Collaborative Research: Developing a multi-scale understanding of microtubule dynamic instability

合作研究：发展对微管动态不稳定性的多尺度理解

• 批准号: 1817966
• 负责人: Holly Goodson
• 金额: $ 98.77万
• 依托单位: University of Notre Dame
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-15 至 2023-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1817966&HistoricalAwards=false
• 关键词: Collaborative; Research; Developing; multi; scale

Most cells in a wide range of organisms contain a dynamic substructure called the cytoskeleton, a network of protein-based polymers and associated proteins that has fundamental roles in cell movement, DNA partitioning, and internal cell organization. A key aspect of many cytoskeletal polymers is that they require chemical energy in the form of ATP (or GTP) to maintain a polymerized state. The harnessing of this energy allows the cytoskeletal filaments to do work, respond dynamically to internal and external signals, and self-organize. The major goal of the work in this project is to use a combination of experiments and computational modeling to develop an improved theoretical framework for understanding and predicting the behaviors of these dynamic cytoskeletal polymers as observed at different scales. More specifically, the proposed work sets out to establish how the biochemical properties of the polymer subunits (including the rate at which they burn ATP or GTP) relate to the behaviors of the individual filaments and to the overall behaviors of populations of filaments. In addition, the project studies how filament binding proteins work together to regulate filament dynamics. While this work is basic science, it has the potential to have practical applications in nanotechnology and synthetic biology. Through this project, graduate students and undergraduates will receive interdisciplinary training in both computational modeling and experimental biology. High school teachers and students will also be engaged in the research process. Freely available, open-source software and tutorials produced through this project will help students and researchers at all levels gain an intuitive understanding of dynamic polymer systems.From a technical perspective, the project has four specific goals, most of which focus on a type of cytoskeletal filaments known as microtubules. Individual microtubules exhibit a dramatic behavior known as dynamic instability, in which they stochastically alternate between extended periods of growth and depolymerization. (1) The first project goal is to develop and test hypotheses for the mechanisms of the transitions in microtubule dynamic instability by relating the behaviors of the filaments to the subunit-level structure of their tips. The approach will utilize a combination of work with a previously established detailed computational model of microtubule dynamics, a novel data analysis tool for identifying and statistically categorizing the microtubule behaviors, and experimental data acquired at high temporal and spatial resolutions. (2) The second goal is to establish a predictive understanding of the relationships between the biochemical characteristics of the subunits (kinetic rate constants), the behaviors of the filaments (e.g., dynamic instability, treadmilling) and the attributes of the polymer systems (e.g., critical concentrations, steady states). The approach will utilize a combination of computational modeling (performed with variants of the model used in Goal 1) and experiments with a bacterial relative of tubulin called PhuZ (chosen because wildtype and altered versions of this protein can be expressed in bacteria and characterized in vitro). (3) The third goal is to use a combination of experiments and computational models to test a set of hypotheses for how a group of filament binding proteins known as +TIPs (microtubule plus-end tracking proteins) work together to regulate microtubule behavior. (4) The final goal is to create for broad distribution packages of our software and associated analysis tools used in Goals 1 to 3. These packages will include software targeted at both the research and teaching communities. While the focus of our studies is on microtubules, the resulting multi-scale understanding of polymerizing filament systems should apply to steady-state (energy-utilizing) polymers more generally, including actin, bacterial filaments, and polymers created through biotechnology.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.

