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PubMedDate not supplied

[Effect of actinomycin D on salinity acclimation of Paramecium calkinsi (Ciliophora, Peniculia)].

Salinity acclimation of euryhaline ciliate Paramecium calkinsi evaluated by swimming velocity was over in 4-5 days. In two series of the main experiment, the acclimation was over in two and three days when actinomycin D (an inhibitor of DNA-dependent RNA polymerase) was added after various periods of the ciliate incubation at a changed salinity (0, 3, 6, and 24 h). The obtained data indicate that the inhibiting effect of actinomycin D on salinity acclimation of P. calkinsi is manifested only within the first day of the ciliate incubation in an altered environment; by the end of this period their swimming velocity was similar to the control one (acclimation in the absence of the inhibitor). Thus, expression of genes associated with salinity acclimation in paramecium largely completes within the first day, while the acclimation continues for at least 4-5 days. We propose that the acclimation-competent species possess a correspondingly rich genetic program.

Acclimatization↗
PubMed2004

Cell proliferation of Paramecium tetraurelia under clinorotation.

It has been reported that Paramecium proliferates faster under microgravity in space, and slower under hypergravity (Kato et al., 2003). Effects of gravity on cell proliferation could be discussed in terms of energetics of swimming. Because of the characteristics of 'gravikinesis' as well as 'gravitaxis', Paramecium would decrease the energy expenditure under microgravity and increase it under hypergravity. The larger stock of energy would enhance the proliferation under microgravity. In order to simulate the effect of microgravity, we investigated the proliferation under clinorotation. When cells were rotated at 2.5 rpm, the proliferation rate decreased. Similar but less pronounced decrease was also found under low speed clinorotation (0.2 rpm).

Animals↗
PubMed1990

High yield isolation procedure for intact secretory organelles (trichocysts) from different Paramecium tetraurelia strains.

The secretory organelles (trichocysts) of Paramecium tetraurelia, wild type (7S) and some mutant cells (non-discharge strains, nd), were isolated with their intact membranes using a self-generating Percoll gradient. This has become possible by detaching trichocysts before (nd) or during (7S) homogenization from the cell membrane, where they are normally firmly attached. Membrane integrity was tested in several ways (acridine orange trapping, Ca2+ insensitivity). Measurement of several marker enzymes, DNA and tubulin content showed considerable enrichment of trichocysts over other subcellular constituents. Calculation of the yield showed that more than 50% of the whole amount of trichocysts per cell can be isolated. However, with the method described trichocysts can be isolated in an intact state from a variety of Paramecium tetraurelia strains so that they now have become amenable to further analyses.

Animals↗
PubMed1978

Physiological consequences of mitochondrial antibiotic-resistant mutations in Paramecium: growth-rates, cytochromic defects and cyanide-insensitive respiration of mutant and erythromycin-treated wild-type strains.

A set of mitochondrial antibiotic-resistant mutants of Paramecium have been analyzed with respect to their growth-rates, cytochromic content and respiratory properties. The mutants could be arranged in a continuous series ranging from strains equivalent to wild-type to severely affected ones; affected strains display longer generation times, reduced amount of cytochrome oxidase and very high levels of cyanideinsensitive respiration. Perfect phenocopies of the mutants were obtained by treating wild-type cells with low concentrations of erythromycin suggesting that the mutations exert their pleiotropic effect by perturbating mitochondria protein synthesis in agreement with the idea that these mutations affect the mitochondrial ribosomes. In the mitochondria of some of the mutants, electrons can be channelled with equal efficiency into the "classical" cyanide-sensitive pathway and the alternate cyanide insensitive (and SHAM-sensitive) one, providing direct demonstration of the branching of these two respiratory pathways. In the absence of any added inhibitor, however, electrons tend to be channelled in the cyanide-sensitive pathway. All the physiological data fit perfectly the genetic data concerning the "stability" of the various mutations in "mixed mitochondrial populations", i.e., markers that were known to be strongly counter-selected with respect to wild-type in such populations correspond to severely affected strains, while markers that were known to be "stable" correspond to "healthy" strains. A more quantitative analysis of the data shows that that there is little or no "complementation" between wild-type and mutated mitochondria in mixed cells indicating a high extent of functional autonomy of mitochondria in Paramecium.

