Physiological responses of Zygosaccharomyces rouxii to osmotic stress

When cell suspensions of Zygosaccharomyces rouxii were subjected to osmotic shock with NaCl, the cell volume decreased sharply and plasmolysis was observed. The cell subsequently recovered and volumes similar to those of cells growing at the respective water activity (a_w) values were found. Cycloheximide prevented cell recovery, indicating the involvement of protein synthesis in the recovery process. The intracellular glycerol concentration of Z. rouxii incubated in the presence of [¹⁴C]glycerol increased from 13 to 96 mmol/l during the initial 20 min after an upshock from 0.998 a_w to 0.96 a_w. All the intracellular glycerol was labelled and therefore derived from the medium. Labelled glycerol was subsequently utilized and replaced by unlabelled glycerol produced by the cell within 90 min. The initial increase in glycerol concentration following the upshock was confirmed by ¹³C-nuclear magnetic resonance (NMR) spectroscopic studies of cell extracts. The combined dihydroxyacetone and dihydroxyacetone phosphate concentrations fluctuated during this period, whereas glycerol-3-phosphate initially increased and then remained constant. This indicates that the production of glycerol is regulated. Decreases in ATP and polyphosphate levels were observed following osmotic upshock and may reflect a greater demand for ATP during the period of adjustment to decreased a_w. The changes in cell volume and in ATP concentration following osmotic upshock may serve as osmoregulatory signals in Z. rouxii, as suggested previously for other microorganisms. Correspondence to: S. G. Kilian     

Lipid composition of the nitrogen starved green alga Neochloris oleoabundans

Neochloris oleoabundans has been cultivated in mineral medium deficient in nitrogen. The yield of lipids was 35–54% of cell dry weight. Triglycerides comprised 80% of the total lipids. Aliphatic hydrocarbons, sterols, pigments, glycolipids and phospholipids comprised the remaining lipid fraction. Saturated, monounsaturated and diunsaturated octodecanoic acid represented approximately one-half of the total fatty acids.     

Neochloris oleoabundans oil production in an outdoor tubular photobioreactor at pilot scale

Journal of Applied Phycology - Oil production was tested with Neochloris oleoabundans in a 6 m3, horizontal soft sleeve tubular reactor from 22 October to 7 November in Matalascañas,...     

Division in Isolated Generative Cells of Allemanda neriilolia

An osmotic shock method of isolating generative cells from Allemanda neriifolia was described. Fresh pollen grains were first placed ill a Brewbaker and Kwack's medium (BK medium) containing 50% sucrose, incubated at 28℃ for 2 hours. During this incubation period pollen grains germinated and produced pollen tubes measuring about 200 μm long. After this initial incubation period, a fixed amount of BK medium without sucrose was added thus diluting the original medium to a sucrose concentration of 30% – an optimum concentration for generative cell growth. The addition, of the BK medium without sucrose brought about an osmotic shock effect on the pollen tubes and caused most of the tubes to burst at the tip region thus releasing the contents together with the generative cell from the tube into the 30% sucrose + BK medium. After isolation and filtering into a fresh lot of 30% sucrose + BK medium, generative cells changed from spindle into spherical-shaped cells. In the 30% sucrose + BK medium, the generative cells divided and within a short period of 3 to 5 hours a laege number of cells at various stages of mitosis was obtained.     

Dynamic changes of protoplasmic volume and of fine structure during osmotic adaptation in the intertidal red alga Porphyra umbilicalis

Abstract Cells of Porphyra umbilicalis show a biphasic osmotic regulatory response. After transfer from 1 × into 3.5 × artificial seawater medium (osmotic upshock) the protoplasts shrink rapidly, then recover their original size within 3 h and continue to increase over the next 14 d. After retransfer from 3.5 × into 1 × medium (osmotic downshock) the protoplasts swell immediately and then adjust to the normal size in 1 x medium. Parallel to the shrinkage of the protoplasts after osmotic upshocks the plasmalemma at first gets a wavy surface which becomes smooth again during the following adaptation process. Immediately after osmotic upshock the vacuolar volume increases and it decreases drastically after osmotic downshock. After osmotic upshocks only small vacuoles are present at first. In adapted plants, however, the vacuolar system is mainly composed of large vacuoles. The volume of the protoplasm without the vacuoles is regulated osmotically. Parallel to the increase in the vacuolar volume after osmotic upshocks there is an increase in the number of intramembraneous particles on the PF-face of the tonoplast. This high value is reduced rapidly to the original number after osmotic downshock. The findings are discussed in relation to the function of the vacuoles as compartments of inorganic ion accumulation during osmotic adaptation.     

