Protonmotive force in Brevibacterium flavum: the lack of intracellular pH homeostasis

Brevibacterium flavum 22LD-P cells were shown to maintain a transmembrane pH gradient (pH) from 0.6 to 1.8–2 units and a transmembrane electric potential difference () from 0 to 200 mV depending on the pH and ionic composition of the incubation medium, grwoth substrate and concentration of cells. decreased from 120–140 mV to 0 when medium pH was lowered from neutral to 5.0–5.5 and increased to 180–200 mV when medium pH was raised to 8–9 in cells utilizing acetate or endogenous substrate. Cells growing on sucrose, kept around 100–120 mV at neutral as well as acidic medium pH. Intracellular pH in the acetate utilizing or endogenously respiring cells was maintained with the range of 8.9 to 5.5 at medium pH ranging from 9.1 to 4.0, respectively. Sucrose grown cells were able to maintain a more stable intracellular pH. Endogenously respiring cells in potassium phosphate buffer at high biomass concentrations maintained larger pH and relatively smaller , than the same cells in diluted suspensions. Cells in sodium phosphate buffer possessed larger and almost no pH, but was still dependent on biomass concentration.The lack of intracellular pH homeostasis and the collapse of at acid medium pH are discussed in the context of cell membrane proton permeability.     

The ability of acidic pH,growth inhibitors,and glucose to increase the proton motive force and energy spilling of amino acid-fermenting <Emphasis Type="Italic">Clostridium sporogenes</Emphasis> MD1 cultures

Clostridium sporogenes MD1 grew rapidly with peptides and amino acids as an energy source at pH 6.7. However, the proton motive force (p) was only –25 mV, and protonophores did not inhibit growth. When extracellular pH was decreased with HCl, the chemical gradient of protons (ZpH) and the electrical membrane potential () increased. The p was –125 mV at pH 4.7, even though growth was not observed. At pH 6.7, glucose addition did not cause an increase in growth rate, but increased to –70 mV. Protein synthesis inhibitors also significantly increased . Non-growing, arginine-energized cells had a of –80 mV at pH 6.7 or pH 4.7, but was not detected if the F₁F₀ ATPase was inhibited. Arginine-energized cells initiated growth if other amino acids were added at pH 6.7, and and ATP declined. At pH 4.7, ATP production remained high. However, growth could not be initiated, and neither nor the intracellular ATP concentration declined. Based on these results, it appears that C. sporogenes MD1 does not need a large p to grow, and p appears to serve as a mechanism of ATP dissipation or energy spilling.Mandatory disclaimer: Proprietary or brand names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product, and exclusion of others that may be suitable.     

The effect of humidity and light on cellular water relations and diffusion conductance of leaves ofTradescantia virginiana L.

Turgor (_p) and osmotic potential (_s) in epidermal and mesophyll cells, in-situ xylem water potential (-xyl) and gas exchange were measured during changes of air humidity and light in leaves ofTradescantia virginiana L., Turgor of single cells was determined using the pressure probe. Sap of individual cells was collected with the probe for measuring the freezing-point depression in a nanoliter osmometer. Turgor pressure was by 0.2 to 0.4 MPa larger in mesophyll cells than in epidermal cells. A water-potential gradient, which was dependent on the rate of transpiration, was found between epidermis and mesophyll and between tip and base of the test leaf. Step changes of humidity or light resulted in changes of epidermal and mesophyll turgor (_p-epi, _p-mes) and could be correlated with the transpiration rate. Osmotic potential was not affected by a step change of humidity or light. For the humidity-step experiments, stomatal conductance (g) increased with increasing epidermal turgor.g/_p-epi appeared to be constant over a wide range of epidermal turgor pressures. In light-step experiments this type of response was not found and stomatal conductance could increase while epidermal turgor decreased.Symbols E transpiration - g leaf conductance - w leaf/air vapour concentration difference - -epi water potential of epidermal cells - -mes water potential of mesophyll cells - -xyl water potential of xylem - _p-epi turgor pressure of epidermal cells - _p-mes turgor pressure of mesophyll cells - _s-epi osmotic potential of epidermal cells - _s-mes osmotic potential of mesophyll cells     

An investigation of the role of solutes in the xylem sap and in the xylem parenchyma as the source of root pressure

