The Arabidopsis gp91-phox Gene Family and NADPH Oxidase Function

植物无法采取行动来保护自己免受环境的侵害。 它们也不像脊椎动物那样具有复杂的、循环的、基于细胞的免疫防御系统。然而，植物已经进化出一种用于病原体识别的免疫系统，该免疫系统启动一系列细胞反应，这些细胞反应共同阻止入侵者，或者可以对压力情况做出反应和反应。活性氧物质（ROI）的产生是植物中病原体感染后观察到的早期反应之一。 产生的ROI可以直接参与杀死入侵的病原体，但也可以作为诱导进一步防御的信号。质膜结合的NADPH氧化酶是植物中ROI的可能来源。这种氧化酶，也称为呼吸爆发氧化酶（RBO），最初在哺乳动物巨噬细胞中描述，是一种介导杀死微生物的多酶复合物。这种蛋白质是NADPH氧化酶的酶亚基，利用分子氧产生超氧化物。拟南芥有一个由10个成员组成的Atrboh（Arabidopsis thaliana respiratory burst oxidase homologs）基因家族，与动物同源的蛋白质称为gp91phox。然而，NADPH氧化酶的精确亚基结构及其激活的调节似乎与哺乳动物巨噬细胞不同，ROI不是执行者，而是介导防御激活以及对环境和发育过程的其他反应的信号。在哺乳动物中，gp91phox的多种亚型在不同的细胞类型中起作用，并执行不同的功能。类似地，植物中不同的NADPH氧化酶也可能介导不同组织中的质外体ROI的产生以及对不同环境刺激的响应。例如，我们的小组已经表明，AtrbohD和AtrbohF负责在防御反应期间产生ROI，并且通过Atrboh家族的这些成员产生的ROI在气孔关闭期间作为阿坝信号传导中的第二信使是必需的。此外，由AtrbohC介导的氧化爆发通过激活允许钙进出细胞的通道来调节根毛形成期间的植物细胞扩增。有趣的是，一氧化氮（NO），另一种介导哺乳动物巨噬细胞中细胞死亡的反应分子，可能与ROI一起调节植物的反应。研究表明，与病原体的相互作用触发NO的释放，ROI和NO似乎，然后，协同工作，以控制过敏反应。本项目的目标是破译，使用功能基因组学工具，植物NADPH氧化酶基因家族的功能。要测试的假设是，不同成员的Atrboh，组件的植物NADPH氧化酶，控制生产的ROI在防御反应和发育过程中，PCD和其他对环境的反应。ROI和NO之间的连接，PCD和其他反应的另一个重要调节器，也将被调查。Atrboh基因的鉴定也使得能够研究激活植物NADPH氧化酶的机制。这是特别相关的，因为哺乳动物NADPH氧化酶的重要调节元件似乎在植物氧化酶中不存在。将进行研究以鉴定ROI信号传导的更多组分，特别是参与调节Atrboh功能的信号转导途径的早期步骤的那些组分，以及ROI依赖性信号传导的推定靶标或介质。这些研究将允许进一步了解ROI在抗病反应期间的功能，以及在由ROI和NO介导的一系列发育反应中的功能。

Plants cannot move to defend themselves from aggressions in their environment. Also they do not posses a complex, circulating, cell-based immunological system of defense like vertebrates. However, plants have evolved an immune system for pathogen recognition that initiates a set of cellular responses that collectively stop the intruder or can react and respond to stress situations. Production of Reactive Oxygen Species (ROI) is one of the earlier responses observed after pathogen infection in plants. ROI produced may directly participate in the killing of the invading pathogens, but could also act as a signal that induces further defenses. A plasma membrane bound NADPH oxidase is the likely source of ROI in plants. This oxidase, also known as the respiratory burst oxidase (RBO), was initially described in mammalian macrophages and is a multienzymatic complex that mediate the killing of microbes. This protein is the enzymatic subunit of the NADPH oxidase that uses molecular oxygen to make superoxide. Arabidopsis has a 10 member Atrboh (Arabidopsis thaliana respiratory burst oxidase homologs), gene family, homologous to the animal protein called gp91phox. However, the precise subunit structure of the NADPH oxidase and the regulation of its activation appear to be different than in mammalian macrophages.It is becoming evident that ROI is not an executioner but a signal that mediate the activation of the defenses as well as other responses to the environment and developmental processes. In mammals, multiple isoforms of gp91phox act in different cell types, and perform different functions. Similarly, different NADPH oxidases in plants may also mediates production of apoplastic ROI in different tissues and in responses to different environmental stimuli. For example, our group has shown that AtrbohD and AtrbohF are responsible for the production of ROI during the defense response and that ROI generated through these members of the Atrboh family are required as second messengers in ABA signaling during stomata closure. Also, the oxidative burst mediated by AtrbohC regulates plant cell expansion during root hair formation through the activation of channels that allow calcium to move in and out of the cell.. Interestingly, Nitric Oxide (NO), another reactive molecule that mediates cell death in mammalian macrophages, may regulate plant responses in conjunction to ROI. Studies show that interaction with pathogens trigger NO release, and ROI and NO seem to, then, work synergistically to control the hypersensitive response.The goal of this project is to decipher, using functional genomics tools, the functions of the plant NADPH oxidase gene family. The hypothesis to be tested is that different members of Atrboh, components of the plant NADPH oxidase, control production of ROI during defense response and in developmental processes, PCD and other responses to the environment. The connections between ROI and NO, another important regulator of PCD and other responses, will be also investigated. The identification of the Atrboh genes also enables to study the mechanism that activates the plant NADPH oxidase. This is particularly relevant since important regulatory elements of the mammalian NADPH oxidase appear to be absent in the plant oxidase. Studies will be conducted to identify more components of the ROI signaling, especially those involved in earlier steps of the signal transduction pathway that regulate Atrboh function, as well as putative targets or mediators of ROI-dependent signaling. These studies will allow to further understand the function of ROI during disease resistance responses, as well as in a range of developmental responses mediated by ROI and NO.