Transmembrane Protein (Perfringolysin O) Association with Ordered Membrane Domains (Rafts) Depends Upon the Raft-Associating Properties of Protein-Bound Sterol

The concentration of chemicals inside the bacterial cytoplasm generates an osmotic pressure, termed turgor, which inflates the cell and is necessary for cell growth and survival. In Escherichia coli, a sudden increase in external concentration causes a pressure drop across the cell envelope that drives changes in cell shape, such as plasmolysis, where the inner and outer membranes separate. Here, we use fluorescence imaging of single cells during hyperosmotic shock with a time resolution on the order of seconds to examine the response of cells to a range of different conditions. We show that shock using an outer-membrane impermeable solute results in total cell volume reduction with no plasmolysis, whereas a shock caused by outer-membrane permeable ions causes plasmolysis immediately upon shock. Slowly permeable solutes, such as sucrose, which cross the membrane in minutes, cause plasmolysis to occur gradually as the chemical potential equilibrates. In addition, we quantify the detailed morphological changes to cell shape during osmotic shock. Nonplasmolyzed cells shrink in length with an additional lateral size reduction as the magnitude of the shock increases. Quickly plasmolyzing cells shrink largely at the poles, whereas gradually plasmolyzing cells invaginate along the cell cylinder. Our results give a comprehensive picture of the initial response of E. coli to hyperosmotic shock and offer explanations for seemingly opposing results that have been reported previously.     

Osmotic relations during elongation growth in hypocotyls of Helianthus annum L.

The relationship between growth, change in cell osmotic pressure and accumulation of osmotic solutes was investigated in hypocotyls of sunflower (Helianthus annum L.) seedlings. During growth in darkness the osmotic pressure decreased by 50% between days 2 and 6 after sowing. After irradiation of dark-grown seedlings with continuous white light (WL) an inhibition of hypocotyl growth was measured, but the osmotic pressure of the growing cells was not lower than in the dark-grown control. Growth in darkness and after WL irradiation was accompanied by an increase in the amount of osmotic substances (soluble sugars) which was proportional to the increase in length of the organ. During growth in continuous WL the cell osmotic pressure decreased by 45 % between days 2 and 6 after sowing. The transfer of WL-grown seedlings to darkness (“re-etiolation”) resulted in a rapid acceleration of hypocotyl growth, but the cell osmotic pressure was the same as that of the WL grown control. Growth in continuous WL was accompanied by a corresponding accumulation of osmotic substances (soluble sugars). The transition from WL to darkness resulted in an enhanced accumulation of osmotica and an increase in cell-wall extensibility. The results indicate that the relative maintenance of cell osmotic pressure during rapid hypocotyl growth in darkness is caused by an enhanced accumulation of soluble sugars into the growing cells of the organ.     

Osmotic responses of unicellular blue-green algae (cyanobacteria): changes in cell volume and intracellular solute levels in response to hyperosmotic treatment

Abstract Changes in cell volume and solute content upon hyperosmotic shock have been studied for six unicellular blue-green algae (cyanobacteria): Synechococcus PCC 6301, PCC 6311; Synechocystis PCC 6702, PCC 6714, PCC 6803 and PCC 7008. The extent of change in volume was shown to be dependent upon the solute used to establish the osmotic gradient, with cells in NaCl showing a reduced shrinkage when compared to cells in media containing added sorbitol and sucrose. Uptake of extracellular solutes during hyperosmotic shock was observed in Synechocystis PCC 6714, with maximum accumulation of external solutes in NaCl and minimum solute uptake in sucrose solutions. Conversely, solute loss from the cells (K⁺ and amino acids) was greatest in sucrose-containing media and least in NaCl. The results show that these blue-green algae do not behave as ‘ideal osmometers’ in media of high osmotic strength. It is proposed that short-term changes in plasmalemma permeability in these organisms may be due to transient membrane instability resulting from osmotic imbalance between the cell and its surrounding fluid at the onset of hyperosmotic shock.     

