Thansplasmalemma redox reactions and ion transport in photosynthetic and heterotrophic plant cells

Extracellular ferricyanide reduction, NADH and ferrocyanide oxidation were investigated by spectrophotometrical method on photosynthetic freshwater plants ( Elodea canadensis Rich., Vallisneria spiralis L., Nitella flexilis L.) and heterotrophic tissues (roots of Triticum vulgare L., Hordeum vulgare L., Zea mays L., Pisum sativum L., Avena sativa L., Allium sativa L., Allium cepa L.). All species had ferricyanide reductase activity. The roots of land plants also carried out extracellular oxidation of NADH and ferrocyanide in contrast to leaves of the freshwater plants. External NADH stimulated ferricyanide reductase activity, but only with those objects that had external NADH oxidase activity. In all species ferricyanide decreased the membrane potential (MP), decreased the membrane resistance measured at a fixed current and inhibited K⁺ influx measured by flame photometry. The factors affecting ferricyanide reductase activity also influenced the inhibitory effect of ferricyanide on the MP and K⁺ transport. These results demonstrate a connection between transport, electrogenic and redox functions of the plasmalemma.     

Membrane-related processes and overall energy metabolism inTrypanosoma brucei and other kinetoplastid species

An electrochemical proton gradient exists across the plasma membrane and the mitochondrial membrane of the bloodstream form ofTrypanosoma brucei. The membrane potential across the plasma membrane and the regulation of the internal pH depend on the temperature.Leishmania donovani regulates its internal pH and maintains a constant electrochemical proton gradient across its plasma membrane under all conditions examined. The mitochondrion of theT. brucei bloodstream form is energized, even though the reactions taking place in it do not result in net ATP synthesis and the Kreb's cycle and the respiratory chain are absent. Glucose is transported across the plasma membrane ofT. brucei by a facilitated diffusion carrier, that can transport a wider range of substrates than its mammalian counterparts. Pyruvate exits the cell via a facilitated diffusion transporter as well. Conflicting evidence exists for the mechanism of glucose transport inL. donovani; biochemical evidence suggests proton/glucose symport, while facilitated diffusion is indicated by physiological data.     

The influence of an electric field on growth and trace metal content in aquatic plants

It is known that both natural and artificial electric fields (EF) affect plants physiological parameters as well as germination, growth and yield. The present article describes results of a preliminary experiment on the impact of electric field on aquatic plants biogeochemistry. The objective of the present study was the assessment of the influence exerted by the electric field on growth and trace metals content of Elodea canadensis. In a laboratory experiment plants were exposed to the field intensity of 54?kV m^?1 for 7?days. The plants length was measured and the content of Fe, Mn, Ni, Pb, and Zn was determined using atomic absorption spectrometry (AAS). Results showed that the application of electric field slightly enhanced the growth of E. canadensis shoots. The content of Mn and Ni was significantly lower, and Pb and Zn significantly higher in plants exposed to the electric filed, while Fe content did not differ between control and EF treatment. This provides a rationale for further studies on biological effects of electric field in trace metal contaminated waters and application of an electrically enhanced phytoremediation.     

The site of action of abscisic acid at the guard cell plasmalemma of Valerianella locusta

Abstract. Abscisic acid (ABA) is taken up by guard cells of isolated epidermata of Valerianella locusta only at low external pH values. At pH 8.0, when nearly all ABA molecules are present as the union of ABA (ABA⁻), no uptake can be observed. ABA-dependent movement of stomata was tested at external pH values between 5.0 and 8.0. Independent of the external pH, ABA induced stomatal closure at all tested ABA concentrations. It is concluded that ABA need not be taken up into the cytosol of the guard cells in order to induce slomatal closure. The primary site of ABA action at the guard cell plasmalemma must be located either at the outer surface of the plasmalemma or at least be easily accessible from outside. ABA− is as effective as undissociated ABA (ABAH).     

The proton pumps of the plasmalemma and the tonoplast of higher plants

Studies on intact cells, membrane vesicles, and reconstituted proteoliposomes have demonstrated in higher plants the existence of an ATP-driven electrogenic proton pump operating at the plasmalemma. There is also evidence of a second ATP-driven H⁺ pump localized at the tonoplast. The characteristics of both these ATP-driven pumps closely correspond to those of the plasmalemma and tonoplast proton pumps ofNeurospora and yeasts.     

