Acidocalcisomes Are Functionally Linked to the Contractile Vacuole of Dictyostelium discoideum (original) (raw)
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Folia Biologica, 2007
Cell movement in the amoebae Dictyostelium discoideum has been examined in media differing in monovalent cation concentration (i.e. Na + and K +). Under isotonic or even slightly hypertonic conditions, the cells move equally well in solutions in which either potassium or sodium ions dominate. However, in strongly hypertonic solutions the amoebae showed motility in a 2% potassium chloride solution, but remained motionless in a hypertonic 2% sodium chloride solution. This inhibition of D. discoideum amoebae movement in a hypertonic sodium chloride solution was fully reversible. Such behaviour corresponds to that of plant, fungi, and some invertebrate animal cells rather than protozoan or vertebrate cells. These observations suggest that studies using D. discoideum as a model for cell motility in vertebrate animal tissue cells should be considered with caution, and would seem to confirm the classification of cellular slime moulds as related rather to Fungi than to Protista. This also shows that the cell membrane models should consider the asymmetry in sodium /potassium ion concentrations found in vertebrate animal cells as one of various possibilities.
Identification of organelles in bacteria similar to acidocalcisomes of unicellular eukaryotes
Journal of Biological Chemistry, 2003
Acidocalcisomes are acidic calcium storage compartments described in several unicellular eukaryotes, including trypanosomatid and apicomplexan parasites, algae, and slime molds. In this work, we report that the volutin granules of Agrobacterium tumefaciens possess properties similar to the acidocalcisomes. Transmission electron microscopy revealed that each intracellular granule was surrounded by a membrane. X-ray microanalysis of the volutin granules showed large amounts of phosphorus, magnesium, potassium, and calcium. Calcium in the volutin granules increased when the bacteria were incubated at high extracellular calcium concentration. Immunofluorescence and immunoelectron microscopy, using antisera raised against peptide sequences conserved in the A. tumefaciens proton pyrophosphatase, indicated localization in intracellular vacuoles. Purification of the volutin granules using iodixanol density gradients indicated a preferential localization of the pyrophosphatase activity in addition to high concentrations of phosphate, pyrophosphate, short-and long-chain polyphosphate, but lack of markers of the plasma membrane. The pyrophosphatase activity was potassium-insensitive and inhibited by the pyrophosphate analogs, amynomethylenediphosphonate and imidodiphosphate, by dicyclohexylcarbodiimide, and by the thiol reagent N-ethylmaleimide. Polyphosphate was also localized to the volutin granules by 4,6-diamino-2-phenylindole staining. The organelles were acidic, as demonstrated by staining with LysoSensor blue DND-167, a dye especially used to detect very acidic compartments in cells, and cycloprodigiosin, a compound isolated from a marine bacterium that has been shown to uncouple proton pyrophosphatase activity acting as a chloride/proton symport. The results suggest that acidocalcisomes arose before the prokaryotic and eukaryotic lineages diverged.
Ca2+ chemotaxis in Dictyostelium discoideum
Journal of Cell Science, 2010
Using a newly developed microfluidic chamber, we have demonstrated in vitro that Ca 2+ functions as a chemoattractant of aggregationcompetent Dictyostelium discoideum amoebae, that parallel spatial gradients of cAMP and Ca 2+ are more effective than either alone, and that cAMP functions as a stronger chemoattractant than Ca 2+ . Effective Ca 2+ gradients are extremely steep compared with effective cAMP gradients. This presents a paradox because there is no indication to date that steep Ca 2+ gradients are generated in aggregation territories. However, given that Ca 2+ chemotaxis is co-acquired with cAMP chemotaxis during development, we speculate on the role that Ca 2+ chemotaxis might have and the possibility that steep, transient Ca 2+ gradients are generated during natural aggregation in the interstitial regions between cells. (2006). Stem cell engraftment at the endosteal niche is specified by the calcium-sensing receptor. Nature 439, 599-603. Aguirre, A., Gonzalez, A., Planell, J. and Engel, E. (2010). Extracellular calcium modulates in vitro bone marrow-derived Flk-1+ CD34+ progenitor cell chemotaxis and differentiation through a calcium-sensing receptor. Biochem. Biophys. Res. Commun. 393, 156-161. Alcantara, F. and Monk, M. (1974). Signal propagation during aggregation in the slime mould Dictyostelium discoideum. J. Gen. Microbiol. 85, 321-334. Andrew, N. and Insall, R. (2007). Chemotaxis in shallow gradients is mediated independently of PtdIns 3-kinase by biased choices between random protrusions. Nat. Cell Biol. 9, 193-200. Bandyopadhyay, S., Jeong, K. H., Hansen, J. T., Vassilev, P. M., Brown, E. M. and Chattopadhyay, N. (2007). Calcium-sensing receptor stimulates secretion of an interferon-gamma-induced monokine (CXCL10) and monocyte chemoattractant protein-3 in immortalized GnRH neurons. J. Neurosci. Res. 85, 882-895. Bangerth, F. (1979). Calcium-related physiological disorders of plants. Annu. Rev. Phytopathol. 17, 97-122. Bohme, R., Bumann, J., Aeckerle, S. and Malchow, D. (1987). A high-affinity plasma membrane Ca 2+ -ATPase in Dictyostelium discoideum: its relation to cAMP-induced Ca 2+ fluxes. Biochim. Biophys. Acta 904, 125-130. Bonner, J. T. (1949). The demonstration of acrasin in the later stages of the development of the slime mold Dictyostelium discoideum. J. Exp. Zool. 110, 259-271. Boyden, S. (1962). The chemotactic effect of mixtures of antibody and antigen on polymorphonuclear leucocytes.
