MOLECULAR INTERACTIONS IN MOVING AXONEMES

描述(申请者描述)：本项目的目标是 了解蛋白质运输过程的机制，这是 鞭毛衣藻的组装、功能及维护。这个过程是 被称为鞭毛内转运(IFT)，在运动性和 多细胞生物体中不动的纤毛。因此，所获得的信息 在衣原体等模型系统中的IFT上，可能与 纤毛、鞭毛和感觉外节的生物发生 在人类身上。不动纤毛功能障碍影响感觉传导，而 活动纤毛功能障碍会导致呼吸道疾病、不孕不育或 发育异常，如内翻。 衣藻的鞭毛器比化学感受器更容易获得 线虫的神经元或海胆或小鼠的胚胎纤毛，它们是 用于研究IFT的其他系统。IFT涉及蛋白质颗粒， 称为IFT粒子，它连续双向运动 在基生体和鞭毛的远端之间。这些IFT粒子 被循环利用，改变鞭毛两端的大小和/或结构。 出于这些原因，我们提出了一种函数方案，其中IFT周期是 分为四个阶段，以说明地球的顺行和逆行运动 以及鞭毛两端的颗粒所起的作用。 这项建议的长期目标将从以下几个方面着手 四个具体目标：a.确定每组对温度敏感的突变体 在IFT周期的四个阶段中的一个阶段有缺陷。B.识别 与货物和/或分子马达结合的IFT颗粒的蛋白质。C.至 克隆执行IFT的机械的相关蛋白质编码基因。 D.鉴定利用IFT颗粒达到最终目的的蛋白质 鞭毛内的位置。 该项目开发的关键是一个用于 对显微图像序列进行定量分析，从而实现跟踪 在体内几秒钟内IFT粒子的运动。这 方法正在导致鉴定顺行或不同的突变体 逆行IFT以及鞭毛基体部或远端的突变体。

DESCRIPTION (applicant's description): The goal of this project is to understand the mechanism of the protein transport process that is required for assembly, function and maintenance of Chlamydomonas flagella. This process is referred to as intraflagellar transport (IFT) and is conserved in motile and immotile cilia of multicellular organisms. Therefore, the information obtained on the IFT in a model system such as Chlamydomonas, likely is pertinent to the understanding of the biogenesis of cilia, flagella and sensory outer segments in humans. Dysfunctions of immotile cilia affect sensory transduction, whereas dysfunctions of motile cilia cause respiratory ailments, sterility or developmental abnormalities such as situs inversus. The flagellar apparatus of Chlamydomonas is more accessible than chemosensory neurons of C. elegans or embryonic cilia of sea urchin or mouse, which are other systems used to study the IFT. The IFT involves protein particles, referred to as IFT particles, that move continuously and bidirectionally between the basal bodies and the distal end of flagella. These IFT particles are recycled and change size and/or structure at both flagellar extremities. For these reasons we propose a functional scheme, in which the IFT cycle is divided in four phases to account for anterograde and retrograde motion of the particles and the functions served by the particles at both ends of flagella. The long-term objective of this proposal will be approached with the following four specific aims: A. To identify sets of temperature-sensitive mutants each defective in one of the four phases of the IFT cycle. B. To identify the proteins of an IFT particle that bind to cargo and/or molecular motors. C. To clone genes encoding relevant proteins of the machinery carrying out the IFT. D. To identify the proteins that use IFT particles to reach their final location within flagella. Pivotal for the development of this project is a procedure used for quantitative analysis of sequences of microscopic images that allows tracking of the motion of IFT particles for periods of several seconds in vivo. This approach is leading to the identification of distinct mutants of anterograde or retrograde IFT as well as mutants of the basal body or distal end of flagella.