Summary Solute osmotic potentials (_x) in the vessels of hydroponically grown maize roots were measured to assess the osmotic-xylem-sap mechanism for generating root pressure (indicated by guttation). Solutes in vessels were measured in situ by X-ray microanalysis of plants frozen intact while guttating. Osmotic potentials outside the roots (_o) were changed by adding polyethylene glycol to the nutrient solution. Guttation rate fell when _o was decreased, but recovered towards the control value during 3–5 days when _o was greater than or equal to –0.3 MPa, but not when _o was equal to –0.4 MPa. In roots stressed to _o = –0.3 MPa, _x, was always more positive than _o, and _x changed only slightly (ca. 0.05 MPa). Thus the adjustment in the roots which increased root pressure cannot be ascribed to _x, contradicting the osmotic-xylem-sap mechanism. An alternative driving force was sought in the osmotic potentials of the vacuoles of the living cells (_v), which were analysed by microanalysis and estimated by plasmolysis. _v showed larger responses to osmotic stress (0.1 MPa). Some plants were pretreated with abundant KNO₃ in the nutrient solution. These plants showed very large adjustments in _v (0.4 MPa) but little change in _x (0.08 MPa). They guttated by 4 h after _o was lowered to –0.4 MPa. It is argued that turgor pressure of the living cells is a likely alternative source of root pressure. Published evidence for high solute concentrations in the xylem sap is critically assessed.Abbreviations _o external water potential - _x osmotic potential of xylem sap - _v osmotic potential of vacuolar sap - EDX energy dispersive X-ray microanalysis - CSEM cryo-scanning electron microscope - LN₂ liquid nitrogen - PEG polyethylene glycol     

Tetramethyl Rhodamine Methyl Ester (TMRM) is Suitable for Cytofluorometric Measurements of Mitochondrial Membrane Potential in Cells Treated with Digitonin

A new method for cytofluorometric analysis of mitochondrial membrane potential has been developed by using TMRM as a cationic, mitochondrial selective probe. The method is based on limited treatment of cultured cells with digitonin which permeabilises the plasma membrane and leaves mitochondria intact. The resulting signal of TMRM-stained cells thus represents only the probe accumulated in mitochondria. Fibroblasts and cybrids were used as a model cell systems and optimal conditions for digitonin treatment and staining by TMRM were described. The TMRM signal collapsed by valinomycin, KCN and antimycin A and FCCP titration was used to gradually lower and characterise the stability of . The method is suitable for sensitive measurement of in different types of cultured cells.     

Cyclosporin A-sensitive decrease in the transmembrane potential across the inner membrane of liver mitochondria induced by low concentrations of fatty acids and Ca2+

At low Ca²⁺ concentrations the pore of the inner mitochondrial membrane can open in substates with lower permeability (Hunter, D. R., and Haworth, R. A. (1979) Arch. Biochem. Biophys., 195, 468-477). Recently, we showed that Ca²⁺ loading of mitochondria augments the cyclosporin A-dependent decrease in transmembrane potential () across the inner mitochondrial membrane caused by 10 M myristic acid but does not affect the stimulation of respiration by this fatty acid. We have proposed that in our experiments the pore opened in a substate with lower permeability rather than in the classic state (Bodrova, M. E., et al. (2000) IUBMB Life, 50, 189-194). Here we show that under conditions lowering the probability of classic pore opening in Ca²⁺-loaded mitochondria myristic acid induces the cyclosporin A-sensitive decrease and mitochondrial swelling more effectively than uncoupler SF6847 does, though their protonophoric activities are equal. In the absence of P_i and presence of succinate and rotenone (with or without glutamate) cyclosporin A either reversed or only stopped decrease induced by 5 M myristic acid and 5 M Ca²⁺. In the last case nigericin, when added after cyclosporin A, reversed the decrease, and the following addition of EGTA produced only a weak (if any) increase. In P_i-containing medium (in the presence of glutamate and malate) cyclosporin A reversed the decrease. These data show that the cyclosporin A-sensitive decrease in by low concentrations of fatty acids and Ca²⁺ cannot be explained by specific uncoupling effect of fatty acid. We propose that: 1) low concentrations of Ca²⁺ and fatty acid induce the pore opening in a substate with a selective cation permeability, and the cyclosporin A-sensitive decrease results from a conversion of to pH gradient due to the electrogenic cation transport in mitochondria; 2) the ADP/ATP-antiporter is involved in this process; 3) higher efficiency of fatty acid compared to SF6847 in the Ca²⁺-dependent pore opening seems to be due to its interaction with the nucleotide-binding site of the ADP/ATP-antiporter and higher affinity of fatty acids to cations.     