The role of the plasmalemma in metal tolerance in angiosperms

Evidence for the role of the plasmalemma in metal tolerance of metal resistant ecotypes, cultivars and clones is presented. A range of tolerance mechanisms involving the plasmalemma are discussed including alterations to protein carrier and channel function and synthesis, efflux pumps and maintenance of plasmalemma integrity. Specific examples of such alterations from the literature on Al, As and Cu tolerance, where the plasmalemma has been shown to have a role in tolerance are considered. Tolerance by alterations to plasmalemma function in tolerant ecotypes may also rely on internal metal detoxification mechanisms constitutive in tolerant and non-tolerant plants.     

Increased resistance to copper-induced damage of the root cell plasmalemma in copper tolerant Silene cucubalus

The relation between copper tolerance and the sensitivity of plants with respect to the effect of copper on the plasmalemma of root cells was studied using plants from one copper sensitive and two copper tolerant populations of Silene cucubalus Wib. In each population, the external copper concentration needed to induce ion leakage (a measure of damage to the permeability barrier) was similar to the highest no-effect-concentration of copper for root growth in that population. At higher concentrations, the degree of root growth inhibition paralleled the rate of ion leakage, the degree of trypan blue staining (a measure of plasmalemma integrity) and the accumulation of lipid peroxidation products. The amount of copper taken up by the plants was inversely related to their level of copper tolerance. Compared to copper sensitive plants, copper tolerant plants showed no increased resistance to either the sulfhydryl reagent N-ethylmaleimide or the free radical-producing compound cumene hydroperoxide.
These results indicate that damage to the permeability barrier of root cells constitutes the primary effect of copper toxicity in both sensitive and tolerant plants, and that copper tolerance is coupled to the ability of the plants to prevent such damage. This ability might depend on exclusion of copper by the root cell plasmalemma.     

In vivo binding of auxin to the plasmalemma and tonoplast of parenchymal cells in the wheat coleoptile

Tritiated auxin applied by an agar block on the wheat coleoptile tip for 2 hr was covalently fixed to adjacent protein by treatment with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (DCC). The density of labelled auxin in the nucleus, the cell wall, the cytoplasm, and the vacuole was determined by autoradiography. Localization of tritiated auxin was studied at high resolution at the tonoplast and the plasmalemma lining the transverse (distal and proximal) and the longitudinal walls. The radioactivity along the tonoplast was always less than along the plasma membrane. The distribution of ³H-auxin was different across the longitudinal and transverse regions of the plasmalemma. The labelling was distributed asymmetrically on the longitudinal plasma membrane with a peak observed on the external surface. Tritiated auxin was distributed more symmetrically on the distal and the proximal plasma membranes. Our results are in agreement with the hypothesis that there are 2 different specific binding sites on the plasmalemma. The ratio of auxin present at the proximal and distal regions of the plasmalemma was 1.28.     

Distribution of Plasma Membrane H+-ATPase and Polar Current Patterns in Leaves and Stems of Elodea canadensis*

The proton pumping ATPase in the plasma membrane of Elodea canadensis is believed to play a major role in inorganic carbon acquisition. To investigate potentially different carbon uptake strategies within the same plant, plasma membrane H⁺-ATPase distribution and polar current patterns were investigated in Elodea leaves and stems. Specific activity of plasma membrane H⁺-ATPase in leaf microsomal fractions was tenfold higher than in stem derived microsomes. Probing western blots with a monoclonal antibody specific for plasma membrane H⁺-ATPase, yielded strongly visible double bands at 100 kDa in leaf microsome preparations, whereas little antigen was detected in analogous stem microsome preparations. Using the same plasma membrane H⁺-ATPase specific antibody on tissue sections, the enzyme was found almost exclusively localized at the border of cells at the lower leaf surface. A positive ion current leaving the lower leaf surface was measured, using a vibrating probe device. Part of this current entered the upper leaf surface and part of it the internodes of the stem. The experimental results support the view, that Elodea leaves have different means of inorganic carbon uptake than stem internodes.     

