对死体营养型病原菌的抗性都显著减低。综上所述，RIPK和 RBOHD协同调控了植物抗病过程中广谱的活性氧迸发。