The membrane potential in whole cells of Methanobacterium thermoautotrophicum

of whole cells of Methanobacterium thermoautotrophicum was estimated under varying conditions using an electrode sensitive to the lipophilic cation tetraphenylphosphonium chloride (TPP⁺). Since was found to be extremely sensitive to air, a special reaction vessel was developed to maintain strict anaerobiosis. The cells took up TPP⁺ under energization by H₂ and CO₂ thus allowing to calculate the from the distribution of TPP⁺ inside and outside the cells. The unspecific uptake of deenergized cells was around 10% of the total uptake of energized cells. TPP⁺ itself slightly diminished the , but had no effect on the formation of methane. Typical values of were in the range of-150 to-200 mV. showed a quantitative dependence on both the electron donor H₂ and the electron acceptor CO₂. NaCl stimulated the extent of the , whereas KCl slightly diminished it. Valinomycin resulted in a linear decline of , whereas the methane production rate was only slightly affected. In contrast, monensin reduced both methanogenesis and .Abbreviations pmf proton motive force - membrane potential - TPP⁺ tetraphenylphosphonium (chloride salt) - TPMP⁺ triphenylmethylphosphonium (chloride salt, if not otherwise indicated) - d.w. dry weight - t _d doubling time - PVC polyvinylchloride     

Stomatal response to leaf water potential in almond trees under drip irrigated and non irrigated conditions

Almond plants (Amygdalus communis L. cv. Garrigues) were grown in the field under drip irrigated and non irrigated conditions. Leaf water potential () and leaf conductance (g₁) were determined at three different times of the growing season (spring, summer and autumn). The relationships between and g₁ in both treatments showed a continuous decrease of g₁ as decreased in spring and summer. Data from the autumn presented a threshold value of (approx. –2.7 MPa in dry treatment, and approx. –1.4 MPa in wet treatment) below which leaf conductance remained constant.     

Osmoregulation and the control of phloem-sap composition in Ricinus communis L.

Phloem-sap composition was studied in plants of Ricinus communis L. grown on a waterculture medium. The sap possessed a relatively high K⁺:Na⁺ ratio and low levels of Ca²⁺ and free H⁺. Sucrose and K⁺ (together with its associated anions) accounted for 75% of the phloem-sap solute potential (_s). In plants kept in continuous darkness, a decrease in phloem-sap sucrose levels over 24h was accompanied by an increase in K⁺ levels. Measurements of phloem-sap _s and xylem water potential () indicated that this resulted in a partial maintenance of phloem turgor pressure _p. In darkness there was also a marked decrease in the malate content of the leaf tissue, and it is possible that organic carbon from this source was mobilized for export in the phloem. The results support the concept of the phloem sap as a symplastic phase. We interpret the increase in K⁺ levels in the phloem in darkness as an osmoregulatory response to conditions of restricted solute availability. This reponse can be explained on the basis of the sucrose-H⁺ co-transport mechanism of phloem loading.Abbreviations water potential - _s solute potential - _p pressure potential     

Control of the rate of cell enlargement: Excision,wall relaxation,and growth-induced water potentials

