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

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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.