Cellular regulation of myosin-I

描述（由申请人提供）：本提案的工作重点是确定脊椎动物肌球蛋白-I的调节机制。肌球蛋白-I同种型是肌球蛋白超家族的单头、膜相关成员，在大多数真核细胞中发现。肌球蛋白-Is包括在人类中发现的最大的非常规肌球蛋白家族（8个基因）。脊椎动物肌球蛋白-I家族的大尺寸和表达谱使其成为最多样化的类别之一。肌球蛋白在膜动力学、细胞骨架结构、机械信号转导、内体加工以及可能的核转录中起重要作用。我们现在才了解这类重要马达的分子作用和调节模式。因此，我们的努力在这个建议集中在了解脊椎动物肌球蛋白-I异构体的物理特性和这些特性的调节。我们将使用细胞生物学，生物物理学和生物化学技术的组合来研究以下具体目标： 1.肌球蛋白-I与脂质膜的生化和细胞相互作用。我们将研究肌球蛋白-I亚型的信号脂质的特异性，我们将确定磷酸肌醇结合和释放的动力学，我们将确定肌球蛋白-I亚型与膜的能动相互作用。 2.肌球蛋白-I的微生物学调节。非肌肉原肌球蛋白在调节肌球蛋白-I的定位和活性方面起着重要作用，但对这种基本的调节模式知之甚少。我们将研究原肌球蛋白和肌球蛋白-I相互作用的生物化学细节，目的是了解肌球蛋白-I通过微丝的细胞调节。 3.肌球蛋白-I作为张力传感器的表征。肌球蛋白-I是一个低占空比电机在没有负载。然而，我们的新研究表明，在小负载下，肌动蛋白附着的寿命显着增加，使其成为一个高占空比电机。我们将研究力和运动密度在调节进行性运动中的作用，并研究肌球蛋白I家族内的机械多样性。

DESCRIPTION (provided by applicant): Efforts in this proposal focus on determining the mechanisms of regulation of vertebrate myosin-I. Myosin-I isoforms are the single-headed, membrane-associated members of the myosin superfamily that are found in most eukaryotic cells. Myosin-Is comprise the largest unconventional myosin family found in humans (eight genes). The large size and expression profile of the vertebrate myosin-I family distinguishes it as one of the most diverse classes. Myosin-Is play essential roles in membrane dynamics, cytoskeletal structure, mechanical signal-transduction, endosome processing, and possibly nuclear transcription. We are only now learning the molecular roles and modes of regulation of this important class of motors. Therefore, our efforts in this proposal focus on understanding the physical properties of vertebrate myosin-I isoforms and the regulation of these properties. We will use a combination of cell biological, biophysical, and biochemical techniques to investigate the following specific aims: 1. Biochemical and cellular interactions of myosin-I with lipid membranes. We will investigate the specificity of myosin-I isoforms for signaling lipids, we will determine the kinetics of phosphoinositide binding and release, and we will determine the motile interactions of myosin-I isoforms with membranes. 2. Microfilament-based regulation of myosin-I. Non-muscle tropomyosin plays an important role in regulating myosin-I localization and activity, but very little is known about this fundamental mode of regulation. We will investigate the biochemical details of the tropomyosin and myosin-I interaction with the goal of understanding the cellular regulation of myosin-I by microfilaments. 3. Characterization of myosin-I as a tension-sensor. Myosin-I is a low-duty ratio motor in the absence of load. However, our new investigations show that under small loads, the lifetime of actin attachment is increased dramatically, such that it becomes a high duty ratio motor. We will investigate the role of force and motor density in regulating processive motility, and we will investigate the mechanical diversity within the myosin-I family.