Plasmolysis and Cell Shape Depend on Solute Outer-Membrane Permeability during Hyperosmotic Shock in E. coli

The concentration of chemicals inside the bacterial cytoplasm generates an osmotic pressure, termed turgor, which inflates the cell and is necessary for cell growth and survival. In Escherichia coli, a sudden increase in external concentration causes a pressure drop across the cell envelope that drives changes in cell shape, such as plasmolysis, where the inner and outer membranes separate. Here, we use fluorescence imaging of single cells during hyperosmotic shock with a time resolution on the order of seconds to examine the response of cells to a range of different conditions. We show that shock using an outer-membrane impermeable solute results in total cell volume reduction with no plasmolysis, whereas a shock caused by outer-membrane permeable ions causes plasmolysis immediately upon shock. Slowly permeable solutes, such as sucrose, which cross the membrane in minutes, cause plasmolysis to occur gradually as the chemical potential equilibrates. In addition, we quantify the detailed morphological changes to cell shape during osmotic shock. Nonplasmolyzed cells shrink in length with an additional lateral size reduction as the magnitude of the shock increases. Quickly plasmolyzing cells shrink largely at the poles, whereas gradually plasmolyzing cells invaginate along the cell cylinder. Our results give a comprehensive picture of the initial response of E. coli to hyperosmotic shock and offer explanations for seemingly opposing results that have been reported previously.     

Responses to Water Stress in Adapted and Unadapted Carrot Cell Suspension Cultures

The responses of suspension-cultured cells of carrot to polyethyleneglycol (PEG)-induced water stress were studied after transferto culture medium containing PEG at concentrations between 0%and 25%. Growth characteristics, cellular osmotic potentialand organic solute concentration changes were followed in unadaptedcells and in cell lines adapted to growth in various PEG concentrations.A decline in fresh and dry weight increase occurred in unadaptedcells with decreasing water potential, while dry weight gainwas unaffected in adapted lines. Substantial osmotic adjustmentwas observed in adapted lines, due mainly to increased glucose,fructose and sucrose. Proline concentration increased up to40-fold in adapted and 12-fold in unadapted cells and otheramino acids including alanine, histidine and arginine showedsimilar, though smaller, responses. Polyamines and glycinebetainedid not increase significantly in either adapted or unadaptedcells. Changes leading to long-term adaptation to water stressare discussed in relation to short-term stress—shock responses. Key words: Water stress, cell culture, Daucus carota, osmotic adaptation, solute accumulation     

Cell Division of Escherichia coli: Control by Membrane Organization

Cells of certain strains of Escherichia coli, after transfer from 37 to 45 C and incubation for 16 min, were observed to swell and subsequently divide synchronously. This swelling and the resulting stretching of the membrane are proposed to be the basis for the synchronous division. Four lines of evidence support this hypothesis. First, osmotic protection by the addition of either sodium chloride or sucrose at the time of heat shock prevents both swelling and synchrony. Second, a mutant neither swelled nor divided synchronously after heat shock. Third, cells grown for several generations with 10% sucrose in the medium swelled and divided synchronously upon transfer to medium without sucrose. Fourth, the mutant not synchronized by heat shock also swelled and underwent synchronous division after the osmotic shift. A tentative model is suggested for the normal control of division, based on membrane configuration at the septation site.     

Vesicle formation in the membrane of onion cells (Allium cepa) during rapid osmotic dehydration

Background and Aims

Optimization of osmotic dehydration in different plant cells has been investigated through the variation of parameters such as the nature of the sugar used, the concentration of osmotic solutions and the processing time. In micro-organisms such as the yeast, Saccharomyces cerevisiae, the exposure of a cell to a slow increase in osmotic pressure preserves cell viability after rehydration, while sudden dehydration involves a lower rate of cell viability, which could be due to membrane vesiculation. The aim of this work is to study cytoplasmic vesicle formation in onion epidermal cells (Allium cepa) as a function of the kinetics of osmotic pressure variation in the external medium.

Methods

Onion epidermal cells were submitted either to an osmotic shock or to a progressive osmotic shift from an osmotic pressure of 2 to 24 MPa to induce plasmolysis. After 30 min in the treatment solution, deplasmolysis was carried out. Cells were observed by microscopy during the whole cycle of dehydration–rehydration.