A new guillotine thermocouple psychrometer was used to make continuous measurements of water potential before and after the excision of elongating and mature regions of darkgrown soybean (Glycine max L. Merr.) stems. Transpiration could not occur, but growth took place during the measurement if the tissue was intact. Tests showed that the instrument measured the average water potential of the sampled tissue and responded rapidly to changes in water potential. By measuring tissue osmotic potential ( _s ), turgor pressure ( _p ) could be calculated. In the intact plant, _s and _p were essentially constant for the entire 22 h measurement, but _s was lower and _p higher in the elongating region than in the mature region. This caused the water potential in the elongating region to be lower than in the mature region. The mature tissue equilibrated with the water potential of the xylem. Therefore, the difference in water potential between mature and elongating tissue represented a difference between the xylem and the elongating region, reflecting a water potential gradient from the xylem to the epidermis that was involved in supplying water for elongation. When mature tissue was excised with the guillotine, _s and _p did not change. However, when elongating tissue was excised, water was absorbed from the xylem, whose water potential decreased. This collapsed the gradient and prevented further water uptake. Tissue _p then decreased rapidly (5 min) by about 0.1 MPa in the elongating tissue. The _p decreased because the cell walls relaxed as extension, caused by _p , continued briefly without water uptake. The _p decreased until the minimum for wall extension (Y) was reached, whereupon elongation ceased. This was followed by a slow further decrease in Y but no additional elongation. In elongating tissue excised with mature tissue attached, there was almost no effect on water potential or _p for several hours. Nevertheless, growth was reduced immediately and continued at a decreasing rate. In this case, the mature tissue supplied water to the elongating tissue and the cell walls did not relax. Based on these measurements, a theory is presented for simultaneously evaluating the effects of water supply and water demand associated with growth. Because wall relaxation measured with the psychrometer provided a new method for determining Y and wall extensibility, all the factors required by the theory could be evaluated for the first time in a single sample. The analysis showed that water uptake and wall extension co-limited elongation in soybean stems under our conditions. This co-limitation explains why elongation responded immediately to a decrease in the water potential of the xylem and why excision with attached mature tissue caused an immediate decrease in growth rate without an immediate change in _p Abbreviations and symbols L tissue conductance for water - m wall extensibility - Y average yield threshold (MPa) - _o water potential of the xylem - _p turgor pressure - _s osmotic potential - _w water potential of the elon gating tissue     

Sex identification by male-specific growth hormone pseudogene (GH-Ψ) in Oncorhynchus masou complex and a related hybrid

It is often difficult to identify sexes of many fish species by conventional cytological method because of the lack of heteromorphic sex chromosomes. Isolation of sex-specific molecular markers is thus important for sexing and for understanding sex chromosome evolution in these species. We have identified genetic sexes by PCR-based male-specificity of a growth hormone pseudogene (GH-) in masu and Biwa salmon, two subspecies of the Oncorhynchus masou complex, and their hybrid Honmasu. PCRs with primers designed from sequences of chinook salmon GH genes amplified GH-I and GH-II fragments in both sexes, but a third GH- fragment was detected in predominant proportion of males and very few phenotypic females. The consistency of phenotypic sex with genetic sex identified by GH- for masu salmon, Biwa salmon and Honmasu was 93.1, 96.7 and 94%, respectively. The remaining individuals showed inconsistency or deviation from sex-specificity: a few phenotypic males lacked the GH-, whereas a few phenotypic females possessed the GH-. Sequence of the putative GH- fragment from such females was identical to that from genetic males, and shared about 95% homology with the corresponding GH- fragment from chinook salmon. This result confirmed that these females were really GH--bearing individuals. PCR analyses with primers designed from masu salmon GH- gave identical results, indicating that the absence of GH- in a few males was not resulted from primer mismatching. These GH--bearing females and GH--absent males were more likely to originate from spontaneous sex reversion than from crossing-over between GH- and the sex determination gene/region.     

Electrochemical potential of protons in vesicles reconstituted from purified,proton-translocating adenosine triphosphatase

Summary Measurements were made of the difference in the electrochemical potential of protons ( ) across the membrane of vesicles reconstituted from the ATPase complex (TF ₀ ·F ₁) purified from a thermophilic bacterium and P-lipids. Two fluorescent dyes, anilinonaphthalene sulfonate (ANS) and 9-aminoacridine (9AA) were used as probes for measuring the membrane potential () and pH difference across the membrane ( pH), respectively.In the presence of Tris buffer the maximal and no pH were produced, while in the presence of the permeant anion NO ₃ ^– the maximal pH and a low were produced by the addition of ATP. When the ATP concentration was 0.24mm, the was 140–150 mV (positive inside) in Tris buffer, and the pH was 2.9–3.5 units (acidic inside) in the presence of NO ₃ ^– . Addition of a saturating amount of ATP produced somewhat larger and pH values, and the attained was about 310 mV.By trapping pH indicators in the vesicles during their reconstitution it was found that the pH inside the vesicles was pH 4–5 during ATP hydrolysis.The effects of energy transfer inhibitors, uncouplers, ionophores, and permeant anions on these vesicles were studied.     

