Study of chloroplast stromules during PCD and inter-organellar communication

项目摘要 这个项目的重点是叶绿体作为免疫信号产生的中心参与者，以及 针对病原体感染的先天性免疫反应所需的程序性细胞死亡(PCD)的调节。 虽然线粒体在哺乳动物PCD中起着核心作用，但新的证据表明，在植物中 叶绿体在执行限制病原体传播的局部性PCD方面具有关键作用。黄瓜叶绿体 除了参与免疫信号的产生，如活性氧物种(ROS)和 防御激素水杨酸(SA)也直接参与病原菌的识别。有趣的是， 叶绿体在免疫反应中动态改变其形态，并发出管状投射 被称为闪光灯。这些诱导的球茎利用细胞骨架伸展，然后固定在细胞核上， 促进叶绿体核周聚集，运输叶绿体产生的ROS和防御 蛋白质进入细胞核。这个应用程序的总体目标是使用新的细胞生物学、遗传学、 蛋白质组学和计算方法解开球茎驱动周围症的机制基础 并确定从叶绿体释放的免疫信号的身份和功能 原子核。具体地说，目标1将检查植物天然免疫期间的细胞器和细胞骨架动力学。 将进一步开发量化闪电特性和动力学的新的计算方法，并 应用于了解天然免疫过程中的叶绿体、叶绿体的基质和细胞骨架的动态。两者都有 植物和动物的病原体以细胞骨架为目标，这个目标将检查病原体效应器如何改变 作为毒力策略的细胞骨架和相关细胞器动力学。AIM 2将研究免疫信号 是球茎诱导和释放叶绿体信号所必需的。不同RO的关系 免疫反应期间的来源、细胞器运动和PCD将使用以下组合进行检查 在目标1中开发的遗传工具和计算方法。在目标3中，已知叶绿体来源的免疫 信号将被表征，新的蛋白质信号将被识别。H_2O_2的下游指标 我们将对原子核进行解释和研究。质谱学将被用来识别被运输的蛋白质 从叶绿体到细胞核。将使用遗传方法来检查它们在PCD期间的作用。最后， 将研究先天免疫过程中叶绿体释放的机制，以确定它是否类似于 哺乳动物细胞凋亡过程中线粒体的释放。了解不同细胞器在PCD中的作用 先天免疫将为细胞死亡和细胞存活过程提供统一的机制基础 对传染性病原体的反应。我们的模型系统的结果将广泛影响对 细胞器到核的通讯，影响对传染病的先天免疫。

Project Summary This project focuses on chloroplasts as central player in the generation of immune signals and the regulation of programmed cell death (PCD) required for innate immune responses against pathogen infection. Although mitochondria play a central role during mammalian PCD, emerging evidence suggests that in plants chloroplasts have a critical function in executing localized PCD that limits pathogen spread. Chloroplasts in addition to being involved in the generation of immune signals such as reactive oxygen species (ROS) and defense hormone salicylic acid (SA), also participate directly in the recognition of pathogen. Interestingly, the chloroplasts dynamically change their morphology during immune responses and send out tubular projections called stromules. These induced stromules use the cytoskeleton to extend and then anchor to the nucleus, which facilitate perinuclear clustering of chloroplasts and transport chloroplast-generated ROS and defense proteins to the nucleus. The overall goal of this application is to use combination of novel cell biology, genetics, proteomics and computational approaches to unravel the mechanistic basis of stromule driven periunclear chloroplast clustering and determine the identity and function of immune signals released from chloroplasts to nuclei. Specifically, Aim 1 will examine organelle and cytoskeleton dynamics during plant innate immunity. Novel computational methods quantifying stromule features and dynamics will be further developed and applied to understand chloroplasts, their stromules, and cytoskeleton dynamics during innate immunity. Both plant and animal pathogens target the cytoskeleton, and this aim will examine how pathogen effectors alter the cytoskeleton and related organelle dynamics as a virulence strategy. Aim 2 will investigate immune signals required for stromule induction and the release of chloroplast signals. The relationship of different ROS sources, organelle movement and PCD during immune responses will be examined using a combination of genetic tools and the computational methods developed in Aim 1. In Aim 3, known chloroplast-derived immune signals will be characterized and novel protein signals will be identified. The downstream targets of H2O2 in the nucleus will be elucidated and studied. Mass spectrometry will be used to identify proteins that are transported from chloroplasts to nuclei. Genetic approaches will be employed to examine their role during PCD. Lastly, the mechanism of release from chloroplasts during innate immunity will be studied to determine if it is akin to mitochondrial release during apoptosis in mammals. Understanding the role of different organelles during PCD and innate immunity will provide a unified mechanistic basis of cell death and cell survival process that occur in response to infectious pathogens. The results from our model systems will impact broadly on understanding of organelle-to-nuclear communication that influence innate immunity against infectious diseases.