Animals↗
PubMed1989

Purification and characterization of a calcium-dependent ATPase from Paramecium tetraurelia.

We report the purification of a CaATPase of high specific activity from Paramecium tetraurelia. The enzyme is preferentially released into solution upon deciliation of cells by a Ca2+ shock procedure. Purification by ion exchange and gel filtration chromatography yields major peptides of 68 and 53 kDa and a minor peptide of 58 kDa, as determined by electrophoresis on sodium dodecyl sulfate polyacrylamide gels. These three peptides yield similar proteolytic peptide maps. Rabbit antisera to the purified enzyme inhibit enzyme activity and specifically label 68- and 53-kDa bands on nitrocellulose blots of the deciliation supernatant from which the enzyme is isolated. Concanavalin A-Sepharose precipitates about 60% of ATPase activity; only the 53-kDa band binds concanavalin A on nitrocellulose blots. The purified enzyme has a specific activity of 620 +/- 70 mumol/min/mg with ATP as substrate in the presence of Ca2+, which is required for enzyme activity. As substrates, ATP and GTP are strongly preferred to UTP and CTP. The Km for ATP in the presence of 3 mM Ca2+ is approximately 20 microM. Enzyme activity is strongly inhibited by the calmodulin antagonists trifluoperazine, fluphenazine, W7, and calmidazolium. However, calmodulin is not associated with the purified enzyme, based on the enzyme's inability to bind anti-calmodulin antibodies or to stimulate brain phosphodiesterase. The intracellular origin of this ATPase, its possible function, and its relationship to several other ATPases of Paramecium are discussed.

Adenosine Triphosphate↗
PubMed1989

Cytotoxicity of certain organic solvents and organophosphorus insecticides to the ciliated protozoan Paramecium caudatum.

Responses of Paramecium caudatum, a ciliated protozoan, to acute exposures of certain organic solvents and organophosphorus insecticides (OPI) were studied by determining their lethal concentration (10 min-LC100) and median lethal concentration (4 h-LC50). The solvents and OPI evoked a distinct sequence of responses. Among the five solvents tested, acetone proved most toxic [LC-2.9% and LC50-0.68% (v/v)], while dimethyl sulphoxide (DMSO) showed least toxicity [LC-11.0% and LC50-3.16% (v/v)]. The order of toxicity of solvents was: acetone greater than ethanol greater than methanol greater than N, N-dimethylformamide greater than dimethylsulphoxide. The LC values of six OPI dissolved in either acetone or DMSO indicated that they were more toxic when dissolved in acetone and least toxic in DMSO. Among the OPI, bromophos proved most toxic (LC-10 ppm) while malathion showed least toxicity (LC-200 ppm) in DMSO. The order of toxicity of OPI was: bromophos greater than pirimiphos-methyl greater than parathion methyl greater than dichlorvos greater than fenitrothion greater than malathion. The 4 h-LC50 values computed for bromophos and malathion (dissolved in DMSO) were 575 ppb and 19.9 ppm, respectively, indicating the high susceptibility of P. caudatum to bromophos. The results indicate that the Paramecium toxicity assay could be used as a complementary system to rapidly elucidate the cytotoxic potential of compounds.

Animals↗
PubMedDate not supplied

[Paramecium test for toxic substances in the blood of humans exposed to conditions simulating weightlessness].

The Paramecium test was for the first time used to measure the build-up of toxic substances in the healthy man exposed to simulated weightlessness. The most distinct changes were seen on bed rest days 3 and 7-8 in all test subjects (41 subjects). After bed rest day 8 the parameter reached a plateau, the toxicity level stopped to increase, but the Paramecium test time remained shorter than normal. It can therefore be suggested that the prognosis of any disease that may develop in this situation will be worse. In view of this it is important to improve the prophylaxis and treatment of endogenous and exogenous intoxications during space flight and to provide active detoxication based on the purification of biological fluids by sorption.

Adult↗
PubMed1985

Modulation of the digestive lysosomal system in Paramecium caudatum. III. Morphological effects of cytochalasin B.

