Structure and function of the contractile vacuoles of Chlamydomonas reinhardtii (Chlorophyta)

收缩液泡（CVs）是许多单细胞生物中发现的渗透调节细胞器。一般来说，人们对CVs的功能知之甚少，特别是在绿藻中。我最近开始分析两种绿藻莱茵衣藻（Chlamydomonas reinhardtii）和雪菲藻（Scherffelia dubia）以及绿中柱头链藻（Mesostigma viride）中CVs的结构和功能。虽然这三种藻类很可能代表了三种不同的淡水栖息地入侵，但这些属的CV的结构和功能显示出一些相似之处。衣藻是这三种藻类中唯一的遗传易感生物。出于这个原因，我计划使用衣藻进行这项拨款提案中概述的研究。本工作的主要重点将是利用插入诱变方法分离衣藻的CV突变体。然后我们将对分离的突变体进行功能分析。此外，我们还将研究衣藻基因组中v - atp酶、空泡焦磷酸酶（V-PPase）和单一水通道蛋白的功能。这些蛋白与包括衣藻在内的各种系统的CV功能有关。我将使用RNAi， GFP/抗体技术结合光学和电子显微镜来研究这些蛋白质的功能。

Contractile vacuoles (CVs) are osmoregulatory organelles found in many unicellular organisms. Little is known about the function of CVs in general and especially in green algae. I have recently started to analyze the structure and function of CVs in the two chlorophyte algae Chlamydomonas reinhardtii and Scherffelia dubia, and the streptophyte Mesostigma viride. Although the three algae represent most likely three different invasions of the freshwater habitat, the structure and function of the CV in these genera show several similarities. Chlamydomonas is the only genetically tractable organism of these three algae. For this reason, I plan to use Chlamydomonas for the studies outlined in this grant proposal. The major focus of this work will be the isolation of CV mutants in Chlamydomonas using an insertional mutagenesis approach. We will then perform a functional analysis of the isolated mutants. In addition we will investigate the function of V-ATPase, vacuolar pyrophosphatase (V-PPase), and the single aquaporin present in the genome of Chlamydomonas. These proteins have been implicated in CV function in various systems including Chlamydomonas. I will investigate the function of these proteins using RNAi, GFP/antibody technology combined with light- and electron microscopy.