Mutational Studies of Processive Myosin Motors

描述（由申请人提供）：这是对母体拨款GM078097，进程性肌球蛋白运动的突变研究的竞争性修订。它试图通过在培养的哺乳动物细胞中提出运动蛋白的实验来扩大原始应用的范围，与纯粹涉及体外表征的母体应用形成对比。此外，它还发挥了创造就业机会（将雇用一名新的博士后）的作用，并通过这种人力投入加快了研究的节奏。第一个目标是确定肌凝蛋白V的哪些特征对于它在活细胞中有效地通过构成肌动蛋白皮层的网络是必不可少的。基于肌凝蛋白V的货物运输中，电机占空比和每货物电机数量的重要性将在COS-7细胞中确定。其次，我们将研究如何在细胞内促进货物的转移，从微管上基于动力蛋白的运动到肌动蛋白上基于肌凝蛋白v的运输。运动蛋白和肌凝蛋白V会附着在相同的货物上，跟随运输从细胞中心到外围。这项工作的影响将是确定我们在体外研究中与肌球蛋白V和肌动蛋白相关的基本特性在细胞内也起作用的程度。许多人类疾病是由于沿肌动蛋白或微管的货物运输中断，或由于疾病蛋白作为这些运输蛋白的货物。更好地了解分子马达在细胞中沿着其轨迹运动的机制对于进一步了解这些疾病的基础至关重要。

DESCRIPTION (provided by applicant): This is a competitive revision of parent grant GM078097, Mutational Studies of Processive Myosin Motors. It seeks to expand the scope of the original application by proposing experiments with motor proteins in cultured mammalian cells, in contrast to the parent application which purely involved in vitro characterization. Moreover, it fulfills both the role of job creation (a new postdoctoral associate will be hired) as well as accelerating the tempo of research by this input of manpower. The first aim seeks to establish what features of myosin V are essential for it to effectively maneuver through the meshwork that constitutes the actin cortex in a living cell. The importance of duty cycle of the motor and motor number per cargo for myosin V based cargo-transport will be determined in COS-7 cells. Secondly, we will investigate how the handoff of cargo, from kinesin-based movement on microtubules to myosin Vbased transport on actin, is facilitated within the cell. Kinesin and myosin V will be attached to the same cargo to follow transport from the cell center out to the periphery. The impact of this work will be to establish the extent to which the basic properties we associate with myosin V and kinesin from in vitro studies are also operative within the cell. A number of human diseases result from disruption of cargo transport along actin or microtubules, or from disease proteins acting as cargo for these transport proteins. A better understanding of the mechanism by which molecular motors move along their track in the cell is critical to further understand the basis of these diseases. PUBLIC HEALTH RELEVANCE: A number of human diseases result from disruption of cargo transport along actin or microtubules, or from disease proteins acting as cargo for these transport proteins. A better understanding of the mechanism by which molecular motors move along their track in the cell is critical to further understand the basis of these diseases.