Morphological explanations for the effects of cytochalasin B (CB) on the digestive vacuole system of Paramecium caudatum were sought using electron-microscopic studies. Cytochalasin B (0.3 mM) essentially stopped vacuole release without stopping vacuole growth. Acidosome fusion with the new vacuoles was inhibited and was accompanied by the formation of discoidal vesicle-lined clumps of microfilament-like material next to these vacuoles. Vacuole membrane retrieval at the open cytoproct was inhibited and was accompanied by the loss of the microfilament network typical of this region. At 0.15 mM CB vacuole release and fusion of the acidosomes and lysosomes with the vacuoles were not stopped, but membrane retrieval at the cytoproct was affected. Cytochalasin B at the higher concentration also affected the movement of the horseradish peroxidase (HRP)-labeled vesicles away from the nascent vacuole. Membrane retrieval by the formation of tubules of 35 to 50 nm diameter from membrane of early and late vacuole stages was also affected by CB. It is conceivable that all of these effects were the result of the disruption of membrane associated actin networks. By contrast, several highly organized microfilamentous systems in the cortex and oral region of these cells appeared to be unaffected by CB or dimethyl sulfoxide (DMSO) at the concentrations used. These results show that CB-susceptible structures assume a key role in the normal functioning of the digestive vacuole system and in vacuole membrane retrieval and reutilization in Paramecium.

Animals↗
PubMed1979

The estimation of velocity distribution profile of Paramecium cytoplasmic streaming.

Cytoplasmic streaming of Paramecium bursaria and Paramecium tetraurelia was investigated by cinematographic techniques. Analysis of the records reveals the paraboidal character of the velocity distribution profiles in all arbitrarily chosen zones along the whole route of moving cytoplasm in the cell. According to the date obtained from cytoplasmic streaming analysis and food vacuole path, the geometry of the "channel" was described in terms of ellipsoid axes. Total volume changes of cytoplasm flowing through a given cross section in a given time unit were computed and no significant differences were found. The participation of a pressure gradient in motive force generation in cytoplasmic streaming is discussed.

Animals↗
PubMed1981

Ionic channels of Paramecium: from genetics and electrophysiology to biochemistry.

This paper reviews the combined genetical, electrophysiological and biochemical analysis of excitation that has been carried out in Paramecium. Paramecium cells display graded Ca++ action potentials in response to a variety of stimuli. These action potentials regulate the orientation of the ciliary beat hence the cell's swimming behaviour. A large array of mutants displaying altered behaviour have been isolated and mapped to over 20 loci. Detailed electrophysiological analyses have been carried out on several classes of mutants revealing defects in specific ion channels in some cases. Mutants have proven very useful to analyze channel properties, to unravel interactions between channels and to discover the function of these channels in a variety of cellular processes. Some important channels are located in the ciliary membranes and cilia as well as ciliary membranes can now be purified in high purity and reasonable yield. These fractions have been used recently in a variety of biochemical approaches to gain insight into the molecular components of the excitation machinery. Specific alterations in some minor membrane proteins have been found in two mutants as well as a specific defect in sphingolipids in a third mutant. Those alterations had to be distinguished from large scale variations in membrane proteins and lipids that occur in this organism in response to modifications in growth conditions. Several other recent biochemical developments are reviewed and the advantages as well as the difficulties of the genetic approach to the molecular study of biological processes are discussed.

Animals↗
PubMed1983

Phosphorylation of endogenous proteins of cilia from Paramecium tetraurelia in vitro.

Several endogenous substrate proteins of cilia from axenically grown Paramecium tetraurelia were phosphorylated in vitro by inherent protein kinases (PKs). Labeling was stimulated by cAMP and to a lesser extent by cGMP. ATP breakdown was most rapid in cilia and subciliary fractions. Using multiple substrate additions during incubations it was shown that phosphorylation was almost completed within 30 s. Very little dephosphorylation by phosphoprotein phosphatases occurred during 5 min of incubation. Proteins of molecular weight of 103 000 and 46 000 were shown to be particularly associated with axonemal structures of the cilia. No distinct differences in phosphorylation patterns were apparent in ciliary membrane vesicles of low and high buoyant density, which exhibit differential enzyme patterns. cAMP receptor proteins were identified by use of the photoaffinity label 8-azido-[32P]cAMP. Receptor proteins with apparent molecular weights of 43 000, 39 000, 37 000, 31 000 and 30 000 were probably related to the regulatory subunits of cAMP-dependent protein kinases as evidenced by inhibition of incorporation of the photoaffinity label by low concentrations of cAMP. Tagging of a protein of 85 000 molecular weight was specifically inhibited by cGMP, thus in all likelihood it corresponded to a cGMP-dependent protein kinase. Corresponding autophosphorylated protein bands were observed with gamma-[32P]ATP. A functional role for protein phosphorylation in cilia of Paramecium remains to be established.

