Structure-Function of Nucleo-Cytoplasmic Communication

核-细胞质通讯的结构-功能

• 批准号: 10475615
• 负责人: Thomas Schwartz
• 金额: $ 37.99万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10475615
• 关键词: Address; Cardiac; Cell Nucleus; Cell physiology; Cells; Communication; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Cytoskeleton; Drug Design; Elements; Emery-Dreifuss Muscular Dystrophy; Eukaryotic Cell; Functional disorder; Genetic Materials; Genetic Transcription; Goals; Human; Individual; Malignant Neoplasms; Maps; Membrane; Methods; Molecular; Muscular dystrophy cardiomyopathy; Myopathy; Nuclear Envelope; Nuclear Pore Complex; Organelles; Positioning Attribute; Premature aging syndrome; Primary Dystonias; Process; Proteins; Resolution; SWP29; Structure; Technology; Translations; X-Ray Crystallography; grasp; human disease; innovation; mechanotransduction; protein complex; skeletal; success

PROJECT SUMMARY / ABSTRACT Eukaryotic cells are defined by their organelles, membrane-enclosed compartments in which specific cellular processes are carried out. The nucleus is the largest organelle, contains all genetic material, and enables separation of gene transcription from protein translation. As the nuclear envelope (NE) serves as a tight barrier enclosing the nucleus, the cell requires machinery to establish and control nucleo-cytoplasmic communication. There are two principally different components to this machinery. On one hand, nuclear pore complexes (NPCs) serve as the main conduit for molecular exchange across the NE. On the other hand, universally conserved linker of nucleo- and cytoskeleton (LINC) complexes serve as physical tethers across the NE, which are necessary for positioning the nucleus and for mechano-sensing in a diverse set of circumstances. Dysfunction of the machinery is at the core of important human diseases, including skeletal and cardiac myopathies, premature aging, and cancer. Our goal is to understand the structure of the protein complexes involved in nucleo-cytoplasmic communication at high (atomic) resolution. Such information helps to identify and separate the myriad functions this machinery carries out and that we are still only beginning to fully grasp. High resolution information further provides the basis for structure-guided drug design to interfere with the salient human diseases, such as Emery-Dreifuss Muscular Dystrophy (EDMD) and Primary Dystonia, which are still not cured. The structural characterization of the NPC and the LINC complex are challenging, because of the size and complexity of these multi-MDa assemblies. Over the past 15 years, we have made significant advances on both problems. For the NPC, we have chosen a highly productive bottom-up approach, in which we characterized multi-subunit complexes predominantly by X-ray crystallography, the building blocks of the massive, 40-100 MDa NPC. Those structures have now been used in combination with cryo-electron tomographic (cryo-ET) maps of assembled NPCs to generate composite structures that attempt to position the roughly 500 individual proteins within one NPC. For the LINC complex, we solved the universally conserved core component and have started to untangle the diverse network of its components, the Sad1/UNC-84 (SUN) and Klarsicht/ANC1/Syne-Homology (KASH) proteins. Going forward, the challenge is the structural characterization of large and dynamic assemblies, which is true for both, the NPC and the LINC complex, for the latter particularly when including the connection to the nucleo- and cytoskeletal components. The dramatic advances in cryo-electron microscopy (cryo-EM) over the recent past make this technology particularly important for our studies. We anticipate combining X-ray crystallography and cryo-EM for studying the most relevant structures going forward. The success of this will depend upon innovative, tailored methods to address the particular challenges that come with each project. We have repeatedly shown over the past decade how to successfully approach such challenges and have devised methods to meet them.

项目摘要 /摘要 真核细胞由它们的细胞器，膜封闭的室定义，其中特定的细胞在其中 进行过程。核是最大的细胞器，包含所有遗传物质，并可以 基因转录与蛋白质翻译的分离。由于核信封（NE）充当紧密的障碍 封闭细胞核，该细胞需要机械才能建立和控制核总质质通信。 该机械有两个主要不同的组件。一方面，核孔复合物 （NPC）是整个NE的分子交换的主要管道。另一方面，普遍 核和细胞骨架（LINC）配合物的保守接头是整个NE的物理系数， 对于在各种情况下定位核和机械感应是必要的。 机械功能障碍是重要人类疾病的核心，包括骨骼和心脏 肌病，过早衰老和癌症。我们的目标是了解蛋白质复合物的结构 参与高（原子）分辨率的核总质通信。这样的信息有助于识别 并分开该机械执行的无数功能，并且我们仍开始完全掌握。 高分辨率信息进一步为结构引导的药物设计提供了基础 显着的人类疾病，例如emery-dreifuss肌肉营养不良（EDMD）和原发性肌张力障碍，这些疾病 仍未治愈。 NPC和LINC综合体的结构表征具有挑战性，因为 这些多MDA组件的大小和复杂性。在过去的15年中，我们变得很重要 这两个问题的进展。对于NPC，我们选择了一种高效的自下而上的方法，其中 我们以X射线晶体学为主要的多生产络合物表征了多支化合物络合物。 巨大，40-100 MDA NPC。这些结构现已与冷冻电子结合使用 组装NPC的层析成像（冷冻-ET）地图，以生成试图定位的复合结构 一个NPC内约有500种单独的蛋白质。对于林克建筑群，我们解决了普遍保守的 核心组件，并开始解开其组件的各种网络，SAD1/UNC-84（SUN） 和klarsicht/anc1/syne-hosology（KASH）蛋白质。展望未来，挑战是结构性 大型和动态组件的表征，对于NPC和LINC复合物都是正确的 后者特别是在包括与核和细胞骨架成分的连接时。戏剧性 最近的冷冻电子显微镜（Cryo-EM）的进步在最近的过去使得这项技术特别使得 对我们的研究很重要。我们预计将X射线晶体学和冷冻EM结合在一起，以研究最多 相关结构将前进。这一成功将取决于创新的，量身定制的方法 解决每个项目带来的特定挑战。我们在过去反复展示 十年如何成功应对此类挑战并设计了满足它们的方法。