The electrochemical proton gradient and its influence on citrate uptake in tonoplast vesicles of Hevea brasiliensis

The relationship between the electrochemical proton gradient, _H+ ^– , and citrate transport has been studied in tonoplast vesicles from Hevea brasiliensis (the rubber tree). Vesicles were generated from lyophilized samples of fresh vacuoles obtained from the latex sap. Methylamine was used to measure intravesicular pH and lipophilic ions to determine the electrical potential difference () across the tonoplast. When incubated at pH 7.5 in the absence of ATP, the tonoplast vesicles showed a pH of 0.6 units (interior acid) and a of about-100 mV (interior negative). This potential is thought to be made up of contributions from an H⁺ diffusion potential, diffusion potentials from other cations and a Donnan potential arising from the presence of internal citrate. In the presence of 5 mol m^-3 MgATP the pH was increased to about 1.0 unit and the to about-10 mV. Under these conditions the proton-motive force ( _{p H+} ^– /F) became positive and reached +50 mV. These effects were specific to MgATP (ADP and Mg²⁺ having no significant effect) and were prevented by the protonophore p-trifluoromethoxycarbonylcyanidephenylhydrazone (FCCP). Citrate uptake by the vesicles was markedly stimulated by MgATP; ADP and Mg²⁺ again had no effect. Nigericin greatly increased pH and this was associated with a large increase in citrate accumulation. The results indicate that the vesicle membrane possesses a functional H⁺-translocating ATPase. The _H+ ^– generated by this ATPase can be used to drive citrate uptake into the vesicles. The properties of the tonoplast vesicles are compared with those of the fresh latex vacuoles.Abbreviations _H+ ^– electrochemical proton gradient - electrical potential difference across membrane - p proton-motive force ( _H+ ^– /F) - FCCP p-trifluoromethoxycarbonylcyanidephenylhydrazone - TPMP⁺ triphenylmethylphosphonium ion     

Effects of moisture stress on the multiplication and expansion of cells in leaves of sugar beet

Summary Sugar beets were subjected to moisture stress by decreasing the water potential of the culture solution osmotically with polyethylene glycol by a known amount, , and, alternatively by applying matric potential, , at the plant roots. Lowering the water potential at the root surface less than 200 millibars by either method resulted in significant decreases in the rate of cell multiplication. The final number of cells per leaf at = -372 mb the final was 165% of that at = -473 mb ( = –101 mb); similarly at = –15 mb the final cell number was 198% of that at = –196 mb ( = –181 mb). The mean cell volume of leaves was not significantly affected by these levels of moisture stress.     

Transmembrane signal defect and absence of cancer extract induced leukocyte adherence inhibition (LAI) for leukocytes from patients with advanced cancer

Summary Leukocytes from patients with early cancer exhibit leukocyte adherence inhibition (LAI) when incubated with extracts of cancer of the same organ and histogenesis, whereas leukocytes from patients with advanced cancer seldom do. To understand the reason for this refractory state, tumor antigen-induced LAI and transmembrane signalling were measured in the same leukocytes. Transmembrane signalling was measured by changes in membrane potential () by the [³H]tetraphenylphosphonium equilibration technique. When leukocytes from patients with early breast cancer were incubated with extracts of breast cancer and malignant melanoma they showed changes consisting of depolarization and hyperpolarization beginning within 0.5 min after addition of the breast cancer extract and finishing 15 min later. Moreover, they showed no changes when incubated with extracts of normal breast tissue. Leukocytes from subjects without cancer seldom showed changes. In criss-cross experiments, leukocytes from patients with melanoma only exhibited changes when incubated with the melanoma extract. There was a strong correlation between cancer extract-induced change and LAI. The change was triggered by leukotriene-like mediators from antibody-dependent monocytes. Authentic leukotrienes triggered changes in all subpopulation of leukocytes. Leukocytes from patients with advanced breast cancer when incubated with breast cancer extract did not transmit a signal or show LAI. Brief elevation of intracellular cyclic AMP restored both change and LAI induced by breast cancer extracts, indicating that reactive leukocytes are present but in a refractory state. We conclude that leukocytes from patients with advanced cancer do not react in LAI because tumor antigen does not trigger a transmembrane signal to initiate the cascade of biochemical reactions and physiological changes for LAI.Abbreviations CCCP carbonyl cyanide m-chlorophenylhydrazone - cyclic AMP cyclic adenosine monophosphate - ETYA eicosatetraynoic acid - HEPES 4-(2-hydroxyethyl)-1-piperazine ethansulfonate - LAI leukocyte adherence inhibition - NAI nonadherence index - OSN organ-specific cancer neoantigen - PBL peripheral blood leukocytes - PGE₂ prostaglandin E₂ - [³H]TPP⁺ [phenyl³H]tetraphenylphosphonium bromide - transmembrane potential     