多种生物体中的大多数细胞都含有称为细胞骨架的动态子结构，这是一种基于蛋白质的聚合物和相关蛋白质的网络，在细胞运动、DNA 分区和内部细胞组织中发挥着重要作用。许多细胞骨架聚合物的一个关键方面是它们需要 ATP（或 GTP）形式的化学能来维持聚合状态。利用这种能量可以使细胞骨架丝发挥作用，动态响应内部和外部信号并进行自组织。该项目工作的主要目标是结合实验和计算模型来开发改进的理论框架，以理解和预测在不同尺度下观察到的这些动态细胞骨架聚合物的行为。更具体地说，拟议的工作旨在确定聚合物亚基的生化特性（包括它们燃烧 ATP 或 GTP 的速率）如何与单个丝的行为以及丝群体的整体行为相关。此外，该项目还研究丝结合蛋白如何共同作用来调节丝动力学。虽然这项工作是基础科学，但它有可能在纳米技术和合成生物学中得到实际应用。通过该项目，研究生和本科生将接受计算建模和实验生物学方面的跨学科培训。高中教师和学生也将参与研究过程。通过该项目制作的免费开源软件和教程将帮助各级学生和研究人员对动态聚合物系统有直观的了解。从技术角度来看，该项目有四个具体目标，其中大部分集中于一类细胞骨架丝称为微管。单个微管表现出一种称为动态不稳定性的戏剧性行为，其中它们在长时间的生长和解聚之间随机交替。 (1) 第一个项目目标是通过将细丝的行为与其尖端的亚基级结构联系起来，开发和测试微管动态不稳定性转变机制的假设。该方法将结合先前建立的微管动力学详细计算模型、一种用于识别和统计分类微管行为的新型数据分析工具，以及以高时间和空间分辨率获取的实验数据。 (2)第二个目标是建立对亚基生化特性（动力学速率常数）、细丝行为（例如动态不稳定性、跑步）和聚合物系统属性（例如运动）之间关系的预测性理解。 ，临界浓度，稳态）。该方法将结合计算模型（使用目标 1 中使用的模型的变体进行）和微管蛋白 PhuZ 的细菌相关实验（选择该蛋白是因为该蛋白的野生型和改变版本可以在细菌中表达并在体外进行表征） ）。 (3) 第三个目标是结合实验和计算模型来测试一组假​​设，即一组称为+TIP（微管正端跟踪蛋白）的丝结合蛋白如何协同工作来调节微管行为。 (4) 最终目标是为目标 1 至 3 中使用的软件和相关分析工具创建广泛的分发包。这些包将包括针对研究和教学社区的软件。虽然我们研究的重点是微管，但由此产生的对聚合丝系统的多尺度理解应该更广泛地适用于稳态（能量利用）聚合物，包括肌动蛋白、细菌丝和通过生物技术创建的聚合物。该奖项反映了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，NSF 的法定使命被认为值得支持。

项目成果

Overexpression of the microtubule-binding protein CLIP-170 induces a +TIP network superstructure consistent with a biomolecular condensate

微管结合蛋白 CLIP-170 的过度表达诱导与生物分子凝聚体一致的 TIP 网络超结构

• DOI: 10.1371/journal.pone.0260401
• 发表时间: 2021
• 期刊: PLOS ONE
• 影响因子: 3.7
• 作者: Wu YO;Bryant AT;Nelson NT;Madey AG;Fernandes GF;Goodson HV
• 通讯作者: Goodson HV

The CLIP-170 N-terminal domain binds directly to both F-actin and microtubules in a mutually exclusive manner

CLIP-170 N 末端结构域以相互排斥的方式直接结合 F-肌动蛋白和微管

• DOI: 10.1016/j.jbc.2022.101820
• 发表时间: 2022-05
• 期刊: Journal of Biological Chemistry
• 影响因子: 4.8
• 作者: Wu, Yueh-Fu O.;Miller, Rachel A.;Alberico, Emily O.;Huang, Yaobing A. P.;Bryant, Annamarie T.;Nelson, Nora T.;Jonasson, Erin M.;Goodson, Holly, V
• 通讯作者: Goodson, Holly, V

Quantification of microtubule stutters: dynamic instability behaviors that are strongly associated with catastrophe

微管口吃的量化：与灾难密切相关的动态不稳定行为

• DOI: 10.1091/mbc.e20-06-0348
• 发表时间: 2022-03-01
• 期刊: Molecular Biology of the Cell
• 影响因子: 3.3
• 作者: Mahserejian, Shant M.;Scripture, Jared P.;Mauro, Ava J.;Lawrence, Elizabeth J.;Jonasson, Erin M.;Murray, Kristopher S.;Li, Jun;Gardner, Melissa;Alber, Mark;Zanic, Marija;Goodson, Holly V.
• 通讯作者: Goodson, Holly V.

Behaviors of individual microtubules and microtubule populations relative to critical concentrations: dynamic instability occurs when critical concentrations are driven apart by nucleotide hydrolysis

个体微管和微管群相对于临界浓度的行为：当临界浓度因核苷酸水解而分开时，会发生动态不稳定

• DOI: 10.1091/mbc.e19-02-0101
• 发表时间: 2020-03
• 期刊: Molecular Biology of the Cell
• 影响因子: 3.3
• 作者: Jonasson, Erin M.;Mauro, Ava J.;Li, Chunlei;Labuz, Ellen C.;Mahserejian, Shant M.;Scripture, Jared P.;Gregoretti, Ivan V.;Alber, Mark;Goodson, Holly V.;Mogilner, Ale
• 通讯作者: Mogilner, Ale

Developing Evolutionary Cell Biology

发展进化细胞生物学

• DOI: 10.1016/j.devcel.2018.11.006
• 发表时间: 2018-11
• 期刊: Developmental Cell
• 影响因子: 11.8
• 作者: Titus, Margaret A.;Goodson, Holly V.
• 通讯作者: Goodson, Holly V.