Animals↗
PubMed1984

Paramecium mitochondrial genes. I. Small subunit rRNA gene sequence and microevolution.

The sequences of the small subunit mitochondrial rRNA genes from two divergent species of Paramecium (primaurelia and tetraurelia) were determined. The gene lies near the center of the linear mitochondrial genome, on the same strand as are all other currently identified genes. The sequences generally resemble their counterparts found in cytoplasmic, procaryotic, and other mitochondrial sources. The rDNA gene boundaries were located by nuclease S1 protection. Small subunit rDNA spans about 1680 nucleotides, including an extraneous 83-base pair sequence very near the 3' end which is unique to Paramecium mitochondria. This "insert" occurs at the apex of the highly variable in length penultimate helix, according to proposed models for small subunit rRNA secondary structure. A discontinuity occurs in isolated rRNA near the start of the insert, resulting in a stable 13 S RNA species and a small segment containing the remaining 3' portion of the gene. The overall rRNA gene sequence was 94% conserved between the two species, and the nucleotide differences consisted of 53% transitions, 37% transversions, and 9% insertions plus deletions. These substitutions were somewhat clustered, and the two most divergent regions coincided with the gene boundaries. The sequence was aligned with Escherichia coli 16 S rRNA for direct comparison of sequence and structure.

Animals↗
PubMed1984

Modulation of the digestive-lysosomal system in Paramecium caudatum. I. Effects of temperature.

The heterophagic pathway of the digestive-lysosomal system in axenically grown Paramecium caudatum is divisible into vacuole formation, vacuole acidification-condensation, lysosomal fusion-digestion and defecation. These four processes can be separated in time, thus permitting the study of the effects of temperature on each process. The optimal growth temperature for this cell was 27 degrees C. The rate of digestive vacuole (DV) formation at varying temperatures was represented by a skewed bell-shaped curve having an optimum between 28 and 30 degrees C. The time course for the acidification-condensation step was lengthened below 26 degrees C, but was not accelerated above this temperature. The rate but not the extent of vacuole condensation was decreased at 19 and 22 degrees C. Temperature increase above 22 degrees C shortened, slightly, the duration of the lysosomal fusion-digestion process, whereas below 22 degrees C small temperature decreases greatly extended this period. Within a given experiment the rates of defecation were proportional to temperatures above 17 degrees C. However, these rates varied widely among different experiments. Interestingly, the activation energies for both the formation and defecation processes averaged 19 kcal/mol. Furthermore, Paramecium appeared to readily adapt to environmental temperature changes, since the length of the processing periods and the rates of defecation were similar in cells with or without a 24 h acclimation. These results indicated that the four processes in the digestive cycle in P. caudatum are distinct but each is energy-dependent.

Animals↗
PubMed1983

Biochemical studies of the excitable membrane of Paramecium tetraurelia. VII. Sterols and other neutral lipids of cells and cilia.

The neutral lipid content of cells and cilia of Paramecium tetraurelia was determined as a function of growth stage and of growth medium composition. The major sterol(s) of deciliated cells and of cilia were the sterol provided in the growth medium (e.g., stigmasterol) and its 7-dehydro derivative. Body sterol esters and triglycerides accumulated during exponential cell growth and became depleted during stationary phase. Isolated cilia contained neither sterol esters nor triglycerides. The ratio of phospholipid to free sterol in cilia was constant (approximately 5:1) throughout exponential growth in axenic or bacterized medium, but the relative proportions of free sterol and 7-dehydrosterol varied with growth phase and with growth medium composition. Cholesterol did not support the growth of Paramecium, but was taken up rapidly into cells and cilia as the free sterol and its 7-dehydro derivative. The fatty acids of sterol esters and of triglycerides were similar to those of membrane phospholipids.

Animals↗
PubMed1983

Pattern determination and pattern regulation in Paramecium tetraurelia.

