MECHANISMS AND FUNCTIONS OF SUBCELLULAR MOTILITY

亚细胞运动的机制和功能

• 批准号: 2723350
• 负责人: WILLIAM M SAXTON
• 金额: $ 0.29万
• 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 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2723350
• 关键词: MECHANISMS; FUNCTIONS; SUBCELLULAR; MOTILITY

Most intracellular motility processes employ motor proteins to move organelles or other cargo along cytoskeletal filaments. There are many essential cargoes, many specific destinations for them, and a large number of different motors. The key questions are: How do motors create movement? and Which motors accomplish what tasks? The proposed research will focus on the microtubule-based motor kinesin. It will investigate how kinesin interacts with microtubules, how its two motor subunits interact with one another, and what its functions are in a developmentally complex organism (Drosophila). In addition, a genetic screen will be used to identify new proteins that interact with kinesin as cargo linkers or regulators. The specific goals of the proposed research are as follows: 1) To identify amino acids of the kinesin heavy chain (KHC) that mediate binding to microtubules. This will be done by: a) identifying the amino acid changes in four mutated Drosophila kinesin heavy chain genes (khc) that show allele specific genetic interaction with a mutation in the Beta2 tubulin gene and b) testing the effects of those changes and others on the in vitro microtubule binding activity of dimerized KHC motor domains. 2) To identify amino acids of KHC that are important for the exceptionally processive movement of kinesin on microtubules. This will be done by: a) screening a large collection of lethal khc mutations for amino acid changes in a small region that is thought to be responsible for coordinating the stepping activities of the two motor domains of a KHC dimer and b) testing the effects of those changes on processive movement in vitro. 3) To determine if kinesin is important for male or female germ cell development/function or for embryogenesis. This will be done by creating chimeric flies that have germlines with no KHC and analyzing gametogenesis. fertility, and embryogenesis. 4) To study newly discovered neuronal defects caused by a loss of kinesin function: distal axonal neuropathy and photoreceptor specific retinal degeneration. These experiments may identify some of kinesin's cargoes in axons and may provide a new understanding of the role of axonal transport motors in the pathology of vertebrate neuropathies. 5) To identify and study the functions of proteins that are involved in kinesin-cargo linkage and in kinesin regulation. This will be done using a fast genetic screen of the Drosophila genome for deficiencies that cause synthetic distal neuropathy when combined with khc mutations in double heterozygotes.

大多数细胞内运动过程采用运动蛋白移动 沿细胞骨骼细丝的细胞器或其他货物。有许多 必需的货物，许多特定目的地以及一个大的 不同电动机的数量。关键问题是：电动机如何创建 移动？哪些电动机完成了什么任务？拟议的研究 将重点放在基于微管的运动动力蛋白上。它将调查 运动蛋白如何与微管相互作用，其两个电动机亚基如何 彼此互动，其功能在一个 发育复杂的生物（果蝇）。另外，遗传 屏幕将用于识别与动力蛋白相互作用的新蛋白质 作为货物接头或监管机构。 拟议研究的具体目标如下： 1）鉴定介导的驱动蛋白重链（KHC）的氨基酸 与微管结合。这将通过以下方式完成：a）识别氨基 四个突变的果蝇驱动蛋白重链基因（KHC）的酸变化 这显示了等位基因的特异性遗传相互作用与突变 beta2小管蛋白基因和b）测试这些变化的影响以及其他变化的影响 在体外微管二聚体KHC电动机上 域。 2）鉴定KHC的氨基酸对 驱动蛋白在微管上的特殊操作运动。这会 可以通过：a）筛选大量致命的KHC突变 被认为是负责的小区域的氨基酸变化 为了协调两个电动机域的垫脚活动 KHC二聚体和B）测试这些变化对过程的影响 体外运动。 3）确定驱动蛋白对于男性还是 女生殖细胞发育/功能或用于胚胎发生。这将是 通过创建具有没有KHC的种系的嵌合蝇和 分析配子发生。生育能力和胚胎发生。 4）新学习 发现的神经元缺陷是由驱动蛋白功能丧失引起的：远端 轴突神经病和感光特异性视网膜变性。这些 实验可以识别轴突中的一些动力蛋白货物，可能 对轴突运输电机在 脊椎动物神经病的病理学。 5）识别和研究 涉及运动蛋白 - 碳连接的蛋白质功能和 动力素调节。这将使用快速的遗传屏幕完成 果蝇基因组，用于引起合成远端神经病的缺陷 当与双杂合子中的KHC突变结合使用时。