Molecular Mechanisms Governing Cooperating Motors

控制协作电机的分子机制

• 批准号: 8496827
• 负责人: Michael R Diehl
• 金额: $ 27.2万
• 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/8496827
• 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; 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.

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/或动力蛋白马达蛋白组沿微管细胞骨架沿着转运。已知与疾病（特别是神经变性）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力产生的特定运输挑战是重要的。此外，许多货物结合驱动蛋白和动力蛋白，并沿沿着微管双向移动。已知这种行为会影响细胞中货物的时空演变和最终分布。然而，机制管理集体汽车运输没有得到很好的理解，总体而言，现有的方法来调查这些问题是有限的能力，以精确地表征或控制货物上的电机的数量。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，以创建相互作用的马达分子的结构定义系统，以及允许在单分子水平上监测集体马达动力学的生物物理测定。通过结合这些能力，我们已经确定了多个驱动蛋白-1分子之间的相互作用对集体运输行为有显著贡献（例如，货物运行长度和力的产生），并且尽管它们能够产生大的力，驱动蛋白马达组倾向于消极地合作。拟议的工作将采用我们的合成技术沿着与精确的粒子跟踪和光学捕获方法，以进一步评估负协同效应影响细胞中多种驱动蛋白的货物运输的程度（目的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合作效应（目的2）。这两项研究都将绘制单个马达分子的性质，马达组件内马达间相互作用的性质和集体运输参数之间的联系。在每种情况下，多电机功能将使用理论模型进行评估，这些模型可以解释单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，我们将进行检测，监测由驱动蛋白和动力蛋白分子组成的结构定义的电机组件的运动（目的3）。在这里，知识的集体行为之间的每一类电机，再加上有能力系统地调查电机数量和比例的影响，双向货物运动，应该允许双向运输机制得到解决。总的来说，拟议的研究将有助于澄清观察驱动蛋白和动力蛋白的明显功能相互依赖的细胞，并提供新的能力，检查和解释运动功能的缺陷如何影响细胞内的运输过程。