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

项目摘要 核糖核蛋白（RNP）颗粒是由RNA和RNA结合而成的无细胞膜细胞器。 RNA结合蛋白（RBP），通过缩合或液-液的生物物理原理形成 相分离（LLPS）。一组有趣的胞质RNP颗粒是应激颗粒（SG）， 形成于各种不同的压力源。压力导致了一种全局性的翻译抑制， 非翻译RNA浓缩并聚集成SG。各种SG蛋白的突变导致异常的 和组成型SG或RNP聚集体。这种RNP聚集体与肌萎缩侧索硬化相关。 硬化症（ALS）、额颞叶痴呆（FTD）和各种肌营养不良症。因此， 医学需要了解RNP颗粒组装和拆卸的原理。 虽然对SG的性质、机制和功能进行了深入研究，但对SG的性质、机制和功能知之甚少。 SGs是否以及如何耦合转录和转录后机制。因此，我的研究 目的集中在表征SG和/或SG组装支架蛋白G3 BP 1如何调节 转录，从而偶联胞质和核应激反应以维持细胞内稳态。在 这个应用程序，我结合联合收割机全基因组，计算和实验的方法，以1）确定是否 转录受SG形成的影响，并确定SG靶基因，2）详细描述 SGs介导转录变化的机制，以及3）检查G3 BP 1的核功能 包括在调节转录中可能的直接作用。 在K99阶段，在Roy帕克博士和Robin Dowell博士的指导下， 使用全基因组数据分析和细胞技术的方法将用于确定是否存在SGs 调控转录，鉴定SG靶基因，并阐明这种调控的详细机制 通路在Amy Palmer博士的支持下，分析SG和/或G3 BP 1在 将获得金属/离子稳态，以确定SGs和/或G3 BP 1是否调节金属/离子 在转录水平上的应激反应过程中的稳态。此外，约翰·里恩博士的专业知识， 研究RNA在染色质识别中的作用将有助于进一步表征G3 BP 1， G3 BP 1-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培训对于 建议研究和推进我的过渡和独立R 00阶段的进展。 该提议的结果不仅将导致重要SG和G3 BP 1功能的鉴定， 而且还提供了RNP颗粒如何耦合胞质和核应激的基本理解 应答

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.