Purification and characterization of the apical plasma membrane of the rat pancreatic acinar cell

Summary A method is described for the rapid purification of the apical plasma membrane from the rat pancreatic acinar cell. It makes use of wheat germ agglutinin affinity chromatography to selectively bind vesicles with N-acetyl glucosamine present at their surface. Particular conditions (150 mm NaCl) had then to be used to keep membrane vesicles in the coveted orientation, i.e. as right-side-out vesicles. Due to its specific apical location in many epithelial cells, -glutamyltranspeptidase was chosen to monitor the purification procedure. The final fraction was enriched in -glutamyltranspeptidase by a factor of 75 relative to the homogenate. Na,K-ATPase, a strict basolateral membrane marker, was not detectable in the fraction. No membranes originating from other compartments, more particularly expected from zymogen granules, or from other cell types, did contaminate the preparation. As expected for an epithelial cell apical plasmalemma, lipid composition showed a very high ratio of glycolipids (37.5%). The absence of membrane-bound GP-2, and the exceptionally high specific activity of -glutamyltranspeptidase suggest that the apical membrane would not be made up by the exocytosis of secretory granule, but instead by the fusion of specialized secretory vesicles very likely originating from the constitutive secretory pathway. In conclusion, this report describes a method of obtaining a fraction highly enriched in the secretory apex of the pancreatic exocrine cell that would be directly involved in exocytosis with zymogen granules and also in local anion transport.The authors would like to thank Dr. Andrew W. Shyjan (Yale University, New Haven, CT) for his kind gift of anti-Na,K-ATPase 1 subunit, Dr. Yannick Laperche (INSERM, Hôpital Mondor, Paris) for his gift of anti--GT, and finally Mr. Gilles P. Grondin for the production of antibodies against amylase. D.L. is supported by NSERC of Canada, FCAR of Québec, and the Canadian Cystic Fibrosis Foundation.     

Online control of cell culture redox potential prevents antibody interchain disulfide bond reduction

The phenomenon of monoclonal antibody (mAb) interchain disulfide bond reduction during manufacturing processes continues to be a focus of the biotechnology industry due to the potential for loss of product, increased complexity of purification processes, and reduced stability of the drug product. We hypothesized that antibody reduction can be mitigated by controlling the cell culture redox potential and subsequently established a threshold redox potential above which the mAb remained intact and below which there were significant and highly variable amounts of reduced mAb. Using this knowledge, we developed three control schemes to prevent mAb reduction in the bioreactor by controlling the cell culture redox potential via an online redox probe. These control methodologies functioned by increasing the concentration of dissolved oxygen (DO), copper (II) (Cu), or both DO and Cu to maintain the redox potential above the threshold value. Using these methods, we were able to demonstrate successful control of antibody reduction. Importantly, the redox control strategies did not significantly impact the cell growth, viability, mAb production, or product quality attributes including aggregates, C-terminal lysine, high mannose, deamidation, and glycation. Our results demonstrate that controlling the cell culture redox potential is a simple and effective method to prevent mAb reduction.     

物质的内吞、外排与质膜、内膜系统的关系

高中生物必修本第1册第2章在讲到细胞膜的主要功能时,以小字的形式提到细胞的内吞作用和外排作用,表述较简单,读者不甚理解。其实这也是活细胞进行新陈代谢作用,不断地与外界环境交换物质,物质通过细胞膜进出细胞的方式之一。离子和小分子物质进出     

Redox characteristics of normal and habituated cell lines of sugarbeet

Redox properties of cells of normal hormone-requiring (N) and habituated (H) sugarbeet calli have been investigated. It was found that H cells at all ages reduced exogenous ferricyanide at a much higher rate than N cells, but they exhibited a lower chemiluminescence either alone or in presence of luminol. The plasma membrane NADH:fer-ricyanide oxidoreductase measured in vitro was almost identical for the two cell types. This could indicate that the higher reducing power of H cells had an intracellular origin. Among the enzyme activities which could provide electrons, malate dehydrogenase activity was found to be a good candidate, being more active in H cells. The results are discussed in relation to the abnormal structure of the cell wall of habituated cells.     

A link between transport and plasma membrane redox system(s) in carrot cells

Carrot (Daucus carota L.) cells grown in suspension culture oxidized exogeneous NADH. The NADH oxidation was able to stimulate K⁺ (⁸⁶Rb⁺) transport into cells, but it did not affect sucrose transport.N,N'-Dicyclohexyl-carbodiimide, diethylstilbestrol, and oligomycin, which only partially inhibited NADH oxidation, almost completely collapsed the K⁺ (⁸⁶Rb⁺) transport. Vanadate, which is less effective as an ion transport inhibitor, was less effective in inhibiting the NADH-driven transport of K⁺ (⁸⁶Rb⁺).p-Fluormethoxycarbonylcyanide phenylhydrazone inhibits the K⁺ transport over 90% including that induced by NADH. The results are interpreted as evidence that a plasma membrane redox system in root cells is closely associated with the ATPase which can drive K⁺ transport. Because of the inhibitor effects, it appears that membrane components common to the redox system and ATPase function in the transport of K⁺.     

