A contractile vacuole (CV) is an organelle or subcellular structure involved in osmoregulation and waste disposal. Previously, a CV was known as a pulsed or pulsed vacuole. CVs should not be confused with vacuoles that store food or water. A CV is found mainly in protists and single-celled algae. In freshwater environments, the concentration of solutes inside the cell is higher than outside the cell. Under these conditions, water flows from the environment into the cell by osmosis. Thus, the CV acts as a protective mechanism against cell expansion (and possibly explosion) due to too much water; it expels excess water from the cell by contracting. However, not all species that have a CV are freshwater organisms; some marine and soil microorganisms also have a CV. Cv is prevalent in species that do not have a cell wall, but there are exceptions.
Through the evolutionary process, CV has been largely eliminated in multicellular organisms; However, there are still in the single-celled stage of several multicellular fungi and in different types of cells in sponges, including amoebocytes, pinacocytes and choanocytes. When Rab11C is expressed as a labeled protein, it is also associated with cv. However, Rab11C does not interact with disgorgin or drainin, and overexpression of Rab11C only partially suppresses the large vacuol phenotype in disgorginous cells (unpublished data). While Rab11A is highly expressed at the vegetative stage, Rab11C is mainly expressed in 18 h development cells (Dictybase) and therefore may not be relevant for CV function in vegetative cells. To regulate osmotic pressure, most freshwater amoebae have a contractile vacuole (CV) that expels excess water from the cell. Acidocalcisomes have been involved to work alongside the contractile vacuole to respond to osmotic stress. They were detected near Trypanosoma cruzi vacuoles and were shown to merge with vacuole when cells were exposed to osmotic stress. Presumably, acidocalcisomes empty their ion content into contractile vacuoles and thus increase the osmolarity of vacuoles. [6] Protozoa have transient foods or digestive vacuoles. The number of these membrane-bound cellular organelles depends on the body`s eating habits. Some species may have a lot of them, while others may contain only one or two at a time. In ciliates, food vacuoles are formed at the base of the.
The endoplasm contains dietary vacuoles, a granular nucleus and a clear contractile vacuole. The amoeba has no mouth or anus; Food is absorbed and matter is excreted at any point on the cell surface. During feeding, cytoplasm extensions circulate around food particles, surround them and form one. The contractile vacuole, as the name suggests, expels water from the cell by contraction. The growth (water collection) and contraction (water outlet) of contractile vacuoles are periodic. A cycle lasts several seconds, depending on the type and osmolarity of the environment. The stage at which water flows into the CV is called diastole. The contraction of the contractile vacuoles and the expulsion of water from the cell are called systoles. LvsA and LvsD are proteins of the BEACH family that contain an evolutionarily conserved domain and are involved in a number of cellular processes (De Lozanne, 2003). We confirmed the results of De Lozanne, 2003, that the disruption of lvsA causes a loss of visible CV and tubules, while the loss of lvsD (in a wild-type background) does not produce a visible phenotype (Figure 6A and B).
A replacement strain of the LvsA gene in which the endogenous open LvsA reading frame fused with GFP is overexpressed by a more active actin promoter (Gerald et al., 2002) has no vacuolar phenotype. Interestingly, disgorgin`s disturbance in this context (LvsAOE/disgorgin−) causes an improved and expanded vacuol phenotype, similar to observations in cells without Disgorgin and LvsD (lvsD−/disgorgin−; Figure 6A and additional figure S2A). Overexpression of GFP-LvsD in disgorgin− cells, similar to lvsA disturbance, suppresses the large vacuol phenotype of disgorgin− cells (Figure 6A). These results suggest that LvsA and LvsD regulate the CV system, but have different and possibly opposite functions. Disgorgin localized in lvsA− cells to pierce structures next to the plasma membrane that have been co-located with dajumine RFP, suggesting that they are CV structures (Figure 6C; Data not displayed). Since Disgorgin was only located in the CV at the end of the loading phase, this suggests that the aberrant CV structures in the lvsA− cells are remnants of CV that stopped growing after discharge. We suggest that LvsA work to maintain the integrity of the CV during the unloading phase. Since previous reports have shown that LvsA translocates into the CV membrane after the vacuole has reached its maximum diameter (De Lozanne, 2003), we examined the detailed kinetics of the LvsA association with the CV membrane to better understand the function of LvsA. Accelerated video microscopy of cells that co-expressed GFP-LvsA and RFP-Dajumin revealed that GFP-LvsA was only translocated into the CV membrane at the very last stage of discharge immediately before the CV bladder flattened against the plasma membrane (additional film S5). The period during which LvsA was associated with CVs was short (18±5 s (n = 30 cells)) as opposed to 57±18 s (n = 30 cells) between the time of localization of disgorgine on CVs and complete discharge of the vacuole. The complete cycle, from the onset of cvian growth to discharge, is ∼100±22 s (n = 30 cells). The kinetics of the lvsA association with the CV membrane coincide with a role of LvsA in maintaining membrane integrity during the fusion process.
This localization of the LvsA CV membrane is independent of disgorgine (Figure 6C). Surprisingly, LvsD did not localize on CV membranes and was still cytosolic in all cell lines tested, whether the cells were in isotonic or hypotonic media (data not shown). Amoebae survive in hypotonic environments because they have contractile vacuoles to pump excess water out of the cell. To understand the differences between CVs with and without disgorgin, we placed the cells that co-express GFP disgorgin and RFP dajumin in the water. RFP-Dajumin evenly marked all vacuoles and tubules, and we observed the loading and unloading of these CVs (Figure 3C). However, we found that Disgorgin was not located on the CV membrane until the late loading phase. At the same time as the recruitment of other disgorgins, the CV stopped growing, became spherical and initiated its dismissal (Figure 3C). We found that, unlike dajumine, which remained on the CV patch, Disgorgin dissociated from the membrane immediately after vacuol fusion (Figure 3C; 60, 72, 120 and 132 s timings). The kinetics of the association of disgorgine with CVs is consistent with the potential role of Disgorgin in regulating the fusion of the CV plasma membrane. Full expression of disgorgin or disgorgin without the F-Box (DisgorginΔF-Box; Figure 1A) in the cells Disgorgin− completes the large phenotype of vacuol and does not cause an overexpression phenotype when expressed in wild-type cells (Figure 1D, data not shown). . .
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