ANALYSES OF DYNEIN HEAVY CHAIN MUTATION IN TETRAHYMENA

描述(摘自摘要)：纤毛和鞭毛很复杂 真核生物中普遍存在的微管细胞器，包括人类， 当它们被发现在呼吸道，女性生殖道 以及在精子细胞上。纤毛和鞭毛功能障碍由 据报道，动力蛋白臂的缺失会导致呼吸和生育。 人类和其他哺乳动物的问题。这项工作的长期目标是 了解纤毛轴丝中动力蛋白的组织方式 了解每种不同动力蛋白在纤毛运动中的作用(S)。这将是 通过使用靶向基因敲除技术将突变引入到 单细胞原生动物四膜虫的动力蛋白重链编码基因 嗜热菌。为了启动这些研究，在 四膜虫DYH6基因。该突变体(DYH6 KO)的初步鉴定 表明从突变体中分离出来的轴丝缺少这两种动力蛋白 组成18s动力蛋白的重链。本项目的具体目标为1) 为了检验Dyh6p是四膜虫内臂成分的假设 通过确定18S动力蛋白在轴丝中的位置和通过 完成18S动力蛋白的表征；2)确定18S的作用 动力蛋白通过决定滑动频率在纤毛运动中发挥作用 在没有或存在胰蛋白酶的情况下，分离的轴突发生解体 测定微管二联体在 解体，并测定突变纤毛和野生型纤毛的波形； 3)通过测定DYH6KO突变对纤毛运动的影响 测定突变细胞和野生型细胞的游泳速度和行为。这个 这项工作的结果将增加对动力蛋白组织和 纤毛和鞭毛的功能。

DESCRIPTION (adapted from abstract): Cilia and flagella are complex microtubular organelles that are ubiquitous among eukaryotes, including humans, where they are found in the respiratory tract, the female reproductive tract and on sperm cells. Ciliary and flagellar dysfunction resulting from the absence of dynein arms has been reported to cause respiratory and fertility problems in humans and other mammals. The long term objectives of this work are to understand the ways dyneins are organized in ciliary axonemes and to understand the role(s) of each different dynein in ciliary motility. This will be done by using targeted gene knockout techniques to introduce mutations into genes encoding dynein heavy chains in the unicellular protozoan, Tetrahymena thermophila. To initiate those studies a mutation was generated in the Tetrahymena DYH6 gene. Initial characterization of this mutant (DYH6 KO) indicates that axonemes isolated form the mutant are missing the two dynein heavy chains that make up 18S dynein. The specific aims of this project are 1) to test the hypothesis that Dyh6p is a component of Tetrahymena inner arm dynein I by determining where 18S dynein is located in the axoneme and by completing the characterization of 18S dynein; 2) to determine the role 18S dynein plays in ciliary motility by determining the frequency of sliding disintegration of isolated axonernes in the absence or presence of trypsin, by determining the rate at which microtubule doublets slide apart during disintegration, and by determining the wave form in mutant and wild-type cilia; 3) by determining the effect of the DYH6 KO mutation on ciliary motility by determining swimming velocity and behavior in mutant and wild-type cells. The results of this work should add to the understanding of dynein organization and function in cilia and flagella.