IIBR RoL: Applying innovative structural tools to highlight RNA's structural dynamics as RNA-protein complexes self-assemble

IIBR RoL：应用创新的结构工具来突出 RNA-蛋白质复合物自组装时的结构动力学

• 批准号: 1930046
• 负责人: Lois Pollack
• 金额: $ 77.32万
• 依托单位: Cornell University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1930046&HistoricalAwards=false
• 关键词: IIBR; RoL; Applying; innovative; structural

Life functions are supported by the interaction of large biological molecules, including proteins and RNA. Although the shapes or structures of some of these complexes have been visualized, very little is known about their dynamics of assembly. How do these differing components interact to create complexes whose function exceeds that of either alone? This is a challenging problem because of the intertwined motions of the components. Through this project, new tools will be applied to highlight the motions of just one macromolecular species during the assembly of a complex. Although both proteins and RNA are present, only the RNA is visible. Students will design, build and apply the new technology to study the self-assembly of a small plant virus, an intricate structure consisting of a protein shell surrounding a nucleic acid core. These studies will allow us to more clearly define the rules that govern the joining of nucleic acids and proteins into biological complexes. Many important biological assembles are comprised of distinct macromolecular components, such as proteins in conjunction with the nucleic acids. Despite their importance, less than 2% of the static structures in the protein data bank reflect RNA-protein complexes. Even less is known about the dynamic interactions between RNA and protein components. Although much effort has been expended in watching one type macromolecule self-assemble, e.g. protein folding, it is much more difficult to interpret dynamic structural information for multicomponent machines because multiple species are present. The inability to separate the signal from the distinct components precludes a simple interpretation. This project will exploit and innovate existing experimental infrastructure to create tools that highlight the signal of only one species on the background of the other, vastly simplifying measurements of complexes' dynamics. Mixers that rapidly combine the protein and RNA precursors will be constructed and will allow experimenters to track the changing structures of only the RNA component during complex assembly. A small plant virus will serve as a model system for these studies. The development and application of two complementary techniques (small angle x-ray scattering and single molecule fluorescence) will offer a unique perspective on this self-assembly problem. Once established, these methods can be applied other RNA-protein complexes, gathering data to elucidate the rules governing their assembly. Project results, sorted by topic, will be posted at: https://pollack.research.engineering.cornell.edu/all_publications/. This project is jointly funded by the Division of Biological Infrastructure program for Infrastructure Innovation for Biological Research, the Molecular Biophysics and Genetic Mechanism Clusters in the Division of Molecular and Cellular Biosciences, and the Rules of Life initiative in the Emerging Frontiers office of the Biology Directorate.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.

生命功能是由大的生物分子，包括蛋白质和RNA的相互作用支持的。 虽然其中一些复合物的形状或结构已经可视化，但对其组装动力学知之甚少。 这些不同的成分是如何相互作用，创造出功能超过其中任何一种的复合物的？这是一个具有挑战性的问题，因为组件的相互交织的运动。通过这个项目，新的工具将被应用于突出一个大分子物种在组装复合物过程中的运动。 虽然蛋白质和RNA都存在，但只有RNA可见。 学生将设计，构建和应用新技术来研究小型植物病毒的自组装，这是一种复杂的结构，由围绕核酸核心的蛋白质外壳组成。这些研究将使我们能够更清楚地定义核酸和蛋白质连接成生物复合物的规则。许多重要的生物集合体由不同的大分子成分组成，如蛋白质与核酸的结合。尽管它们很重要，但在蛋白质数据库中，只有不到2%的静态结构反映了RNA-蛋白质复合物。关于RNA和蛋白质成分之间的动态相互作用，我们所知更少。虽然已经花费了大量的努力来观察一种类型的大分子自组装，例如蛋白质折叠，但由于存在多个物种，因此解释多组分机器的动态结构信息要困难得多。 由于无法将信号从不同的成分中分离出来，因此无法进行简单的解释。 该项目将利用和创新现有的实验基础设施，以创建工具，突出一个物种的信号在另一个物种的背景下，大大简化了复合物动态的测量。 可以快速联合收割机将蛋白质和RNA前体结合起来，并允许实验者在复杂的组装过程中只跟踪RNA成分的结构变化。 一种小型植物病毒将作为这些研究的模型系统。两种互补技术（小角X射线散射和单分子荧光）的发展和应用将为这种自组装问题提供独特的视角。一旦建立，这些方法可以应用于其他RNA-蛋白质复合物，收集数据以阐明控制其组装的规则。 项目结果，按主题排序，将张贴在：www.example.com。该项目由生物基础设施部门生物研究基础设施创新计划、分子和细胞生物科学部门的分子生物物理学和遗传机制集群联合资助，以及生物学理事会新兴前沿办公室的生命规则倡议。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准。

项目成果

RNA structures and dynamics with Å resolution revealed by x-ray free-electron lasers.

• DOI: 10.1126/sciadv.adj3509
• 发表时间: 2023-09-29
• 期刊: SCIENCE ADVANCES
• 影响因子: 13.6
• 作者: Zielinski, Kara A.;Sui, Shuo;Pabit, Suzette A.;Rivera, Daniel A.;Wang, Tong;Hu, Qingyue;Kashipathy, Maithri M.;Lisova, Stella;Schaffer, Chris B.;Mariani, Valerio;Hunter, Mark S.;Kupitz, Christopher;Moss III, Frank R.;Poitevin, Frederic P.;Grant, Thomas D.;Pollack, Lois
• 通讯作者: Pollack, Lois

Visualizing a viral genome with contrast variation small angle X-ray scattering

通过小角度 X 射线散射对比变化可视化病毒基因组

• DOI: 10.1074/jbc.ra120.013961
• 发表时间: 2020
• 期刊: Journal of Biological Chemistry
• 影响因子: 4.8
• 作者: San Emeterio, Josue;Pollack, Lois
• 通讯作者: Pollack, Lois