The protonmotive force and phosphorylation potential developed by whole cells of the methylotrophic bacteriumMethylophilus methylotrophus

The and the Gp have been measured in whole cells ofMethylophilus methylotrophus during the oxidation of various respiratory chain substrates. The magnitude of the depended on the external pH and the composition of the assay medium, and varied from-109 to-165 mV. The relative contributions of the and the pH to the were found to vary with the external pH such that the internal pH remained constant; depending on the composition of the assay medium, this value was between 6.6 and 7.0. A Gp of approximately-46 kJ/mol was generated during the oxidation of methanol, and either the or pH alone was fully competent to drive ATP synthesis. Respiration and ATP synthesis were found to be poised far from equilibrium under the conditions of these experiments, and the value of the Gp was thus controlled kinetically. Comparison of the with the Gp yielded an H⁺/ATP quotient >2.6 g-ion H⁺/mol ATP.Abbreviations TMPD N,N,N,N-tetramethyl-p-phenylenediamine - FCCP carbonylcyanidep-trifluoromethoxyphenylhydrazone - DMO 5,5-dimethyloxazolidine-2,4-dione - TPMP⁺ triphenylmethylphosphonium (iodide salt); Tween 20, polyoxyethylenesorbitan monolaurate - TPP⁺ tetraphenylphosphonium (bromide salt) - bulk phase transmembrane electrochemical potential difference of protons ( ) - pH bulk phase transmembrane pH difference (pHin-pHout) - bulk phase transmembrane electrical potential difference (in-out) - p true protonmotive force (incorporating both bulk phase and localised protons; )     

Monitoring the mitochondrial transmembrane potential with the JC-1 fluorochrome in programmed cell death during mesophyll leaf senescence

Summary. Analysis of the mitochondrial transmembrane potential (_m) with the help of the JC-1 fluorochrome (5,5,6,6-tetrachloro-1,1,3,3-tetraethylbenzimidazolcarbocyanine iodide) during mesophyll leaf senescence was performed in order to determine whether a reduction of _m takes place during mesophyll senescence and whether plant mitochondria, like mammalian ones, might be involved in the induction of programmed cell death. Fluorescence analysis of mesophyll protoplasts of Pisum sativum in a confocal microscope, fluorescent spectra analysis and time dependence of fluorescence intensity of monomers and of J-aggregates revealed that JC-1 is incorporated and accumulated specifically in plant mitochondria. Analysis of _m during mesophyll protoplast senescence revealed that two subpopulations of mitochondria which differ in _m exist in all analyzed stages of leaf senescence. The first subpopulation contains mitochondria with red fluorescence of J-aggregates due to an unperturbed high _m. The second subpopulation comprises mitochondria with green fluorescence of monomers due to a low _m, proving total depolarization of mitochondrial membranes. Fluorescence analysis demonstrated that even in the latest analyzed stages of leaf senescence, mitochondria with a high _m still exist. Fluorometric measurements revealed that the fluorescence intensity of J-aggregates decreases with the age of plants, which indicates that a reduction of _m during the mesophyll senescence process takes place; however, it does not take place within the whole population of mitochondria of the same protoplast. The reason of this can be due to a dramatic reorganization of mitochondria in mesophyll cells and the appearance of large mitochondria with local heterogeneity of _m in the oldest analyzed stages. All mitochondria in every stage of senescence maintained their membrane organization even when their size, distribution, and spatial organization in protoplasts changed dramatically. We stated that the reduction of _m does not directly induce programmed cell death in mesophyll cells, as opposed to animal apoptosis.Correspondence and reprints: Department of Plant Anatomy and Cytology, Institute of Experimental Biology of Plants, Warsaw University, Miecznikowa 1, 02-096 Warszawa, Poland.     

