Regulation of the Heat Shock Response in E. Coli

描述（由申请人提供）：维持细胞稳态和对抗压力的有效机制对所有生物都至关重要。我们研究两个对所有活细胞都至关重要的问题：如何构建对热应力（热应力）的有效响应？32向热冲击响应；HSR)，以及如何维持细胞离散区室的内稳态(？e向包络应力响应)。我们未来的工作将解决3个关键问题：1)如何自稳态反馈控制？32岁完成了吗?HSR的活性通过稳态反馈控制系统与细胞折叠状态耦合。令人惊讶的是，我们的新研究涉及FtsY，信号识别颗粒（SRP）的受体（SR），这是在该途径中将蛋白质插入内膜所必需的。我们将研究SR/SRP蛋白在反馈调节中的作用，并验证？32在这个过程中起调节作用；可能是通过延长时间？32不能用于结合RNA聚合酶。如果这些研究暗示了SR/SRP机器在细胞控制中，它们也将为这些普遍保守的GTPases的范式改变观点提供基础。2)如何整合包膜应力信号来激活？反应吗?包膜应力控制RseA的降解速率，膜跨越反西格玛因子负调节？E，因此耦合？E活动与细胞包膜的状态有关。以前，我们认为这条通路仅仅是由未组装的孔蛋白激活的，但我们最近的工作表明，激活需要两个额外的信号。我们将识别这些信号，确定三个信号之间的关系，并在体外重建这一调控途径。这些研究将提供对这两个细胞区室如何沟通的关键理解。3) srna的作用是什么？E(和？32)介导的应激反应？我们最近的工作和其他人的工作表明？E反应是下调外膜孔蛋白的srna的产生。我们将识别剩下的？E sRNAs并验证它们的功能是提供外膜孔蛋白含量的全面监测和对应激反应特性的关键控制的假设。此外，我们的初步证据表明，一个sRNA控制DnaK，一个- 32的调节因子。因此，我们还会识别吗？32个受控制的srna并筛选具有调节作用的rna。微生物占世界生物量的整整一半，对地球上的生命极为重要，原因多种多样，如在我们的生态系统中进行矿物回收，生产对生物技术和环境解毒至关重要的产品，以及引起疾病。在过去的十年里，444种细菌的基因组序列已经发表，还有数百种正在进行中。研究所有这些细菌是不可能的，但我们可以对选定的细菌进行深入研究，因为我们开发的范式广泛适用于所有细菌。我们研究了两个关键的应激反应的调节和功能，在模式生物大肠杆菌，在那里前沿的研究是可能的。这些反应是高度保守的，两者都与相关生物的发病机制有关。此外，我们正在确定的一种策略被证明是协调抗生素、毒力和环境清理中重要因素生产的反应范例。最后，我们的研究可能会发现抗生素的新靶点。

DESCRIPTION (provided by applicant): Effective mechanisms to maintain cellular homeostasis and combat stress are vital to all living organisms. We study two issues fundamental to all living cells: how to construct an effective response to thermal stress (the ?32-directed heat shock response; HSR), and how to maintain homeostasis in discrete compartments of the cell (the ?E-directed envelope stress response). Our future work will address 3 critical questions: 1) How is homeostatic feedback control of ?32 accomplished? The activity of the HSR is coupled to cellular folding state by a homeostatic feedback control system. Surprisingly, our new studies implicate FtsY, the receptor (SR) for the Signal Recognition Particle (SRP), which is required for insertion of proteins into the inner membrane in this pathway. We will examine the role of SR/SRP proteins in feedback regulation, and test the provocative hypothesis that membrane localization of ?32 plays a regulatory role in this process; possibly by amplifying the length of time ?32 is unavailable for binding RNA polymerase. If these studies implicate the SR/SRP machine in cellular control, they will also provide the basis for a paradigm changing view of these universally conserved GTPases. 2) How are envelope stress signals integrated to activate the ?E response? Envelope stress controls the rate of degradation of RseA, the membrane spanning antisigma factor that negatively regulates ?E, thereby coupling ?E activity to status of the cell envelope. Previously, we thought that this pathway was activated simply by unassembled porins but our recent work suggests that two additional signals are required for activation. We will identify these signals, determine the relationship between the three signals, and reconstitute this regulatory pathway in vitro. These studies will provide critical understanding of how the two cellular compartments communicate. 3) What is the impact of sRNAs in the ?E (and ?32) mediated stress responses? Our recent work and that of others indicates that a major aspect of the ?E response is production of sRNAs that downregulate outer membrane porins. We will identify the remaining ?E sRNAs and test the hypothesis that they function to provide overall surveillance of outer membrane porin content and critical control of the properties of the stress response. Additionally, our preliminary evidence indicates that an sRNA controls DnaK, a regulator of ?32. Therefore, we will also identify ?32-controlled sRNAs and screen for those with regulatory roles. PUBLIC HEALTH RELEVANCE: Project relevance Microbes account for fully half of the world's biomass and are of immense importance to life on earth, for reasons as diverse as performing mineral recycling in our ecosystem, making products of important for biotechnology and detoxification of the environment, and causing disease. In the last decade, the genomic sequences of 444 bacteria have been published and hundreds more are in progress. It is impossible to study all of these bacteria, but we can perform an intensive study of selected bacteria, as the paradigms we develop are widely applicable to all bacteria. We study the regulation and function of two critical responses to stress, in the model organism E. coli, where cutting edge studies are possible. These responses are highly conserved, and both have been implicated in pathogenesis in related organisms. Moreover, one of the strategies we are defining is proving to be a paradigm for responses that orchestrate production of antibiotics, virulence and agents important in environmental cleanup. Finally, our studies may uncover new targets for antibiotics.