MECHANISMS AND FUNCTIONS OF SUBCELLULAR MOTILITY

亚细胞运动的机制和功能

• 批准号: 6266214
• 负责人: WILLIAM M SAXTON
• 金额: $ 28.15万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-08-01 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6266214
• 关键词: Drosophilidae; arthropod genetics; cell motility; cellular polarity; cytoskeletal proteins; electron microscopy; intracellular transport; invertebrate embryology; kinesin; microtubules; molecular genetics; nerve /myelin protein; neuronal transport; oogenesis; protein structure function; protein transport

DESCRIPTION (applicant's description): In the cells of metazoan animals, motor proteins that move along microtubules drive the active transport of chromosomes, organelles and other macromolecular cargoes. Motor-based transport processes are fundamental for cell growth, division, signaling, and asymmetry. My lab is investigating the mechanisms that generate and control microtubule-based movements, taking advantage of the sophisticated genetics of Drosophila, its sequenced genome, and a variety of biochemical/cell biological approaches. To understand how motors move their cargoes, 3 aspects of each motor's activity must be defined: how productive motion is derived from chemical energy, how specific motor-cargo linkages are made, and how these two activities are controlled. Our knowledge of mechanochemical mechanisms has progressed rapidly, but linkage and regulatory mechanisms remain poorly understood. Our proposed studies are aimed at identifying motor protein functions, defining specific motor-cargo linkages and probing regulatory mechanisms. We previously discovered that ubiquitous kinesin-I, the founder of the kinesin superfamily, is an important anterograde motor for fast organelle transport in Drosophila axons; its specific cargoes remain uncertain. In the last funding period we used genetic enhancer screens to identify proteins that influence kinesin-l function in axons. Eight different loci were identified: Kinesin light chain, dynein, dynactin, ABL tyrosine kinase, ENMIASP, and 3 unknowns that we will characterize (EK2/1O, EK4, EK5). We propose to study specific motor-cargo relationships and the functions of the new fast axonal transport proteins using a GFP-based single organeIIe tracking approach in live neurons, as well as standard biochemical/cell biological approaches. ABL and ENA are particularly exciting; known to interact with the actin cytoskeleton, they now also appear to regulate kinesin-l in axons. Mapping of EK4 has placed it in a small region that includes 2-3 good candidate genes, one of which is a membrane protein implicated in mitochondrial motility. Pursuing kinesin-l functions in other tissues, we have discovered essential roles in axial patterning during oogenesis. Kinesin-l is required specifically for the localization of the posterior determinant oskar mRNA. It is also needed for Gurken secretion by the oocyte, which signals follicle cells to establish the dorsal pole. We propose to use biochemical interaction tests, immunolocalization, and genetic screens to determine how kinesin-l interacts with the oskar transport complex, and how it facilitates Gurken secretion.

描述（申请人的描述）：在后生动物的细胞中 沿着微管移动的蛋白质驱动主动传输的蛋白质 染色体，细胞器和其他大分子货物。基于运动的运输 过程对于细胞生长，分裂，信号传导和不对称性至关重要。 我的实验室正在研究产生和控制的机制 基于微管的运动，利用了复杂的遗传学 果蝇，其测序基因组和多种生化/细胞生物学 方法。 要了解电动机如何移动货物，这是每个电动机活动的三个方面 必须定义：生产性运动是如何从化学能中得出的 建立了特定的电动货车连接，这两个活动是如何 受控。我们对机械化学机制的了解迅速发展， 但是连锁和监管机制仍然很少理解。我们提出的 研究旨在鉴定运动蛋白功能，定义特定 电动货车连锁和探测调节机制。我们以前 发现无处不在的运动蛋白I是超级家族的动机的创始人 是果蝇快速器官传输的重要顺行电动机 轴突；它的具体货物仍然不确定。在最后的资金期内 使用了遗传增强子筛选来鉴定影响驱动蛋白L的蛋白质 轴突的功能。确定了八个不同的基因座：动力素轻链， Dynein，Dynactin，ABL酪氨酸激酶，Enmiasp和3个未知数我们将 表征（EK2/1O，EK4，EK5）。我们建议研究特定的电动机货车 使用新快速轴突运输蛋白的关系和功能 现场神经元中的基于GFP的单器官跟踪方法以及 标准生化/细胞生物学方法。 ABL和ENA特别是 令人兴奋；已知会与肌动蛋白细胞骨架相互作用，现在也出现 调节轴突中的驱动蛋白-L。 EK4的映射将其放置在一个小区域 其中包括2-3个好候选基因，其中之一是膜蛋白 与线粒体运动有关。 在其他组织中追求动力蛋白-L功能，我们发现了必不可少的 在卵子发生过程中的轴向图案中的作用。驱动蛋白-l是需要特别需要的 为了定位后决定因素OSKAR mRNA。也需要 对于卵母细胞的Gurken分泌，该分泌信号是卵泡细胞建立 背杆。我们建议使用生化互动测试， 免疫定位和遗传筛选以确定驱动蛋白L相互作用 随着Oskar运输综合体以及它如何促进Gurken的分泌。