Most freshwater flagellates use contractile vacuoles (CVs) to expel excess water. We have used Chlamydomonas reinhardtii as a green model system to investigate CV function during adaptation to osmotic changes in culture medium. We show that the contractile vacuole in Chlamydomonas is regulated in two different ways. The size of the contractile vacuoles increases during cell growth, with the contraction interval strongly depending on the osmotic strength of the medium. In contrast, there are only small fluctuations in cytosolic osmolarity and plasma membrane permeability. Modeling of the CV membrane permeability indicates that only a small osmotic gradient is necessary for water flux into the CV, which most likely is facilitated by the aquaporin major intrinsic protein 1 (MIP1). We show that MIP1 is localized to the contractile vacuole, and that the expression rate and protein level of MIP1 exhibit only minor fluctuations under different osmotic conditions. In contrast, SEC6, a protein of the exocyst complex that is required for the water expulsion step, and a dynamin-like protein are upregulated under strong hypotonic conditions. The overexpression of a CreMIP1-GFP construct did not change the physiology of the CV. The functional implications of these results are discussed.
In hypotonic media, cells/organisms take up water by osmosis. Water uptake occurs only if the water potential inside the cell is more negative than that outside the cell. The amount of water taken up in a given time depends on (i) the water potential gradient between the medium and the cell interior, (ii) the surface area of the plasma membrane (PM), and (iii) the extent to which the PM is permeable to water. Cells can use three strategies to prevent bursting caused by the osmotic influx of water in a hypotonic environment. First, organisms can modify the composition of its interior, its surface area, and/or its PM permeability to limit water uptake. Second, organisms can use a cell wall to generate a cell wall pressure potential (turgor) to balance the osmotic water potential, as many algae, plants, and fungi do. However, many unicellular protists do not possess a rigid cell wall. Instead, these protists use a third strategy: they employ water pumps called contractile vacuoles (CVs) to remove excess water. CVs are specialized vacuoles that slowly accumulate water during diastole and periodically expel the liquid rapidly into the medium (systole) (1-4).Although CV morphologies and behaviors differ among various organisms (see for a recent comparison of the contractile vacuoles of a variety of protists), the basic mechanism (water uptake into the CV by osmosis) appears to be conserved between different eukaryotes. The same proteins/ cellular processes have been implicated in CV function in Amoeba, Dictyostelium, Paramecium, Trypanosoma, and green algae (e.g., proton pumps and aquaporins) (3, 4, 6). However, we still do not know anything about what other factors (ion transport systems) are needed to build up the necessary osmotic gradient in a...