Molecular mechanisms of coat assembly and regulation in membrane trafficking pathways

膜运输途径中外壳组装和调节的分子机制

• 批准号: 10619604
• 负责人: Lauren Parker Jackson
• 金额: $ 40.81万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10619604
• 关键词: Affinity; Alzheimer' s Disease; Binding; Biochemical; Capsid Proteins; Cell Line; Cellular Membrane; Cellular biology; Clathrin; Coat Protein Complex I; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Data; Defect; Destinations; Ensure; Event; Experimental Designs; Family; Goals; Health; Human; Integral Membrane Protein; Link; Lipids; Malignant Neoplasms; Membrane; Membrane Lipids; Methods; Molecular; Molecular Structure; Movement; Parkinson Disease; Pathway interactions; Phenotype; Physiological Processes; Proteins; Regulation; SNAP receptor; Spastic Paraplegia; Structural Models; Testing; Tubular formation; Vesicle; Work; X-Ray Crystallography; biophysical techniques; genetic regulatory protein; human disease; insight; model organism; nervous system disorder; polypeptide; protein complex; protein protein interaction; protein transport; tool; trafficking

Project Summary The timely delivery of membrane-bound vesicles and tubules bearing transmembrane protein and lipid cargoes to discrete cellular membranes is fundamental to cell biology and human health. Many proteins associated with trafficking pathways are linked to serious and crippling human diseases, especially neurological diseases and disorders. Although many trafficking proteins and some pathways are well characterized, we still do not understand other trafficking pathways that we infer must exist between membranes. This constitutes an enormous gap in our understanding of fundamental cell biology. Our goal is to elucidate the molecular structures and functions of important coat protein complexes that initiate trafficking pathways by forming coats around vesicles or tubules. Coat proteins recognize and package relevant cargoes, and they promote efficient assembly of additional required protein components, like accessory proteins and SNAREs. Clathrin coats have long served as an important paradigm, but increasing evidence demonstrates how other coats use distinct mechanisms. We investigate the retromer and Assembly Polypeptide (AP) family of coat complexes (COPI, AP-4, AP-5) by using a variety of tools to ascertain molecular mechanisms of coat assembly and regulation. Biochemical approaches allow us to identify and test new interactions in coat complexes, including how accessory and regulatory proteins drive function. Integrated structural methods including X-ray crystallography, cryo-electron microscopy (cryo-EM), and cryo-electron tomography (cryo-ET) provide detailed evidence for how coats interact with key partners and allow us to generate specific hypotheses to test function. Biophysical techniques enable us to quantify binding affinities and to probe interfaces identified in structural models. With collaborators, we use molecular data to design experiments in cultured cell lines or in model organisms to explore how protein-protein interactions drive phenotypes at the cellular or organismal levels. Ultimately, we hope to gain a molecular understanding of how coats assemble at distinct membranes to drive different trafficking pathways. We anticipate this work will reveal new mechanisms of coat assembly and regulation and will provide fundamental insights into the protein networks that underlie key cellular events on membranes.

项目摘要 带有跨膜蛋白和脂质货物的膜结合囊泡和小管的及时输送 离散的细胞膜是细胞生物学和人类健康的基础。许多蛋白质与 贩运途径与严重和严重的人类疾病有关，尤其是神经系统疾病和 疾病。尽管许多贩运蛋白和一些途径的特征都很好，但我们仍然没有 了解我们推断的其他贩运途径必须存在于膜之间。这是一个 我们对基本细胞生物学的理解巨大差距。我们的目标是阐明分子 重要外套蛋白质复合物的结构和功能，这些复合物通过形成大衣启动运输途径的结构和功能 囊泡或小管周围。涂上蛋白质识别并包装相关的货物，并提高高效 组装额外的蛋白质成分，例如辅助蛋白和贪食剂。网状蛋白外套有 长期以来一直是一个重要的范式，但越来越多的证据表明了其他外套如何使用不同 机制。我们研究了外套复合物（Copi，copi， AP-4，AP-5）使用多种工具来确定外套组装和调节的分子机制。 生化方法使我们能够识别和测试外套复合物中的新互动，包括如何 附件和调节蛋白驱动功能。综合结构方法，包括X射线晶体学， 冷冻电子显微镜（Cryo-EM）和冷冻电子层析成像（Cryo-ET）提供了详细的证据 大衣如何与关键伙伴相互作用，并允许我们生成特定的假设以测试功能。生物物理 技术使我们能够量化结合亲和力和探测结构模型中识别的界面。和 合作者，我们使用分子数据在培养的细胞系或模型生物中设计实验 探索蛋白质 - 蛋白质相互作用如何在细胞或生物水平上驱动表型。最终，我们 希望对外套如何在不同的膜上组装以驱动不同的分子了解不同 贩运途径。我们预计这项工作将揭示外套组装和调节的新机制，并且 将提供对膜上关键细胞事件的蛋白质网络的基本见解。