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Collaborative Research: Integrating Theory and Experiment to Unravel Protein Transport in the ER

合作研究：结合理论和实验来解开内质网中的蛋白质转运

基本信息

批准号：
2034486
负责人：
Laura Westrate
金额：
$ 35.11万
依托单位：
Calvin University
依托单位国家：
美国
项目类别：
Standard Grant
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-15 至 2024-12-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=2034486&HistoricalAwards=false
关键词：
Collaborative Research Integrating Theory Experiment

项目摘要

The endoplasmic reticulum, or ER, is a specialized intracellular structure responsible for a myriad of critical cellular functions, including protein synthesis, quality-control, and export. It is estimated that about one third of all mammalian proteins are created and then exported from the ER as the first step on their way to being secreted from the cell to become the signal carriers and receptors that interface with the extracellular world. The ER itself forms an interconnected network of sheets and hollow tubules that hosts a variety of protein processing pathways. This project aims to address a fundamental question in cell biology -- how does the structural complexity of the ER regulate and support its functional role as the protein processing hub of the cell? An interdisciplinary approach combines imaging within live cells, predictive physical modeling, and state-of-the-art simulation and data-analysis techniques to explore how proteins navigate through the ER maze. This collaborative effort seeks to unravel how the unique physical structure of the ER regulates its critical biological functions of protein sorting and export. The Broader Impacts of this work includes the intrinsic merit of the research as all eukaryotic cells have ER and dysfunctions of this organelles have been implicated in such human afflictions as diabetes and Parkinson’s disease. Additionally, the work will be integrated with a broad interdisciplinary educational program that spans from an elementary school science club, to a high school robotics team, to undergraduate research interns, aiming to introduce students at all educational levels to the insights that may be gained by applying physical and mathematical approaches to biological problems.The endoplasmic reticulum forms a continuous polymorphic network of stacked sheets and hollow tubules that spans throughout the intracellular space and is responsible for the folding, quality-control, sorting, and export of secreted proteins. The primary goal of this project is to establish a structure-function relationship for this organelle, developing a quantitative understanding of how its unique morphology supports its role as a protein delivery network. This goal will be approached collaboratively from the dual perspectives of physical model-building and dynamic live-cell imaging. A mathematical framework will be developed that combines analytical results for reaction-diffusion processes in complex geometries with novel simulation techniques to establish how network morphology modulates the kinetics of proteins finding binding partners, sorting regions, and exit sites within the network. Rapid confocal imaging of ER structure and photoactivated luminal and membrane proteins will enable quantification of the underlying dynamics of network structures (including tubule junctions and exit sites) and the embedded proteins. Genetic and pharmacological perturbations will be leveraged to test predictions of the effect of structure on intra-ER protein transport. The efficiency of protein sorting and export from the ER will be quantified using a synchronized accumulation-and-release system, and the contribution of physical factors such as bulk luminal flow and exit site distribution will be explored through a confluence of theoretical models and live-cell measurements. Ultimately, a feedback loop between theory and experiment will elucidate the underlying principles that link the complex architecture of the ER with its crucial protein processing functions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

内质网，或ER，是一种专门的细胞内结构，负责无数的关键细胞功能，包括蛋白质合成，质量控制和输出。据估计，大约三分之一的哺乳动物蛋白质被产生，然后从ER输出，作为从细胞分泌的第一步，成为与细胞外世界接触的信号载体和受体。内质网本身形成了一个相互连接的网络片和中空的小管，主机的各种蛋白质加工途径。该项目旨在解决细胞生物学中的一个基本问题-ER的结构复杂性如何调节和支持其作为细胞蛋白质加工中心的功能作用？一种跨学科的方法结合了活细胞内的成像，预测性物理建模以及最先进的模拟和数据分析技术，以探索蛋白质如何通过ER迷宫。这项合作努力旨在揭示ER的独特物理结构如何调节其蛋白质分选和输出的关键生物学功能。这项工作的更广泛影响包括研究的内在价值，因为所有真核细胞都有ER，并且这种细胞器的功能障碍与糖尿病和帕金森病等人类疾病有关。此外，这项工作将与广泛的跨学科教育计划相结合，从小学科学俱乐部到高中机器人团队，再到本科研究实习生，旨在向所有教育水平的学生介绍通过应用物理和数学方法解决生物学问题可能获得的见解。内质网形成一个连续的多形态网络，由堆叠的片状和中空的网状结构组成。横跨整个细胞内空间的小管，负责分泌蛋白的折叠、质量控制、分选和输出。该项目的主要目标是建立这种细胞器的结构-功能关系，定量了解其独特的形态如何支持其作为蛋白质递送网络的作用。这一目标将从物理模型建立和动态活细胞成像的双重角度进行合作。将开发一个数学框架，结合分析结果的反应扩散过程中复杂的几何形状与新的模拟技术，以建立网络形态如何调节动力学的蛋白质找到结合伙伴，排序区域，并退出网站内的网络。ER结构和光活化的管腔和膜蛋白的快速共聚焦成像将使网络结构（包括小管连接和出口位点）和嵌入的蛋白质的潜在动力学的定量成为可能。将利用遗传和药理学扰动来测试结构对ER内蛋白转运的影响的预测。蛋白质分选和出口从ER的效率将使用一个同步的积累和释放系统进行量化，并通过理论模型和活细胞测量的汇合探讨物理因素，如批量管腔流量和出口部位分布的贡献。最终，理论和实验之间的反馈回路将阐明将ER的复杂结构与其关键的蛋白质加工功能联系起来的基本原理。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估来支持。