Molecular Mechanisms Governing Cooperating Motors

控制协作电机的分子机制

• 批准号: 8102716
• 负责人: Michael R Diehl
• 金额: $ 27.86万
• 依托单位: RICE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8102716
• 关键词: Accounting; Address; Affect; Attenuated; Behavior; Binding; Binding Proteins; Biochemical; Biological Assay; Cells; Cooperative Behavior; Coupled; Coupling; Cytoskeleton; Data; Defect; Disease; Dynein ATPase; Engineering; Evolution; Foundations; Grouping; In Vitro; Individual; Intracellular Transport; Kinesin; Knowledge; Laboratories; Length; Life; Link; Mechanics; Methods; Microinjections; Microtubules; Molecular; Molecular Motors; Monitor; Motion; Motor; Mutation; Nature; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Outcome; Pathology; Pharmaceutical Preparations; Physiology; Polymers; Production; Property; Proteins; Public Health; Publishing; Regulation; Relative (related person); Research; Role; Running; System; Techniques; Technology; Testing; Theoretical model; Therapeutic Agents; Time; Transport Process; War; Work; behavior influence; cell motility; design; human disease; insight; novel; optical traps; particle; protein complex; public health relevance; research study; single molecule; stem

DESCRIPTION (provided by applicant): Many important subcellular cargos are transported along the microtubule cytoskeleton by groups of kinesin and/or dynein motor proteins. Specific proteins and cargos that are implicated in diseases, particularly neurodegeneration, are known to interact with multiple motor molecules, and hence, there are likely strong links between collective motor function and human diseases. The grouping of motors is believed to be important for specific transport challenges that may require high force production. Furthermore, many cargos bind to kinesin and dynein and move bidirectionally along microtubules. This behavior is known to influence the spatio-temporal evolution and final distributions of cargos in cells. Yet, mechanisms governing collective motor transport are not well understood, and overall, existing methods to investigate these problems are limited by inabilities to precisely characterize or control the number of motors on cargos. This project addresses these issues by building upon our newly developed biosynthetic strategies to create structurally- defined systems of interacting motor molecules, and biophysical assays that allow collective motor dynamics to be monitored at the single-molecule level. By combining these capabilities, we have established that interactions among multiple kinesin-1 molecules contribute significantly to collective transport behaviors (e.g., cargo run lengths, and force production), and that despite their abilities to produce large forces, groups of kinesin motors tend to cooperate negatively. The proposed work will employ our synthetic technologies along with precision particle tracking and optical trapping methods to further evaluate the extent to which negative cooperativity influences cargo transport by multiple kinesins in cells (Aim 1). We will also examine analogous cooperative effects in motor systems composed of multiple dynein molecules (Aim 2). Both of these studies will draw connections between the properties of individual motor molecules, the nature of inter-motor interactions within motor assemblies, and collective transport parameters. In each case, multiple-motor functions will be evaluated using theoretical models that can account for all relevant biochemical states of individual motors as well as microtubule-bound configurations of motor assemblies. Finally, we will perform assays that monitor the motions of structurally-defined motor assemblies composed of kinesin and dynein molecules (Aim 3). Here, knowledge of collective behaviors among each class of motors, coupled with the ability to systematically investigate the influence of motor number and ratio on bidirectional cargo motility, should allow bidirectional transport mechanisms to be resolved. Overall, the proposed study will help to clarify observations of kinesin and dynein's apparent functional interdependence in cells, and provide new abilities to examine and interpret how defects in motor function influence intracellular transport processes. PUBLIC HEALTH RELEVANCE: Resolving mechanisms of collective motor transport will provide a foundation to interpret how transport defects, genetic and/or environmental, influence the function of motor molecules in cells. Outcomes from this work may also suggest new methods to develop and evaluate therapeutic agents directed at molecular motors.

描述（由申请人提供）：许多重要的亚细胞货物通过驱动蛋白和/或动力蛋白运动蛋白组沿着微管细胞骨架运输。已知与疾病（特别是神经退行性疾病）有关的特定蛋白质和货物与多种运动分子相互作用，因此，集体运动功能与人类疾病之间可能存在密切联系。电机的分组被认为对于可能需要高输出力的特定运输挑战非常重要。此外，许多货物与驱动蛋白和动力蛋白结合并沿着微管双向移动。已知这种行为会影响细胞中货物的时空演化和最终分布。然而，管理集体汽车运输的机制尚不清楚，总体而言，调查这些问题的现有方法因无法精确表征或控制货物上的汽车数量而受到限制。该项目通过建立我们新开发的生物合成策略来解决这些问题，以创建结构定义的相互作用运动分子系统，以及允许在单分子水平上监测集体运动动力学的生物物理测定。通过结合这些能力，我们已经确定多个驱动蛋白-1分子之间的相互作用对集体运输行为（例如，货物运行长度和力产生）有显着贡献，并且尽管它们有能力产生巨大的力，但驱动蛋白马达组往往会消极合作。拟议的工作将采用我们的合成技术以及精密粒子跟踪和光学捕获方法，以进一步评估负协同性对细胞中多个驱动蛋白的货物运输的影响程度（目标 1）。我们还将研究由多个动力蛋白分子组成的运动系统中的类似协同效应（目标 2）。这两项研究都将在单个电机分子的特性、电机组件内电机间相互作用的性质以及集体运输参数之间建立联系。在每种情况下，将使用理论模型来评估多电机功能，该理论模型可以解释各个电机的所有相关生化状态以及电机组件的微管结合配置。最后，我们将进行监测由驱动蛋白和动力蛋白分子组成的结构定义的运动组件的运动的检测（目标 3）。在这里，了解各类电机之间的集体行为，再加上系统地研究电机数量和比率对双向货物运动的影响的能力，应该可以解决双向运输机制。总体而言，拟议的研究将有助于澄清对细胞中驱动蛋白和动力蛋白明显功能相互依赖性的观察，并提供新的能力来检查和解释运动功能缺陷如何影响细胞内运输过程。 公共健康相关性：解决集体运动运输的机制将为解释遗传和/或环境的运输缺陷如何影响细胞中运动分子的功能提供基础。这项工作的结果还可能提出开发和评估针对分子马达的治疗剂的新方法。