Sequence determinants of membraneless organelle rheology

项目概要/摘要 我的实验室的长期愿景是阐明生物学中尺度组装的生物物理原理， 利用这些发现促进生物医学和生物技术的进步。在这种情况下，实验室目前的重点是 阐明细胞内相分离的基本原理。相分离是一个基本的过程 细胞用来将无数的生物分子组织成功能区室， 细胞器如应激颗粒、胚芽颗粒和核仁，在基因调控中具有重要作用， 应激反应，以及许多额外的基本功能。相反，异常相变与 神经退行性疾病和其他疾病。这个MIRA提案的总体目标是建立如何 生物分子序列和组成决定了无膜细胞器的材料性质， 材料特性如何影响生物功能和功能失调。我们将仔细分析 以及决定生物分子的流变性（液体状、固体状或组合）的分子相互作用。 由内在无序蛋白质的相分离形成的缩合物。我们将使用 蛋白质工程，诱变，显微镜，一套生物物理技术和纳米技术，包括 纳米粒子探针工具箱的开发，用于测量无膜细胞器流变学。我们将 然后研究无膜细胞器流变学如何成为生物功能的基础，重点是胚芽颗粒， 我们将研究无膜细胞器流变学的扰动如何扰乱生物功能。 最后，我们将检查某些序列特征是否特别容易出现异常的液-固相 转换和毒性，表明可能的病理学起源和治疗目标。在一起，这项工作 旨在为理解分子序列、流变学和 无膜细胞器在健康和疾病中的功能。

PROJECT SUMMARY/ABSTRACT The long-term vision of my lab is to elucidate biophysical principles of mesoscale assembly in biology and to harness those discoveries for biomedical and biotechnological advances. In that vein, the lab’s current focus is on elucidating principles underlying intracellular phase separation. Phase separation is a fundamental process that cells use to organize their myriad biomolecules into functional compartments, giving rise to membraneless organelles such as stress granules, germ granules, and the nucleolus, with important roles in gene regulation, stress response, and many additional essential functions. Conversely, aberrant phase transitions are associated with neurodegenerative and other diseases. The overall goal of this MIRA proposal is to establish how biomolecular sequence and composition determine the material properties of membraneless organelles, and how material properties contribute to biological function and misfunction. We will dissect the sequence features and molecular interactions that determine the rheology (liquid-like, solid-like, or a combination) of biomolecular condensates formed by phase separation of intrinsically disordered proteins. We will accomplish this using protein engineering, mutagenesis, microscopy, a suite of biophysical techniques, and nanotechnology, including the development of a toolbox of nanoparticle probes for measuring membraneless organelle rheology. We will then investigate how membraneless organelle rheology underlies biological function, focusing on germ granules, and we will investigate how perturbations to membraneless organelle rheology perturb biological function. Finally, we will examine whether certain sequence features are particularly prone to aberrant liquid-solid phase transitions and toxicity, indicating possible origins of pathology and targets for therapies. Together, this work seeks to provide a strong foundation for understanding the links between molecular sequence, rheology, and function of membraneless organelles in health and disease.