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Dynein arm-function studied by mutants and in vitro motility

通过突变体和体外运动研究动力蛋白臂功能

基本信息

批准号：
04454608
负责人：
KAMIYA Ritsu
金额：
$ 3.97万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454608/
关键词：
Chlamydomonas dynein flagella mutant motility 鞭毛運動細胞運動

项目摘要

Dynein plays a most important function in the flagellar beating mechanism. Flagellar dynein comprises as many as ten different heavy chains in the inner and outer arms, but the specific function of each heavy chain is unknown. To approach the function of various heavy chains, we carried out two lines of experiments : isolation Chlamydomonas mutants that lack particular heavy chains, and assay of the motile activity of isolated heavy chains.In the first approach, we isolated a mutant that lacks part of the beta heavy chain of the outer arm and two kinds of mutants that lack different sets of inner-arm heavy chains. Studies on these mutants have indicated that (1) the beta heavy chain is more important than the alpha heavy chain in the function of outer arm dynein, and (2) five out of the eight known inner-arm heavy chains are unnecessary for the generation of flagellar beating.In the second line of study, we first developed a system in which dissociated outer-arm dynein, an alpha/beta subparticle and a gamma subparticle, can be recombined and attached to an outer arm-less mutant, oda1. This study again demonstrated that the beta heavy chain is important in the outer arm function. Finally, we developed an in vitro motility assay system using isolated inner arm dynein subspecies. Six out of the seven subspecies (containing eight heavy chains) were found to have activities to translocate microtubules in vitro. Different subspecies differ in maximal velocity and in the ability to cause microtubule rotation. These studies indicate that the flagella contain various dyneins that differ functionally. How these different dyneins cooperate in producing regular flagellar beating is a fascinating future problem.

动力蛋白在鞭毛的跳动机制中起着重要的作用。鞭毛动力蛋白在内外臂中包含多达10条不同的重链，但每条重链的具体功能尚不清楚。为了探讨各种重链的功能，我们进行了两条实验：分离缺乏特定重链的衣原体突变体，并测定分离的重链的运动活性。在第一种方法中，我们分离了一种缺乏外臂部分β重链的突变体和两种缺乏不同套内臂重链的突变体。对这些突变体的研究表明：（1）在外臂动力蛋白的功能中，β重链比α重链更重要，（2）八条已知的内臂重链中有五条对于鞭毛跳动的产生是不必要的。在第二条研究中，我们首先开发了一个系统，其中解离的外臂动力蛋白，一个α/β亚粒子和一个γ亚粒子，可以被重组并附着到外部无臂突变体ODA 1上。该研究再次证明β重链在外臂功能中是重要的。最后，我们开发了一个体外运动分析系统，使用分离的内臂动力蛋白亚种。发现7个亚种中的6个（含有8条重链）具有体外微管移位活性。不同的亚种在最大速度和引起微管旋转的能力上不同。这些研究表明，鞭毛含有不同功能的动力蛋白。这些不同的动力蛋白如何合作产生有规律的鞭毛跳动是一个迷人的未来问题。