Mechanism and function of microvillar myosin-1

描述（由申请人提供）：该提案的重点是发展我们对1类肌球蛋白的理解：普遍表达的单体、膜结合、基于肌动蛋白的马达，其参与多种细胞功能，包括细胞器运输， 转录、宿主防御、细胞运动和机械感觉。我们对肌球蛋白-1的研究主要集中在“刷状缘”上，这是一种紧密排列的微绒毛阵列，从许多转运上皮细胞类型的顶端表面延伸出来。这个细胞器是许多肌球蛋白超家族成员的家园，其中最丰富的是肌球蛋白-1a（Myo 1a），这是八种脊椎动物1类肌球蛋白之一。我们的实验室利用细胞生物学和生物物理学方法的独特组合来发现：（i）Myo 1a有助于膜-细胞骨架粘附，这对于维持正常的刷状缘结构至关重要，（ii）Myo 1a能够将富含宿主防御机制的膜囊泡从微绒毛尖端释放到肠腔中。最近的研究也强调了Myo 1a功能的生理意义，这些研究表明该马达的突变与人类结直肠肿瘤的形成有关。虽然我们在阐明Myo 1a的生物学作用方面取得了实质性进展，但控制这种和其他肌球蛋白-1功能的分子特性，相互作用和事件仍然缺乏特征。该提案的目标是发展我们对Myo 1a有助于刷状缘功能的基本生物化学和生物物理特性的理解。为此，目标1将检查单一Myo 1a分子与活细胞的质膜和支持的双层在体外相互作用的单一属性，目标2将检查Myo 1a绑定到支持的双层的力产生潜力，和目标3将调查力传感在Myo 1a的动力学和功能的调节中的作用。由于刷状缘形成和维持的缺陷是对人类健康构成重大威胁的许多疾病的核心，因此对管理Myo 1a行为的分子机制的深入了解将提供最终可能用于开发旨在修复畸形或受损刷状缘的治疗方法的信息。

DESCRIPTION (provided by applicant): This proposal is focused on developing our understanding of class 1 myosins: ubiquitously expressed monomeric, membrane binding, actin-based motors that participate in diverse cellular functions, including organelle trafficking, transcription, host defense, cell motility, and mechano-sensation. Our studies of myosin-1 are centered on the 'brush border', a tightly packed array of microvilli that extends from the apical surface of many transporting epithelial cells types. This organelle is home to a number of myosin superfamily members, with the most abundant being myosin-1a (Myo1a), one of eight vertebrate class 1 myosins. Our laboratory has leveraged a unique combination of cell biological and biophysical approaches to discover that: (i) Myo1a contributes to membrane-cytoskeleton adhesion, which is critical for maintaining normal brush border structure, and (ii) Myo1a powers the release of membrane vesicles enriched in host defense machinery from microvillar tips into the intestinal lumen. The physiological significance of Myo1a function is also underscored by recent studies showing that mutations in this motor are linked to colorectal tumor formation in humans. While we have made substantial progress toward elucidating the biological roles of Myo1a, the molecular properties, interactions, and events that govern the function of this and other myosins-1 remain poorly characterized. The goal of this proposal is to develop our understanding of the fundamental biochemical and biophysical properties that enable Myo1a to contribute to brush border function. To this end, Aim 1 will examine the unitary properties of single Myo1a molecules interacting with the plasma membrane of live cells and supported bilayers in vitro, Aim 2 will examine the force generating potential of Myo1a bound to supported bilayers, and Aim 3 will investigate the role of force sensing in the regulation of Myo1a dynamics and function. Because defects in brush border formation and maintenance are at the core of numerous diseases that pose significant threats to human health, developing insight on the molecular mechanisms that govern Myo1a behavior will provide information that may ultimately be used in the development of therapeutics aimed at repairing malformed or damaged brush borders.