The differences in energy metabolism and redox status between sows with short and long farrowing duration

Farrowing duration is a crucial factor affecting survival of piglets and health of sows, and is highly correlated with the incidence of stillbirth. The present study assessed the metabolic characteristics of sows with short farrowing duration (SFD) or long farrowing duration (LFD). A total of 20 Yorkshire sows were screened from 60 sows and were retrospectively allocated into SFD (211 min on average, n = 10) or LFD (388 min on average, n = 10) group. Parameters associated with energy metabolism and redox status were characterised. Results showed that sows at farrowing had decreased plasma concentrations of glucose, triglyceride, acetate, butyrate and total short-chain fatty acids (P < 0.05), but increased concentrations of lactic acid and propionate (P < 0.05), when compared with sows on day 107 of gestation. The SFD sows had shorter time from last meal until the onset of farrowing (P < 0.05) and tended to have less stillbirths (P = 0.08) and lower stillbirth rate (P = 0.07). For the blood metabolites, SFD sows at farrowing had higher concentration of plasma glucose (P < 0.05), but lower concentration of lactic acid (P < 0.05) than LFD sows. Besides, SFD sows tended to have higher plasma malondialdehyde concentration (P = 0.06) than LFD sows. Correlation analysis showed that farrowing duration was negatively correlated with plasma glucose concentration at onset of farrowing. In conclusion, our study strongly suggests that glucose is a key metabolite for energy metabolism of the uterus during farrowing. The farrowing process could be closely related to uterine energy expenditure, and sows with shorter farrowing duration could be resulting from the shorter time from last meal until the onset of farrowing, associated with a greater proportion of energy from glucose.     

Regulated redox processes at the plasmalemma of plant root cells and their function in iron uptake

Plants take up iron as ferric chelates or, after reduction, as ferrous ions. Ferric reduction takes place at the plasma membrane of the root epidermis cells by a transmembrane redox system, which can be activated when iron is getting short. It is proposed that this inducible system, with NADPH as electron donor, is separate from a system, presumably present in all plant cells, which transports electrons from NADH or NADPH to ferricyanide, or,in vivo, oygen.     

Cell metabolism: An essential link between cell growth and apoptosis

Growth factor-stimulated or cancerous cells require sufficient nutrients to meet the metabolic demands of cell growth and division. If nutrients are insufficient, metabolic checkpoints are triggered that lead to cell cycle arrest and the activation of the intrinsic apoptotic cascade through a process dependent on the Bcl-2 family of proteins. Given the connections between metabolism and apoptosis, the notion of targeting metabolism to induce cell death in cancer cells has recently garnered much attention. However, the signaling pathways by which metabolic stresses induce apoptosis have not as of yet been fully elucidated. Thus, the best approach to this promising therapeutic avenue remains unclear. This review will discuss the intricate links between metabolism, growth, and intrinsic apoptosis and will consider ways in which manipulation of metabolism might be exploited to promote apoptotic cell death in cancer cells. This article is part of a Special Issue entitled Mitochondria: the deadly organelle.     

The compartmentalization and translocation of the sphingosine kinases: Mechanisms and functions in cell signaling and sphingolipid metabolism

Members of the sphingosine kinase (SK) family of lipid signaling enzymes, comprising SK1 and SK2 in humans, are receiving considerable attention for their roles in a number of physiological and pathophysiological processes. The SKs are considered signaling enzymes based on their production of the potent lipid second messenger sphingosine-1-phosphate, which is the ligand for a family of five G-protein-linked receptors. Both SK1 and SK2 are intracellular enzymes and do not possess obvious membrane anchor domains within their primary sequences. The native substrates (sphingosine and dihydrosphingosine) are lipids, as are the corresponding products, and therefore would have a propensity to be membrane associated, suggesting that specific membrane localization of the SKs could affect both access to substrate and localized production of product. Here, we consider the emerging picture of the SKs as enzymes localized to specific intracellular sites, sometimes by agonist-dependent translocation, the mechanism targeting these enzymes to those sites, and the functional consequence of that localization. Not only is the signaling output of the SKs affected by subcellular localization, but the role of these enzymes as metabolic regulators of sphingolipid metabolism may be impacted as well.