MOLECULAR AND GENETIC ANALYSIS OF FLAGELLAR DYNEINS

鞭毛动力蛋白ATP酶是一种大型的多亚基马达酶。 最 纤毛和鞭毛上有两排不同的动力蛋白 双峰微管，两者都有助于微管滑动， 鞭毛能动性 外排的动力蛋白更接近于 细胞质动力蛋白含有两个或三个Ca. 500 kd催化重质 链，两个CA。75 kd中间链和多个轻链。 虽然动力蛋白作为微管相关马达的作用， 纤毛和细胞质运动是公认的，很少有人知道 它们的调节或它们与所承载的负载的连接方式 （细胞质中的小泡，纤毛和鞭毛中的双线微管）。 拟议实验的长期目标是确定 鞭毛动力蛋白重链、中间链和轻链 鞭毛运动，并表征动力蛋白之间的相互作用 亚基和双联体微管相关蛋白是重要的 在鞭毛装配过程中用于动力蛋白附着和调节 外排动力蛋白马达。利用衣原体的外排动力蛋白 莱茵氏鞭毛作为一个模型系统，正常动力蛋白所必需的基因 组装和功能正在被克隆、测序和遗传作图。 各亚基在动力蛋白产生和调节中的作用 然后通过表型表征分析运动活动， 每个基因座的突变以及体外改变和体内表达 克隆的基因。 这里提出的实验旨在探索 α和β重链的结构-功能关系。链和 70 kd中间链，并使用插入失活诱变 克隆更多的动力蛋白组装基因。

Flagellar dynein ATPases are large, multi-subunit motor enzymes. Most cilia and flagella have two non-identical rows of dyneins attached to each doublet microtubule, both of which contribute to microtubule sliding and flagellar motility. Dyneins of the outer row more closely resemble cytoplasmic dyneins in containing two or three ca. 500 kd catalytic heavy chains, two Ca. 75 kd intermediate chains and multiple light chains. While the role of dyneins as microtubule-associated motors for both ciliary and cytoplasmic motility is well recognized, little is known about their regulation or their mode of attachment to the loads they carry (vesicles in the cytoplasm, doublet microtubules in cilia and flagella). Long term goals of the proposed experiments are to determine the roles of flagellar dynein heavy chains, intermediate chains and light chains in flagellar motility, and to characterize interactions between dynein subunits and doublet microtubule-associated proteins that are important for dynein attachment during flagellar assembly and for regulation of outer row dynein motors. Using the outer row dynein of chlamydomonas reinhardtii flagella as a model system, genes essential for normal dynein assembly and function are being cloned, sequenced, and genetically mapped. The function of each subunit in the generation and regulation of dynein motor activity is then being analyzed by phenotypic characterization of mutations at each locus and by in vitro alteration and in vivo expression of cloned genes. Experiments proposed here are designed to explore structure-function relationships in the alpha and beta heavy. chains and 70 kd intermediate chain, and to use insertional inactivation mutagenesis to clone additional dynein assembly genes.