A Cell-Based Screen for Inhibitors of Flagella-Driven Motility in Chlamydomonas Reveals a Novel Modulator of Ciliary Length and Retrograde Actin Flow
A Cell-Based Screen for Inhibitors of Flagella-Driven Motility in Chlamydomonas Reveals a Novel Modulator of Ciliary Length and Retrograde Actin Flow
复制标题
DOI：
10.1002/cm.20504
复制
发表时间：
2011-03-01
期刊：
影响因子：
2.9
通讯作者：
Marshall, Wallace F.
中科院分区：
文献类型：
--
作者：
Engel, Benjamin D.;Ishikawa, Hiroaki;Marshall, Wallace F.
纤毛是运动和感觉细胞器，在生理上起着至关重要的作用。纤毛缺陷可引起许多人类疾病症状，包括多囊肾、脑积水和视网膜变性。尽管这些细胞器很重要，但它们的组装和功能尚不完全清楚。单细胞绿藻莱茵衣藻作为纤毛组装和功能研究的模型系统具有许多优点。在这里，我们描述了我们最初的努力，建立一个化学生物学工具包，以增加可用于研究这种生物体纤毛的遗传工具，与传统遗传方法相比，目标是能够在快速的时间尺度上可逆地干扰纤毛功能。我们筛选了5520种化合物，从中确定了4种候选化合物，在无毒剂量下对鞭毛具有可重复的作用。其中三种化合物导致鞭毛麻痹，一种以可逆和剂量依赖的方式诱导鞭毛缩短，并伴有鞭毛内运输速度的降低。后一种化合物还能缩短哺乳动物细胞中纤毛的长度，因此我们将这种化合物命名为“纤毛素”，因为它能缩短纤毛。该化合物还能有效且可逆地抑制果蝇S2细胞的微管运动和逆行肌动蛋白流动。纤毛素可能被证明对研究细胞前沿的逆行肌动蛋白流动特别有用，因为它以可调的剂量依赖性方式减缓逆行流动，直到高浓度时流动完全停止，并且这些作用在药物冲洗后迅速逆转。(C) 2011 Wiley-Liss, Inc。
Cilia are motile and sensory organelles with critical roles in physiology. Ciliary defects can cause numerous human disease symptoms including polycystic kidneys, hydrocephalus, and retinal degeneration. Despite the importance of these organelles, their assembly and function is not fully understood. The unicellular green alga Chlamydomonas reinhardtii has many advantages as a model system for studies of ciliary assembly and function. Here we describe our initial efforts to build a chemical-biology toolkit to augment the genetic tools available for studying cilia in this organism, with the goal of being able to reversibly perturb ciliary function on a rapid time-scale compared to that available with traditional genetic methods. We screened a set of 5520 compounds from which we identified four candidate compounds with reproducible effects on flagella at nontoxic doses. Three of these compounds resulted in flagellar paralysis and one induced flagellar shortening in a reversible and dose-dependent fashion, accompanied by a reduction in the speed of intraflagellar transport. This latter compound also reduced the length of cilia in mammalian cells, hence we named the compound "ciliabrevin'' due to its ability to shorten cilia. This compound also robustly and reversibly inhibited microtubule movement and retrograde actin flow in Drosophila S2 cells. Ciliabrevin may prove especially useful for the study of retrograde actin flow at the leading edge of cells, as it slows the retrograde flow in a tunable dose-dependent fashion until flow completely stops at high concentrations, and these effects are quickly reversed upon washout of the drug. (C) 2011 Wiley-Liss, Inc.