A Systems Biology-aided Investigation of Pathogen-mediated Manipulation of Sugar Metabolism in Arabidopsis

植物-微生物病理系统构成了一个非常复杂的生物网络，在这个网络中，来自病原体和寄主的分子参与者参与了一场争夺优势的战斗。专门的病原体已经进化出一套称为效应器的分子，这些分子调节宿主细胞的生理并支持寄生。在过去的几十年里，大量的文献记录了诱导有效的免疫反应和颠覆效应器介导的宿主防御的分子机制。然而，一个关键的悬而未决的问题是，病原体如何改变细胞新陈代谢，包括操纵源-汇关系，以获得营养。这个跨学科的项目通过整合生物学和快速发展的计算机科学领域来促进研究、教育和基于社区的科学参与，大大超出了湿实验室技术的范围。我们将从机理上理解效应器介导的扰动以从中央液泡中动员糖。阐明病原体感染如何调节全球转录动力学并改变生物信息流是首要关注的问题。同样重要的是，代谢、激素、昼夜节律和免疫信号通路之间的串扰将被揭示，涉及致病和非致病细菌菌株的信号。该项目位于阿拉巴马州的中心，是一所国家公认的多样性大学，该项目的范围扩展到教育途径，高级基因组学课程的学生将获得相关生物信息学分析的经验，PI领导的生物教学计划的城市高中教师将接触到当今的基因创新，热心的招聘工作将寻求吸引邻近的HBCU少数族裔学生。该计划的中心框架，了解植物病原体如何通过其效应者的毒力活动获得糖，比以前单一病原体的限制维度：一种蛋白质分析方法有了实质性的进步。具体地说，这个项目的重点是一种病原体效应物HopD1，它与一组宿主蛋白物理上相互作用，形成一个功能模块，称为“空泡转化酶(Vac-INV)模块。”HopD1扰乱Vac-INV模块以动员储存在中央液泡中的糖，同时减缓质外体糖进入宿主细胞的吸收的机制将被研究。此外，还将研究通过干扰保守的水通道蛋白介导的气孔调节机制来抑制免疫应答的作用。基于Vac-INV依赖的转录回路的电子模拟，还将构建一个植物-病原菌相互作用的高分辨率全球动态转录调控网络。在已经开发了一个基于网络的界面OOCEAN，用于启动子和生物信息学分析，该项目将继续扩大其在快速发展的系统生物学世界中的用途。新颖的基于网络生物学的分析将有助于更广泛地了解单一效应器如何同时针对植物免疫和新陈代谢的不同分支，以获得病原体的优势。这项工作还将产生一个系统，该系统也可以应用于与植物防御和新陈代谢有关的其他重要问题。总而言之，这个系统水平的项目将巩固全球对效应器-宿主相互作用、效应器介导的干扰以及随后的动态转录调控的理解。

The plant-microbe pathosystem constitutes a very complex biological network in which the molecular players from both the pathogen and the host engage in a battle for dominance. Specialized pathogens have evolved suites of molecules called effectors that modulate host cell physiology and support parasitism. Over the past decades, a plethora of literature has documented the molecular mechanisms that underlie the induction of effective immune responses and subversion of effector-mediated host defenses. However, a key unresolved question is how pathogens alter cellular metabolism, including manipulation of the source-sink relationships, to acquire nutrients. This interdisciplinary project extends significantly beyond wet-lab techniques by integrating both biology and the rapidly evolving field of computer science for the advancement of research, education, and community-based scientific engagement. Mechanistic understanding of effector-mediated perturbations to mobilize sugars from the central vacuole will be pursued. Elucidating how pathogen infection modulates the global transcriptional dynamics and alters the flow of biological information is of prime focus. Equally paramount, cross-talk between metabolic, hormonal, circadian and immune signaling pathways, involving signals from pathogenic and non-pathogenic bacterial strains, will be revealed. Situated in the heart of Alabama, at a nationally recognized university for diversity, this project's scope extends into educational avenues as students in advanced level genomics courses will gain experience with relevant bioinformatic analyses, urban high school teachers in the PI's-led BioTeach program will gain exposure to the genetic innovations of today, and zealous recruiting efforts will seek to engage neighboring HBCU-minority students.The central framework of this proposal, understanding how plant pathogens acquire sugars through the virulence activities of their effectors, presents a substantial advancement over the previously limiting dimensions of a one pathogen: one protein analysis approach. Specifically, this project is focused on a pathogen effector, HopD1, that physically interacts with a set of host proteins forming a functional module, termed 'vacuolar invertase (Vac-INV) module.' The mechanisms by which HopD1 perturbs the Vac-INV module to mobilize sugars stored in the central vacuole while slowing down the resorption of apoplastic sugars into the host cell will be investigated. HopD1's immune response suppression through interference of a conserved aquaporin-mediated stomatal regulation mechanism will also be investigated. Based on the in silico modeling of Vac-INV?dependent transcriptional circuits, a high resolution global dynamic transcriptional regulatory network in plant-pathogen interactions will also also constructed. Having already developed a web-based interface, OOCEAN, for promoter and bioinformatics analyses, this project will continue to extend its utility in the fast-evolving world of Systems Biology. Novel network biology-based analyses will help obtain a broad understanding of how single effectors simultaneously target different branches of plant immunity and metabolism for the pathogen's advantage. This work will also result in a system that can be applied to other important questions pertaining to plant defense and metabolism as well. Collectively, this systems-level project will solidify global understanding of effector-host interactions, effector-mediated perturbations, and subsequently, dynamic transcriptional regulation.