Cellular stress sensing via biomolecular condensation

项目摘要 背景：细胞承受广泛的环境压力，必须能够感知和响应 为了生存而做出的改变。感知过程的发生取决于压力本身，我们有 在酿酒酵母中鉴定出150多种热敏蛋白，它们在发酵后表现出生物分子缩合 温度升高。在这一组中，一组保守的GTP酶在没有 对任何其他细胞成分的需求，打开了这些蛋白质有能力 才能感觉到热休克。此外，这组蛋白质与两种基本的功能反应有关 在热应激下发生--热休克基因转录上调和核糖体生物合成关闭。 对热高度敏感的蛋白质可能作为大量功能的上游长期未知的传感器 细胞内的变化，并作为这一提议的热感应候选。 长期以来一直被视为恶劣环境条件的有毒后果，最近的研究表明 生物分子凝析油的形成具有非随机性、适应性和可逆性。有了这种新兴的观点，疾病 如痴呆症和肌萎缩侧索硬化症，与非膜结合蛋白聚集体的积累有关 可能是压力感应通路异常激活的结果，表明我们对压力感知通路的理解 疾病病理可能需要重新评估。 具体目标：1：候选者是否足以在体内诱导转录反应？2：什么是 热敏机制？3：候选缩合在核糖体上的功能相关性是什么 制作？ 研究设计：我将利用一种嗜冷酵母，它在 温度低于酿酒酵母。嗜冷酵母可能含有同源的感受器蛋白 在较低温度下提高敏感度。我将内源传感器候选基因替换为它们的 重组酿酒酵母的嗜冷性同源物及其上调产热能力的检测 特定的转录本和减弱的核糖体生物发生。我将重新构建这些功能组件 并测试候选传感器的冷凝是否会影响任一种反应。 候选传感器的缩合也将在体外使用生化和生物物理分析进行研究，以 研究它们感知热量的内在能力，以描述感知的机械基础。 培训：这项研究将与芝加哥大学的D.Allan Drummond博士一起进行，并将建立 根据我的实验技能，首先描述了生物分子缩合的功能相关性 环境压力传感，并进一步扩展到理解生物物理机制。这次培训 将为我未来的职业生涯做好准备，研究环境压力如何塑造细胞行为。

Project Summary Background: Cells experience a wide array of environmental stresses, and must be able to sense and respond to changes in order to survive. The sensing process which occurs depends on the stress itself, and we have identified over 150 heat-sensitive proteins in S. cerevisiae which exhibit biomolecular condensation after temperature increase. Within this group, a conserved set of GTPases displays significant aggregation without the requirement of any other cellular components, opening up the possibility that these proteins have the ability to sense heat shock. Further, this set of proteins is connected to two fundamental functional responses which occur under heat stress -- transcriptional upregulation of heat shock genes and shutoff of ribosome biogenesis. Proteins that are highly sensitive to heat may act as long-unidentified sensors upstream of massive functional changes within the cell, and serve as heat sensing candidates for this proposal. Long viewed as a toxic consequence of harsh environmental conditions, recent work has shown that biomolecular condensate formation is non-random, adaptive, and reversible. With this emerging view, diseases like dementia and ALS which are associated with the accumulation of non-membrane bound protein aggregates might be the result of an aberrant activation of stress sensing pathways, indicating that our understanding of the disease pathology may need to be reevaluated. Specific Aims: 1: Are candidates sufficient to induce the transcriptional response in vivo? 2: What is the mechanism for heat sensing? 3: What is the functional relevance of candidate condensation on ribosome production? Study Design: I will take advantage of a cryophilic yeast which execute their heat-induced cellular responses at lower temperatures than S. cerevisiae. The cryophilic yeast likely contain homologous sensor proteins with increased sensitivity at lower temperatures. I will replace the endogenous sensor candidate genes with their cryophilic homologs and assay the ability of the recombinant S. cerevisiae to upregulate the production of heat- specific transcripts and attenuate ribosome biogenesis. I will reconstitute the components of these functional responses in vitro and test whether the condensation of candidate sensors can affect either response. Condensation of candidate sensors will also be studied in vitro using biochemical and biophysical assays to investigate their intrinsic ability to sense heat to describe the mechanistic underpinnings of sensing. Training: This research will be performed with Dr. D. Allan Drummond at the University of Chicago and will build upon my experimental skills by first characterizing the functional relevance of biomolecular condensation in environmental stress sensing and further expanding into understanding the biophysical mechanism. This training will prepare me for a future career studying how environmental stresses shape cellular behavior.