Involvement of intracellular calcium in protein secretion in Dictyostelium discoideum
Journal of Cell Science, 1992
We reported previously that Ca2+ depletion of Dictyostelium discoideum cells severely inhibits extracellular cyclic nucleotide phosphodiesterase (PD) synthesis at a post-transcriptional step. In this study, further experiments were performed to learn more about the nature of this phenomenon. Examination of the polysomal distribution of PD transcripts in control cells and in cells depleted of Ca2+ by incubation with EGTA and A23187 (EA) suggested that inhibition of PD production does not involve translational control. Kinetic analysis of this inhibitory process revealed that soluble, intracellular PD activity, synthesized from either the 2.4 or 1.9 kb PD mRNA, decreased very rapidly upon addition of EA. Furthermore, this decrease in activity was accompanied by the preferential loss of PD-related polypeptides, indicating a proteolytic event. EA-induced PD degradation required cellular energy and concomitant protein synthesis but was unaffected by most of the lysosomotropic agents tested. Therefore, PD proteolysis might not occur in the lysosome. In cell fractionation experiments, the EA-sensitive, intracellular PD activity comigrated with a rough ER marker in Percoll/KCl gradients. In addition to its effect on the PD, EA were also observed to inhibit production and rapidly lower the intracellular levels of another secreted glycoprotein, the PD inhibitor. Together, these results suggest that depletion of some intracellular Ca2+ store(s) in Dictyostelium, possibly the ER, disrupts the normal function of the secretory pathway, resulting in selective degradation of certain proteins.
The Role of Acidocalcisomes in Parasitic Protists
Journal of Eukaryotic Microbiology, 2009
Acidocalcisomes of Trypanosoma cruzi are acidic calcium-containing organelles rich in phosphorus in the form of pyrophosphate (PP i ) and polyphosphate (poly P). Acidification of the organelles is driven by vacuolar proton pumps, one of which, the vacuolar proton pyrophosphatase (V-H + -PPase), is absent in mammalian cells. A Ca 2+ -ATPase is involved in calcium uptake and an aquaporin is important for water transport. Enzymes involved in the synthesis and degradation of PPi and poly P are present within the organelle. Acidocalcisomes function as storage sites for cations and phosphorus, participate in PP i and poly P metabolism, and volume regulation, and are essential for virulence. A signaling pathway involving cyclic AMP (cAMP) generation is important for fusion of acidocalcisomes to the contractile vacuole complex (CVC), transference of aquaporin, and volume regulation. This pathway is an excellent target for chemotherapy as shown by the effects of phosphodiesterase C (PDEC) inhibitors on parasite survival.
Journal of Eukaryotic Microbiology, 2011
The structural organization of parasites has been the subject of investigation by many groups and has lead to the identification of structures and metabolic pathways that may represent targets for anti-parasitic drugs. A specific group of organelles named acidocalcisomes has been identified in a number of organisms, including the apicomplexan parasites such as Toxoplasma and Plasmodium, where they have been shown to be involved in cation homeostasis, polyphosphate metabolism, and osmoregulation. Their structural counterparts in the apicomplexan parasite Eimeria have not been fully characterized. In this work, the ultrastructural and chemical properties of acidocalcisomes in Eimeria were characterized. Electron microscopy analysis of Eimeria parasites showed the dense organelles called volutin granules similar to acidocalcisomes. Immunolocalization of the vacuolar proton pyrophosphatase, considered as a marker for acidocalcisomes, showed labeling in vesicles of size and distribution similar to the dense organelles seen by electron microscopy. Spectrophotometric measurements of the kinetics of proton uptake showed a vacuolar proton pyrophosphatase activity. X-ray mapping revealed significant amounts of Na, Mg, P, K, Ca, and Zn in their matrix. The results suggest that volutin granules of Eimeria parasites are acidic, dense organelles, and possess structural and chemical properties analogous to those of other acidocalcisomes, suggesting a similar functional role in these parasites.
Separation and properties of prestalk and prespore cells of Dictyostelium discoideum
Experimental Cell Research, 1981
We describe a method of separating prestalk and prespore cells of Dictyostelium discoideum slugs using a self-generating Percoll gradient. This method gives quantitative recovery of cells and good purity. Separated prestalk and prespore cells possess different levels of the enzymes UDP galactose : polysaccharide transferase, CAMP phosphodiesterase and glycogen phosphorylase. We have used this method, as well as mechanical dissection of slugs, to examine the fate of separated prestalk and prespore cells in Dicfyostelium strains that are able to give rise to mature stalk and spore cells in cell monolayers. The results from such experiments provide direct evidence that prestalk and prespore cells from the migrating slug stage are programmed to differentiate into stalk and spore cells respectively.
Vacuolar H + -ATPase and weak base action in Dictyostelium
Molecular Microbiology, 1996
Amoebae of Dictyostelium discoideum release ammonia during development, and the accumulation of this weak base is believed to be responsible for inhibiting fruiting-body formation and switching aggregates into migrating slugs. Exposure to weak bases can also inhibit aggregation and cell-type specific gene expression. The pathway by which weak bases influence development is not understood. We show here that the development of a set of mutants defective in acidification of intracellular acidic compartments is abnormally sensitive to inhibition by weak bases. Moreover even in the absence of added weak bases these mutants are delayed in aggregation and have a protracted migratory phase. The same behaviour is observed in transformants harbouring an antisense construct for one of the vacuolar H'-ATPase subunits. These results support the idea that weak bases exert their effects by inhibiting acidification of an intracellular acidic compartment.