Cell Organization Through Phase Separation: Mechanisms, Functions and Disease

生物分子凝聚物将选定的大分子组浓缩成真核细胞中离散的焦点 在没有周围膜的情况下。缩合物存在于真核细胞中，并且具有功能 从信号转导到RNA代谢再到基因表达的过程。异常凝聚物 与神经退行性疾病、癌症和皮肤病有关。我们对真核细胞的认识 许多冷凝物似乎是通过液-液形成的这一发现改变了组织结构 多价大分子的相分离（LLPS）。我的实验室在这一发现中发挥了重要作用 建立关键原则，包括多价相互作用在促进生物多样性方面的重要作用 LLPS、共价修饰对 LLPS 的调节以及 LLPS 增加酶促作用的能力 活动。近年来，我们展示了 LLPS 如何产生膜相关簇， 增加信号分子的比活性。我们还表明，本质上无序的区域 蛋白质（IDR）可以经过 LLPS 产生液滴，随着时间的推移，液滴会硬化为固体，这可能是由于 淀粉样蛋白丝的形成，并且失调的硬化可能会导致神经退行性变。 此外，我们提出了第一个模型来解释基于支架/客户端框架的冷凝物成分。 最近，我们发现染色质具有进行 LLPS 的内在倾向，这提供了一个新的观点 真核基因组组织。在这里，我们提出了一项广泛的计划来解决以下领域的主要问题： 生物分子凝聚场。我们将使用微流体技术在一次评估中评估数千个 IDR 的 LLPS 实验，通过以下方法建立了 LLPS 和淀粉样蛋白形成的序列决定因素的预测模型 印尼盾。这项工作将加深我们对这些过程的生物物理学理解，预测哪些 IDR 是可能的 通过自组装促进特定的生物过程（例如转录），并揭示 IDR 如何 突变通过淀粉样蛋白形成导致疾病。我们还将研究各个组成部分如何 酵母 P 体影响 LLPS 阈值和区室组成以及 RNA 解旋酶和 RNA 脱帽活性在其中受到调节。这项工作将带来一种新的凝结水模型 基于组件之间交互模式的组合，并将解释如何组合和 封装可以控制天然缩合物中的酶活性。最后，我们将学习如何 核小体间距和多种组蛋白翻译后修饰控制染色质 LLPS 生成生化和功能上不同的基因组区域，检查是否引起 NUT 癌 通过有缺陷的 LLPS，并开发一种基于靶向小分子的新药物设计方法 凝结物。这项工作将共同揭示生物相分离的新原理，解释相分离如何 分离可用于控制 RNA 代谢以及基因组组织和功能，并提供见解 研究神经退行性疾病和癌症的机制和潜在治疗方法。

Biomolecular condensates concentrate select groups of macromolecules into discrete foci in eukaryotic cells in the absence of a surrounding membrane. Condensates are found throughout eukaryotic cells, and function in processes ranging from signal transduction to RNA metabolism to gene expression. Aberrant condensates have been implicated in neurodegeneration, cancer and skin diseases. Our understanding of eukaryotic cell organization was transformed by the discovery that many condensates appear to form through liquid-liquid phase separation (LLPS) of multivalent macromolecules. My lab played an important part in this discovery by establishing key principles, including the essential role of multivalent interactions in promoting biological LLPS, the regulation of LLPS by covalent modifications, and the ability of LLPS to increase enzymatic activity. In recent years, we showed that and how LLPS can produce membrane-associated clusters that increase the specific activity of signaling molecules. We also showed that intrinsically disordered regions of proteins (IDRs) can undergo LLPS to produce liquid droplets that harden to solids over time, likely due to formation of amyloid filaments, and that misregulated hardening may contribute to neurodegeneration. Further, we proposed the first model to explain condensate composition based on a scaffold/client framework. Most recently, we showed that chromatin has an intrinsic propensity to undergo LLPS, providing a new view of eukaryotic genome organization. Here, we propose a broad program to address leading questions in the biomolecular condensate field. We will use microfluidics to assess LLPS of thousands of IDRs in a single experiment, leading to a predictive model for the sequence determinants of LLPS and amyloid formation by IDRs. This work will deepen our biophysical understanding of these processes, predict which IDRs are likely to contribute to specific biological processes (e.g. transcription) through self-assembly, and reveal how IDR mutations lead to disease through amyloid formation. We will also examine how individual components of yeast P bodies impact the LLPS threshold and composition of the compartments and how RNA helicase and RNA decapping activities are modulated within them. This work will lead to a new model of condensate composition based on the patterns of interaction between components, and will explain how composition and encapsulation can control enzymatic activities in native condensates. Finally, we will learn how internucleosome spacing and diverse histone post-translational modifications control chromatin LLPS to generate biochemically and functionally distinct genomic regions, examine whether NUT carcinoma is caused by defective LLPS, and develop a new approach to drug design based on targeting small molecules to condensates. Together, the work will reveal new principles of biological phase separation, explain how phase separation can be used to control RNA metabolism and genome organization and function, and provide insights into the mechanisms and potential treatments of neurodegeneration and cancer.