What Is the Main Function of Contractile Vacuole in Protozoa

In Chlamydomonas, two contractile vacuoles are located near the anterior end of the cell, that is, near the bases of the flagella. Chlamydomonas, with a diameter of 1.5 μm, includes the smallest known contractile vacuoles. Home / Cell Biology / What is the function of contractile vacuol in protozoa? What is contractile vacuole? Learn more about it! We know that GAP activity and membrane localization are necessary for disgorgin function, but we do not know the function of the F-box disgorgin domain. The loss of F-box does not affect the ability of overexpressed disgorgine to complete disgorgine phenotypes. We found two orthologs of Dictyostelium SKP1 in complexes containing disgorgines and showed that they interact with Disgorgin in an F-box dependent manner. F-box proteins and SKP proteins are components of the SCF-ubiquitin-E3 ligase complex (Skp-Cullin-F-Box). Disgorgin is most likely either a target of ubiquitination or it acts as an adapter to bring another protein into the SCF complex, which is then ubiquitinated. Alternatively, the Disgorgin F-Box domain can mediate the formation of a complex that does not contain CFS (Kipreos and Pagano, 2000). However, we have no direct evidence for either model and have not been able to demonstrate ubiquitination of disgorgine. The contractile vacuol remains firm or moves through the cytoplasm while being closely related to the endoplasm. In the broadest sense, the morphologies and behaviors of the CV differ from one organization to another.

The contractile vacuole complex (CV) is an osmoregulatory organelle of free amoebae and protozoa that controls intracellular fluid balance by accumulating and expelling excess water from the cell so that the cells can survive under hypotonic stress as in pond water. In the absence of a functional CV complex, cells cannot expel water, swell sharply and lyse. Extensive work has been done to characterize the properties and functions of the CV system. In dictyostelium, the CV system consists of tubules and vacuoles (or bubbles) that are interconvertible (Gerisch et al., 2002). Tubular structures act as collecting channels to accumulate excess water while bubbles fuse with the plasma membrane, allowing bladder contents to be released into the extracellular medium, thus expelling water from the cell body (Heuser et al., 1993). When the cells are in an isotonic medium, the CV system shows limited activity, but when the cells are placed in a hypotonic medium, the CV system is quickly activated: the bubbles fill with water and then fuse with the plasma membrane, discharging its contents (Heuser et al., 1993; Gabriel et al., 1999). Heuser (2006) proposed that the CV should not disappear during the discharge phase, but should collapse and flatten against the plasma membrane, thus preserving its various membrane components. It has been implied that azidocalciums act 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, azidocalciums empty their ionic content into the contractile vacuole, which increases the osmolarity of the vacuoles. [6] Disgorgin has some similarities with drainin, another regulator of CV discharge. Both contain areas of tuberculosis and the elimination of both proteins causes an increase in CV formation.

However, cell morphologies and CV flow differ in the two null strains. CVs are less enlarged and vacuol sizes are more uniform in disgorginous cells than in drainin cells (Figure 4A and Additional Figure S2A). Drainin cells are partially hypoosmotically sensitive and exhibit two types of abnormal discharge, one of which is similar to that observed in disgorginous cells (vacuole forms a bleb) (Becker et al., 1999). Although disgorginous cells do not have active CV plasma fusion as described above, cells can discharge and are insensitive to hypotonic stress (Figure 2C; Data not displayed). Try PMC Labs and let us know what you think. Find out more. Thus, it is quite clear that the contractile vacuole is the organelle that protects the cell from cell expansion and explosion due to too much water in the cytoplasm. It protects this by expelling excess water from the cell by contracting. If you look at a protozoan like Ameoaba, Paramecium, etc. under a microscope, then the contractile vacuol simply looks like a transparent, spacious, rounded cellular organelle, bound to the membrane and filled with water in the cell.

