The role of Stress Granules and G3BP1 in coupling cytosolic and nuclear stress responses

项目摘要 核糖核蛋白(RNP)颗粒是一种无膜细胞器，由RNA和 RNA结合蛋白(RBP)，通过凝聚或液-液的生物物理原理形成 相分离(LLP)。一组有趣的胞浆RNP颗粒是应激颗粒(SGS)，即 形成于各种不同的压力源之上。压力导致全局翻译抑制，导致暴露 取消翻译RNA以浓缩和聚合成SGS。不同SG蛋白的突变导致异常 和构成的SGS或RNP聚集体。这种RNP聚集体与肌萎缩侧索硬化症相关 硬化症(ALS)、额颞叶痴呆(FTD)和各种肌肉营养不良症。因此，它是高的 医学上需要了解RNP颗粒的组装和拆解原理。 虽然人们对SGS的性质、机制和功能进行了高度的研究，但对SGS的研究还很少 如果以及如何，SGS结合转录和转录后机制。因此，我的研究 目的研究SGS和/或SG装配支架蛋白G3BP1如何调控 转录，从而耦合胞质和核应激反应，以维持细胞内稳态。在……里面 在这一应用中，我结合了全基因组、计算和实验方法来1)确定 转录受SGS形成的影响，确定SG的靶基因，2)详细表征 SGS介导转录变化的机制；3)检测G3BP1的核功能 包括在转录调控中可能的直接作用。 在K99阶段，在Roy Parker博士和Robin Dowell博士的指导下，全面 使用全基因组数据分析和细胞技术的方法将被用于确定SGS 调控转录，鉴定SG靶基因，阐明这种调控的详细机制 路径。在艾米·帕尔默博士的支持下，分析SGS和/或G3BP1在 为了确定SGS和/或G3BP1是否调节金属/离子，将获得金属/离子的动态平衡 转录水平上的应激反应中的动态平衡。此外，约翰·里恩博士在 研究RNA在染色质识别中的作用将促进G3BP1的特性及其重要性 在G3BP1-染色质结合中的RNA。 在基因组技术和计算分析方面的额外K99培训将是必不可少的 建议进行研究，并在独立R00阶段推进我的过渡和进展。 这一建议的结果不仅将导致识别重要的SG和G3BP1功能， 同时也提供了对RNP颗粒如何耦合胞质和核压力的基本理解 回应。

Project Summary Ribonucleoprotein (RNP) granules are cellular membrane-less organelles comprised of RNA and RNA binding proteins (RBPs), which form through the biophysical principle of condensation or liquid-liquid phase separation (LLPS). An interesting group of cytosolic RNP granules are stress granules (SGs), that form upon a variety of different stressors. Stress leads to a global translation inhibition, causing exposed untranslating RNAs to condense and aggregate into SGs. Mutations in various SG proteins lead to aberrant and constitutive SGs or RNP aggregates. Such RNP aggregates are associated with amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and various muscular dystrophies. Therefore, it is of high medical need to understand the principles of RNP granule assembly and disassembly. While the properties, mechanisms and functions of SGs are highly studied, nothing is known about if, and how, SGs couple transcriptional and posttranscriptional mechanisms. Therefore, my research objectives focus on characterizing how SGs and/or the SG assembly scaffold protein G3BP1 regulate transcription, thereby coupling cytosolic and nuclear stress responses to maintain cellular homeostasis. In this application, I combine genome-wide, computational and experimental approaches to 1) determine if transcription is affected by the formation of SGs and identify SG target genes, 2) characterize the detailed mechanism by which SGs mediate transcriptional changes, and 3) examine nuclear functions of G3BP1 including possible direct roles in regulating transcription. During the K99 phase, under the mentorship of Dr. Roy Parker and Dr. Robin Dowell, comprehensive approaches using genome-wide data analysis and cellular techniques will be used to determine if SGs regulate transcription, identify SG target genes, and elucidate the detailed mechanism of this regulatory pathway. With support from Dr. Amy Palmer, expertise in analyzing the role of SGs and/or G3BP1 in metal/ion homeostasis will be acquired in order to identify if SGs and/or G3BP1 regulate metal/ion homeostasis during stress response on the transcriptional level. Further, expertise from Dr. John Rinn in studying the role of RNA in chromatin recognition will advance characterizing G3BP1 and the importance of RNA in G3BP1-chromatin binding. Additional K99 training in genomic techniques and computational analysis will be essential for the proposed research and advance my transition into and progress during the independent R00 phase. The results of this proposal will not only lead to identification of important SG and G3BP1 function, but also provide a fundamental understanding of how RNP granules couple cytosolic and nuclear stress responses.