Microtubule bundles in the mitotic spindle: probing how mechanical and functional robustness emerge from molecular architecture

有丝分裂纺锤体中的微管束：探讨分子结构如何产生机械和功能稳健性

• 批准号: 10226166
• 负责人: Mary Williard Elting
• 金额: $ 36.6万
• 依托单位: NORTH CAROLINA STATE UNIVERSITY RALEIGH
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10226166
• 关键词: Address; Affect; Architecture; Axon; Biochemistry; Biological Process; Building Codes; Bundling; Cell division; Cells; Chromosome Segregation; Chromosomes; Cilia; Congenital Abnormality; Coupling; Crosslinker; Disease; Engineering; Ensure; Equilibrium; Fiber; Fission Yeast; Goals; In Vitro; Kinetochores; Lead; Malignant Neoplasms; Maps; Measures; Mechanics; Microtubule Bundle; Microtubules; Mitotic spindle; Molecular; Molecular Probes; Morphology; Property; Spontaneous abortion; Stereotyping; Structure; Sum; Time; Work; base; daughter cell; flexibility; genetic information; in vivo; insight; mechanical properties; molecular scale; novel; programs; reconstitution; response; tool; transmission process

Project Summary/Abstract The mitotic spindle is a microtubule-based machine that segregates chromosomes into two new daughter cells when cells divide. Accurate spindle function is critical: mistakes lead to extra or missing chromosomes, which are associated with cancer, birth defects, and miscarriage. Spindle function requires robust coupling of biochemistry and mechanics. Yet, understanding how this self-organizing machine generates the required forces in the right place at the right time remains a challenge. Our long term goal is to determine how micron-scale mechanical properties of the spindle emerge from molecular-scale biochemistry. We focus on microtubule bundles, which provide organization and underpin rigidity in the spindle and in other microtubule-based structures. We do not understand what material properties bundling molecules impart to spindle bundles, how their molecular properties allow them to do so, or how these emergent mechanical properties are tuned for biological functions. To address these questions, we will measure quantitative readouts of how bundles respond to perturbations that alter mechanics. We take a multi-system approach to understanding bundle mechanics in mammalian kinetochore-fibers (k-fibers), which attach and segregate chromosomes; in fission yeast S. pombe spindles, whose stereotyped organization facilitates probing how specific crosslinker properties affect bundles overall; and in vitro, where we have more precise control. Our approach is organized into two programs: (1) probing the molecular and mechanical organization of spindle microtubule bundles, and (2) controlling spindle microtubule bundles to alter function through novel mechanics. In Program 1, we will determine how k-fiber organization balances competing mechanical constraints of robust force-transmission for chromosome segregation with flexibility to adapt to changing spindle morphology. We will also develop new tools to measure force between microtubules within spindle bundles, determining how these bundles effectively transmit force to achieve their mechanical functions. In Program 2, we will determine how the geometric and mechanical properties of microtubule crosslinkers impart bundle-scale properties that are adapted to particular functions. We will create engineered crosslinkers whose mechanical and geometric properties we will control, and use them to build reconstituted microtubule bundles in vitro, and to alter bundle properties in vivo. By measuring the response of these bundles to molecular-scale changes, we will determine how micron-scale properties emerge. In sum, the proposed work will map how molecular scale parts impart spindle bundles with properties that balance competing mechanical constraints. In the long term, this approach may lead to new insight into how altering the cell’s “building code” can be harnessed to target microtubule architectures with key roles in disease, or to build novel architectures. This approach can extend to understand the emergent mechanics of microtubule bundle architectures beyond the spindle, such as in cilia and axons.

项目摘要/摘要 有丝分裂主轴是基于微管的机器，将染色体分离为两个新的机器 细胞分裂时子细胞。准确的主轴功能至关重要：错误会导致额外或缺失 与癌症，先天缺陷和流产有关的染色体。主轴功能需要 生物化学和力学的强大耦合。但是，了解这种自组织的机器 在正确的时间在正确位置产生所需的力仍然是一个挑战。我们的长期目标是 确定纺锤体尺度生物化学的微米尺度机械性能。 我们专注于微管束，这些束为主轴和其他方面提供组织和基础 基于微管的结构。我们不了解捆绑分子赋予的物质特性 主轴束，它们的分子特性如何允许它们这样做，或者这些新兴机械如何 对生物学功能进行了调整。为了解决这些问题，我们将衡量定量 读数捆绑包如何响应改变力学的扰动。我们采用多系统的方法 了解哺乳动物动型纤维（K纤维）中的束机械师，它们附着并分离 染色体；在裂变酵母菌S. pombe纺锤体中，其定型组织有助于探测如何 特定的交联属性总体上影响束；在体外，我们拥有更精确的控制。 我们的方法分为两个程序：（1）探测的分子和机械组织 纺锤微管束，（2）控制纺锤微管束以通过新颖的方式改变功能 力学。在节目1中，我们将确定K纤维组织如何平衡竞争机械 强大的力传输对染色体隔离的限制，灵活地适应了变化 纺锤形态。我们还将开发新工具来测量主轴内微管之间的力 捆绑包，确定这些捆绑包如何有效传输力以实现其机械功能。在 程序2，我们将确定微管交联的几何和机械性能如何赋予 适合特定功能的束规范属性。我们将创建工程的交联器 我们将控制机械和几何特性，并使用它们来构建重构的微管束 体外，并在体内改变束性能。通过测量这些束对分子尺度的响应 变化，我们将确定微米级特性如何出现。 总而言之，拟议的工作将绘制分子尺度零件如何赋予特性的主轴束 平衡竞争的机械约束。从长远来看，这种方法可能会导致对 如何利用更改单元格的“建筑代码”来针对微管体系结构，并在 疾病，或建立新型建筑。这种方法可以扩展以了解 微管束架构超出主轴，例如纤毛和轴突。