In Paramecium, you will observe a very complex and advanced contractile vacuole compared to other protozoa. Similar to other small GTPases, Rabs switches between GDP-related (inactive) and GTP -active) states. The hydrolysis of GTP is stimulated by GTPase activating proteins (RabGAPs), most of which contain a conserved catalytic tuberculosis domain (Tre/Bub2/Cdc16) (Bernards, 2003). By studying the regulation of the CV function of dictyostelium, we have identified a new Rab8A-GAP, disgorgin, which, together with drainin and its Rab11A regulator, controls CV discharge. Drainin and disgorgin/Rab8A are successively located in the late loading phase at the CV membrane and control different phases of the process. We show that two proteins of the BEACH family, LvsA and LvsD, exert an effect as suppressors or enhancers of disgorgine phenotypes to regulate CV formation. By studying the cellular phenotypes of different mutated strains and the genetic and biochemical interactions of different CV components, we provide new insights into the signaling pathways that regulate CV function and formation. Our results suggest that Disgorgin is a GAP for Rab8A, but not for other Rabs, who are known to locate in the RESUME, and that Disgorgin`s GAP activity is necessary for their function.

In addition, Rab8A and Disgorgin locate simultaneously on CV bulbs. We suggest that the fusion of the CV plasma membrane, which allows the flow of CV bladder contents into the extracellular medium, is mediated by disgorgin and Rab8A and this event requires the Rab8A cycle from the GTP state to the GDP-related state. In yeast, Sec4 (a counterpart of Rab8) binds to Sro7p, which interacts with the t-SNARE sec9p protein and is involved in membrane fusion (Grosshans et al., 2006a). Another Sec4 effector, Sec15p, is a component of the exocyst that mediates the docking and fusion of exocytic vesicles to the plasma membrane and interacts with the Sro7p signaling pathway (Guo et al., 1999; Grosshans et al., 2006a). Rab8A may have a similar function in the fusion of the CV plasma membrane into dictyostelium. Our proposed role for Rab8A results from its co-location with Disgorgin and the fact that Rab8A-GTP is a Disgorgin substrate. We have not been able to obtain definitive genetic evidence that Rab8A is needed for this process because we have not been able to produce a rab8A-free strain, most likely because Rab8A is essential for growth. Similarly, we could not test a dependency for the localization of disgorgin on Rab8A. GRAPH 49.3. Contractile vacuol complex in the parameterizing showing ampoules (amp), a collecting channel (cc), a contractile vacuole (cv), a pore (pv), spongiomatous tubules (sp), fluid segregation organelles (fs) and microtubular ligaments (mtr).

In other protozoan species such as Tokophrya, as in Suctoria and Ciliata in general, the contractile vacuole has a permanent channel that connects it to the outside world. The canal seems to have a very elaborate structure and consists of a pore, a canal and a narrow tubule in a papilla that protrudes well into the cavity of the contractile vacuole. TPS or TPS fused proteins were produced in BL21 (DE3) bacteria and purified for glutathione sepharis (Amersham Biosciences) according to the manufacturer`s instructions. The GST reduction test was a modified form of the ras bond test described above (Sasaki et al., 2004). In short, the cell extract of Ax2 cells expressing disgorgine v5 or drainin v5 was incubated with 10 μg of TPS or GST-Rabs on glutathione agarose beads at 4°C for 1 h. The pearls were washed three times. The proteins were separated on an SDS-PAGE gel and immunopopulated with the anti-v5 antibody (sigma). We performed the GAP test with or without 250 nM GAN using the EnzChek® phosphate dosing kit (Invitrogen) according to the protocol described above (Pan et al., 2006). Absorption at 360 nm was monitored with the SpectraMax Plus384 microtiter plate detection system using SoftMax Pro 4.3LS (Molecular Devices). Three independent experiments were conducted. The kinetic data was analyzed by simultaneously adapting it with MATLAB (MachWorks Inc.) to the first-order Michaelis-Menten model function. To identify the properties of large vacuoles, we labeled disgorginous cells with markers for different types of organelles: TRITC-dextran (endosomes), lysotrackers (lysosomes) or RFP-dajumin (CV system) (Gabriel et al., 1999; Insall et al., 2001).

Dajumine RFP, but not the other markers, clearly marked the large vacuol structures corresponding to those observed in phase contrast microscopy, suggesting that the large vacuoles in the disgorginous cells are enlarged CVs (Figure 2A and Additional Film S1). . . . .

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