Key Results

The application of an osmotic shock to onion cells, from an initial osmotic pressure of 2 MPa to a final one of 24 MPa for <1 s, led to the formation of numerous exocytotic and osmocytic vesicles visualized through light and confocal microscopy. In contrast, after application of a progressive osmotic shift, from an initial osmotic pressure of 2 MPa to a final one of 24 MPa for 30 min, no vesicles were observed. Additionally, the absence of Hechtian strand connections led to the bursting of vesicles in the case of the osmotic shock.

Conclusions

It is concluded that the kinetics of osmotic dehydration strongly influence vesicle formation in onion cells, and that Hechtian strand connections between protoplasts and exocytotic vesicles are a prerequisite for successful deplasmolysis. These results suggest that a decrease in the area-to-volume ratio of a cell could cause cell death following an osmotic shock.     

Effects of Sucrose on Water Relations of Cut, Senescing, Carnation Flowers

Carnation flower stems were stood in water or sucrose solutionand changes in water content, water and osmotic potential, turgorpressure and solutes (sugars, nitrogen, phosphorus, potassium)of petals were measured throughout the flower life. In bothtreatments the petals had a higher specific water content atincipient wilting than when the flowers were first cut. In water,turgor pressure decreased rapidly after the seventh day becauseof a decrease in tissue solute content. In sucrose solution,loss, of solutes was delayed probably because the sugar provideda respiratory substrate to maintain cell membrane integrity.In these cells, sugars and water accumulated causing decreasesin water potential and osmotic potential. Solutes and waterwere lost at about day 15 and turgor pressure decreased. Therewas some evidence that from about day 11 cells were so gorgedwith sugars that they burst when they were placed in water duringthe adjustment of water content prior to water potential measurements. Most of the initial petal osmotic energy content could be accountedfor by sugar, potassium, and anions associated with potassium,but in water, as the petals aged and sugar content decreased,so the potassium ions contributed a larger proportion of theosmotic energy; with stems in sucrose, the endogenous sugarcontent (reducing sugars plus sucrose) contributed an increasingproportion of the total osmotic energy. Dianthus caryophyllus, carnation, flowers, water relations, senescence     

Autolysis of Escherichia coli

Autolysis of unwashed exponential-phase Escherichia coli cells was efficiently promoted by first submitting them to a quick downshock with distilled water before an upshock with 0.5 M sodium acetate, pH 6.5. The association of these two osmotic shocks had a remarkable synergistic effect and led to significant decreases in turbidity and viability. Different factors influencing the rate of cell lysis were examined. A close correlation was established between autolysis and the degradation of peptidoglycan. Both phenomena were induced by the same shock treatment, followed similar kinetics, and were efficiently blocked by addition of divalent cations. Cell lysis was also inducible by a shock treatment with 10(-3) M ethylenediaminetetraacetic acid or ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid and blocked by the addition of divalent cations.     

Effect of osmotic pressure on membrane energy-linked functions in Escherichia coli

Osmotic upshock of E. coli cells in NaCl or sucrose medium resulted in a large decrease in the cytoplasmic volume and the inhibition of growth, of the electron transfer chain and of four different types of sugar transport system: the lactose proton symport, the glucose phosphotransferase system, the binding-protein dependent maltose transport system and the glycerol facilitator. In contrast to NaCl and sucrose, the permeant solute glycerol had no marked effect. These inhibitions could be partially relieved by glycine betaine. Despite these inhibitions, the internal pH, the protonmotive force and the ATP pool were maintained. It is concluded that inhibition of electron transfer and of sugar transport is the consequence of conformational changes caused by the deformation of the membrane. It is also concluded that the arrest of growth observed upon osmotic upshock is not due to energy limitations and that it cannot be explained by the inhibition of carbohydrate transport.     