Sodium ions are necessary for growth and energy transduction in the marine cyanobacterium Oscillatoria brevis

The maximal growth rate of the marine cyanobacterium Oscillatoria brevis was reached at 200–400 mM NaCl and pH 9.0–9.6. NaCl was found (i) to stimulate the rate of the light-supported generation across the cytoplasmic membrane of the cells and (ii) to decrease the sensitivity of level and motility of the O. brevis trichomes to protonophorous uncouplers. The Na⁺/H⁺ antiporter, monensin, increased both and the uncoupler sensitivity of the cells. The data obtained agree with the assumption that O. brevis possesses a primary Na⁺ pump in its cytoplasmic membrane.Abbreviations ATP adenosine-5-triphosphate - TTFB tetrachlortrifluoromethylimidazol - CCCP carbonyl cyanide m-chlorophenylhydrazone - Na⁺ transmembrane electrochemical potential differences of Na⁺ - transmembrane electric potential difference - pNa transmembrane pNa difference     

Osmotic adjustment and the inhibition of leaf,root, stem and silk growth at low water potentials in maize

The expansion growth of plant organs is inhibited at low water potentials ( _w), but the inhibition has not been compared in different organs of the same plant. Therefore, we determined elongation rates of the roots, stems, leaves, and styles (silks) of maize (Zea mays L.) as soil water was depleted. The _w was measured in the region of cell expansion of each organ. The complicating effects of transpiration were avoided by making measurements at the end of the dark period when the air had been saturated with water vapor for 10 h and transpiration was less than 1% of the rate in the light. Growth was inhibited as the _w in the region of cell expansion decreased in each organ. The _w required to stop growth was-0.50,-0.75, and-1.00 MPa, in this order, in the stem, silks, and leaves. However, the roots grew at these _w and ceased only when _w was lower than-1.4 MPa. The osmotic potential decreased in each region of cell expansion and, in leaves, roots and stems, the decrease was sufficient to maintain turgor fully. In the silks, the decrease was less and turgor fell. In the mature tissue, the _w of the stem, leaves and roots was similar to that of the soil when adequate water was supplied. This indicated that an equilibrium existed between these tissues, the vascular system, and the soil. At the same time, the _w was lower in the expanding regions than in the mature tissues, indicating that there was a _w disequilibrium between the growing tissue and the vascular system. The disequilibrium was interpreted as a _w gradient for supplying water to the enlarging cells. When water was withheld, this gradient disappeared in the leaf because _w decreased more in the xylem than in the soil, indicating that a high flow resistance had developed in the xylem. In the roots, the gradient did not decrease because vascular _w changed about the same amount as the soil _w. Therefore, the gradient in _w favored water uptake by roots but not leaves at low _w. The data show that expansion growth responds to low _w differently in different growing regions of the plant. Because growth depends on the maintenance of turgor for extending the cell walls and the presence of _w gradients for supplying water to the expanding cells, several factors could have been responsible for these differences. The decrease of turgor in the silks and the loss of the _w gradient in the leaves probably contributed to the high sensitivity of these organs. In the leaves, the gradient loss was so complete that it would have prevented growth regardless of other changes. In the roots, the maintenance of turgor and _w gradients probably allowed growth to continue. This difference in turgor and gradient maintenance could contribute to the increase in root/shoot ratios generally observed in water-limited conditions.Symbols _s osmotic potential - _w water potential     

Subcellular heterogeneity of mitochondrial membrane potential in anther tapetum

Studies on animal material have revealed that changes in the mitochondrial permeability transition pore (PTP), which cause a reduction in the mitochondrial transmembrane potential (_m) followed by release of cytochrome c, belong to the earliest manifestations of some types of apoptosis. We have attempted to monitor the _m of mitochondria during programmed cell death (PCD) of the secretory tapetum using JC-1, a fluorochrome dye that detects mitochondrial membrane potential and to relate changes in this potential to mitochondrial ultrastructure. Analysis of tapetal cells isolated from Ornithogalum virens anthers revealed that the _m of mitochondria in the tapetal cells alters during development; the change, however, is not uniform in the mitochondrial population within a single tapetal cell. In young tapetal cells, at the tetrad stage, we detected only the red fluorescence of JC-1 aggregates in all tapetal mitochondria, which indicates highly negative _m. In an advanced stage of PCD at the late microspore stage, in each tapetal cell we detected both mitochondria with red (as formerly) and mitochondria with green fluorescence. The green fluorescence of JC-1 monomers indicates mitochondria with depolarised membranes. These changes in _m are related to observed changes in mitochondria ultrastructure. This is the first documentation of intracellular heterogeneity of _m during anther tapetum development. Alteration in _m suggests a relationship between mitochondrial function and PCD processes in tapetal cells.