Pattern regulation was investigated in the progeny of laterally fused cells of Paramecium tetraurelia. The immediate progeny of such fused cells (doublets) reveal two sets of cortical organelles arranged roughly symmetrically. Doublets tend to transform gradually into cells with only one set of organelles (singlets). At least two different and mutually exclusive pathways of doublet-to-singlet transformation are reported. In intermediate stages of regulation the cortical areas bearing different cortical landmarks may be brought into an abnormal neighbourhood. Differentiated cortical bands of cortex, bearing organellar landmarks, are faithfully propagated even if they are improperly and asymmetrically located on the cell. The confrontation of such cortical bands may lead to the transient appearance of additional duplicated organelles. It is suggested that pattern regulation in Paramecium during doublet-to-singlet transformation results from at least three factors: the regression of some part of the cortical areas, the interaction of the juxtaposed parts remaining and the slow regulatory shift of positions of the cortical structures.

Animals↗
PubMed1982

Protein phosphorylation/dephosphorylation and stimulus-secretion coupling in wild type and mutant Paramecium.

Axenic cultures of Paramecium tetraurelia take up 32Pi and phosphorylate a number of polypeptides as determined by autoradiography following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The most heavily labeled polypeptide has an apparent Mr of approximately 65,000. Wild type cells stimulated to secrete with picric acid, the standard secretagogue for these cells, show a marked reduction in labeling of the 65,000 Mr polypeptide. There is no change in the Coomassic blue staining protein pattern after addition of picric acid. Addition of picric acid to cells solubilized in sample buffer containing 10% sodium dodecyl sulfate, significantly lowers the pH but does not induce dephosphorylation of the 65,000 Mr polypeptide. Dephosphorylation of the 65,000 Mr polypeptide is further correlated with secretion in two types of experiments. 1) Preincubation of cells in Mg2+ (no added Ca2+) inhibits both secretion and dephosphorylation in response to picric acid. 2) A temperature-sensitive mutant, nd 9, when grown at 18 degrees C (permissive temperature) has the normal intramembrane particle array (rosette) at the secretory site and secretes and dephosphorylates the 65,000 Mr polypeptide in response to picric acid, but when grown at 27 degrees C (nonpermissive temperature) does not have assembled rosettes at the secretory site, and does not secrete nor dephosphorylate the 65,000 Mr polypeptide in response to picric acid. This represents the first correlation between a phosphoprotein and a physiological activity (secretion) in Paramecium. Our results show the presence of an in vivo stimulus-sensitive phosphoprotein of Mr 65,000 which appears related to Ca2+-mediated exocytosis. Inhibition of dephosphorylation occurs when secretion is blocked, either by Mg2+ or by a mutation affecting an intramembrane particle array, the rosette.

Animals↗
PubMed1981

Localization of actin in the cortex of Paramecium tetraurelia cells by immuno- and affinity-fluorescence microscopy.

The cortex of the ciliate Paramecium tetraurelia has been examined both with whole cells and with isolated surface complexes (pellicles) for the presence and distribution of actin, using direct fluorescence labeling with antibodies monospecific for Paramecium actin, with DNase I or with heavy meromyosin, all tagged with FITC as a fluorochrome. The results obtained with the three different methods were similar. Actin is concentrated in the cilia and--clearly visible only after deciliation--in the basal bodies. The docking site of secretory vesicles (trichocysts) onto the cell membrane does not display any selective actin content. This does not preclude the interaction with actin at other sites of the trichocysts, since diffuse fluorescence labeling in deeper layers of the cell did not allow us yet to analyze the distribution of actin throughout the cell body. However, our data would preclude any direct involvement of actin at the site of exocytotic membrane fusion proper.

Actins↗
PubMed1995

Cortical memory in Paramecium: a theoretical approach to the structural heredity.

Structural heredity is frequently encountered in ciliates. In particular, during division of Paramecium the cortical characteristics of the 2 daughter cells are inherited from those of the mother cell. Genesis of new structures is therefore only possible if there are pre-existing older structures. We attempted to simulate this phenomenon with a model already used to describe the structure and morphogenesis of the cortex of Paramecium. We show that to initiate spatial organization it is enough to choose special starting conditions which comprise the primer of the structure, even if highly localized and at a low concentration of morphogens. This property does not question the basic concepts of genomic heredity.

Animals↗