Effects of accumulation of 3-O-methylglucose on levels of endogenous osmotic solutes in Chlorella emersonii

Abstract. Regulation of the concentration of osmotic solutes was studied in Chlorella emersonii grown at external osmotic pressures (II) ranging between 0.08 and 1.64MPa. NaCl was used as osmoticum. The total solute content of the cells was manipulated by applying 2 mol m⁻³ 3- O -methylglucose (MG), which was not metabolized, and accumulated at concentrations ranging between 60 and 230 mol m⁻³ within 4 h after its addition to the medium. Methylglucose uptake resulted in decreases in concentrations of proline and sucrose, the two solutes mainly responsible for osmotic adaptation of C. emersonii to high external II. The responses were consistent with the hypothesis that proline and sucrose concentrations are controlled by a system of osmotic regulation, with turgor and/or volume as a primary signal. Short-term experiments showed that even very small increases in turgor and/or volume, due to accumulation of methylglucose, resulted in large decreases in proline and sucrose. Over the first 30-60 min the total solute concentration in the cells increased by at most 15 osmol m⁻³ which would represent an increase in turgor pressure of at most 0.04 M Pa. Yet, the decreases in proline and sucrose were as fast as those in cells exposed to a sudden decrease of 0.25 MPa in external II, when the turgor pressure would have increased by at least 0.15 MPa. High concentrations of methylglucose in cells grown at high II did not affect the rapid synthesis of proline and sucrose which started when the cells were transferred to yet higher II. Thus, methylglucose had no direct effects on proline and sucrose metabolism, and it has been assumed that it acted solely as an inert osmotic solute within the cell.     

多种被子植物花粉生殖细胞大量分离技术的比较研究

从11种植物的成熟花粉粒中分离出大量生活的生殖细胞。比较了四种分离方法(一步渗透压冲击法、二步渗透压冲击法、低酶法和花粉原生质体释放法)在不同植物中的效果。归纳出三类植物适于采用三种分离方法。研究了影响分离效果的若干重要因素。对5种植物的分离生殖细胞进行了纯化。经鉴定,纯化的细胞群体中80%以上的细胞是生活的。     

Adaptive characteristics of salt-induced myceloids of Arthrobacter globiformis

When Arthrobacter globiformis is grown in medium containing increased concentrations of NaCl or decreased levels of cations, the bacteria grow as clusters of branching myceloid cells. The sensitivities of salt-induced and citrate-induced myceloids to several environmental stresses were compared to those of normal exponential-phase bacilli and stationary-phase cocci. Salt-induced myceloids were more resistant than normal cells to ultraviolet light or heat shock at 45°C but not to osmotic upshock or pH 4.3; citrate-induced myceloids showed an intermediate rate of heat inactivation. Carbon or nitrogen starvation of myceloids in the absence of added NaCl or citrate led to their division into single cells. Both myceloids and the single cells derived from them were more resistant than normal bacteria to nitrogen starvation. Salt-induced and citrate-induced myceloids showed reduced metabolism of many different carbon compounds in Biolog GP plates. These studies suggest that the formation of multicellular structures by A. globiformis is an adaptive response which increases its potential for survival.     

Enhancement of phenylethanoid glycosides biosynthesis in cell cultures of <Emphasis Type="Italic">Cistanche deserticola</Emphasis> by osmotic stress

The effect of osmotic stress on cell growth and phenylethanoid glycosides (PeGs) biosynthesis was investigated in cell suspension cultures of Cistanche deserticola Y. C. Ma, a desert medicinal plant grown in west region of China. Various initial sucrose concentrations significantly affected cell growth and PeGs biosynthesis in the suspension cultures, and the highest dry weight and PeGs accumulation reached 15.9 g l⁻¹-DW and 20.7 mg g⁻¹-DW respectively at the initial osmotic stress of 300 mOsm kg⁻¹ where the sucrose concentration was 175.3 mM. Stoichiometric analysis with different combinations of sucrose and non-metabolic sugar (mannitol) or non-sugar osmotic agents (PEG and NaCl) revealed that osmotic stress itself was an important factor for enhancing PeGs biosynthesis in cell suspension cultures of C. deserticola. The maximum PeGs contents of 26.9 and 23.8 mg g⁻¹-DW were obtained after 21 days at the combinations of 87.6 mM sucrose with 164.7 mM mannitol (303 mOsm kg⁻¹) or 20 mM PEG respectively, which was higher than that of C. deserticola cell cultures grown under an initial sucrose concentration of 175.3 mM after 30 days. The stimulated PeGs accumulation in the cell suspension cultures was correlated to the increase of phenylalanine ammonium lyase (PAL) activity induced by osmotic stress.