Heterogenous cytoplasmic RNA-protein aggregates enhance cellular survival under stress

异质细胞质 RNA 蛋白聚集体增强细胞在应激下的存活率

• 批准号: 9910869
• 负责人: Srivats Venkataramanan
• 金额: $ 6.53万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-02 至 2022-01-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9910869
• 关键词: Address; Area; Biological; Biological Assay; Brain; Cancerous; Cell Line; Cell Survival; Cells; Cellular Stress; Cytoplasmic Granules; Cytoplasmic Organelle; Data; Dependence; Dimensions; Environment; Flow Cytometry; G3BP1 gene; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Goals; Heat shock proteins; Heat-Shock Response; Heterogeneity; High-Throughput Nucleotide Sequencing; Homo; Hypoxia; Knowledge; Left; Measures; Messenger RNA; Molecular; Molecular Biology; Monitor; Mus; Neoplasm Metastasis; Normal Cell; Oxidative Stress; Pathology; Phase; Physiological; Polyribosomes; Population; Principal Investigator; Protein Isoforms; Proteins; RNA; RNA-Binding Proteins; Recovery; Regimen; Regulation; Research; Resistance; Ribosomes; Role; Specific qualifier value; Specificity; Starvation; Stress; Technology; Testing; Therapeutic; Tissues; Transcript; Translational Regulation; Translations; Triage; Variant; Virus Diseases; Work; anti-cancer; biological adaptation to stress; cancer cell; fitness; interdisciplinary approach; optogenetics; programs; protein aggregation; ras GTPase-Activating Proteins; response; stress granule; stressor; tool; tumor

Project Summary/Abstract Plasticity of gene expression programs allows cells to respond and adapt to stress. While adaptation involves both transcriptional and post-transcriptional changes, translational regulation is particularly crucial in situations that demand a rapid response. Cells respond to stress such as hypoxia, starvation, oxidative stress, heat shock or viral infections by shutting down translation and sequestering mRNA into stress granules; dynamic, non-membrane bound, phase-separated cytoplasmic organelles. The ability of cells to form stress granules is particularly important in the ability of cancer cells to survive chemotherapeutic treatments. However, stress granules are not monotypic. G3BP, a core nucleator protein of stress granules, has at least three known variants - G3BP1, G3BP2a and G3BP2b. G3BP variants can form homo or hetero-multimers in the context of stress granules, and have also been shown to have different tissue specificities. However, the functional consequences of this variation amongst granule nucleating proteins has not been studied. The absence of this knowledge renders our understanding about the cellular response to various insults incomplete. G3BP variants also have differential granule-forming capacities, indicating more than one dimension of heterogeneity amongst cellular stress granules. Further, our data suggests that G3BP variants are unable to compensate for each other functionally, indicating divergence in the stress response. Therefore, we hypothesize that the heterogeneity in stress granule composition is important for the cell to be able to respond and adapt to diverse stresses, and that granules of different compositions have different effects on the mRNA fates in response to stress. We will address this hypothesis via the following specific aims: (1) Identify role of stress granule heterogeneity in cellular resistance to exogenous stresses: We will subject cells that can produce only a limited number of or only a single granule subtype(s) to a wide panel of stressors and monitor absolute and relative fitness using a sensitive flow cytometry assay and (2) Elucidate the mechanism(s) of granule-dependent differential fitness in response to stressors: Using a combination of molecular biology and high-throughput sequencing technologies, we will measure localization of transcripts to granules nucleated by specific G3BP variants. A recently developed optogenetic tool will allow us to induce granule formation in the absence of stress, thereby independently measuring the effect of granules on cytoplasmic translation. My goal is to become a principal investigator pursuing an independent research program focused on deciphering cellular responses to stress. UCSF is an ideal environment to pursue an interdisciplinary approach to answering these fundamental biological questions. Completion of the proposed work will expand our knowledge of the mechanisms underlying the cellular response to stress, informing on responses to pathologies and therapeutic regimens.

项目概要/摘要 基因表达程序的可塑性使细胞能够对压力做出反应和适应。适应的同时 涉及转录和转录后变化，翻译调控尤其重要 需要快速响应的情况。细胞对缺氧、饥饿、氧化应激等应激做出反应， 通过关闭翻译并将 mRNA 隔离到应激颗粒中来引起热休克或病毒感染； 动态的、非膜结合的、相分离的细胞质细胞器。细胞形成压力的能力 颗粒对于癌细胞在化疗中存活的能力特别重要。 然而，应力粒并不是单一型的。 G3BP 是应激颗粒的核心成核蛋白，至少具有 三个已知变体 - G3BP1、G3BP2a 和 G3BP2b。 G3BP变体可以形成同源或异源多聚体 应激颗粒的背景，并且还被证明具有不同的组织特异性。然而， 颗粒成核蛋白之间这种差异的功能后果尚未研究。这 缺乏这方面的知识使我们无法理解细胞对各种损伤的反应 不完整。 G3BP 变体也具有不同的颗粒形成能力，表明不止一种 细胞应激颗粒之间的异质性维度。此外，我们的数据表明 G3BP 变体 无法在功能上相互补偿，表明压力反应存在差异。所以， 我们假设应激颗粒组成的异质性对于细胞能够 不同成分的颗粒对不同的压力有不同的影响 mRNA 命运对压力的反应。我们将通过以下具体目标来解决这一假设：（1）确定 应激颗粒异质性在细胞对外源应激抵抗力中的作用：我们将研究能够 仅产生有限数量或仅产生单一颗粒亚型以适应广泛的压力源和监测 使用灵敏的流式细胞术测定绝对和相对适合度，以及（2）阐明 颗粒依赖性差异适应度对应激源的反应：结合分子生物学和 高通量测序技术，我们将测量转录本到成核颗粒的定位 特定的 G3BP 变体。最近开发的光遗传学工具将使我们能够诱导颗粒形成 没有压力，从而独立测量颗粒对细胞质翻译的影响。我的目标 是成为一名首席研究员，从事一项专注于破译的独立研究项目 细胞对压力的反应。加州大学旧金山分校 (UCSF) 是追求跨学科方法的理想环境 回答这些基本的生物学问题。完成拟议的工作将扩大我们的 了解细胞对压力反应的机制，了解对压力的反应 病理学和治疗方案。