Sugar transport in Coprinus cinereus

Two transport systems for glucose were detected: a high affinity system with a K_m of 27 μM, and a low affinity system with a K_m of 3.3 mM. The high affinity system transported glucose, 2-deoxy-d-glucose (K_m = 26 μM), 3-O-methylglucose (K_m = 19 μM), d-glucosamine (K_m = 652 μM), d-fructose (K_m = 2.3 mM) and l-sorbose (K_m = 2.2 mM). All sugars were accumulated against concentration gradients. The high affinity system was strongly or completely inhibited by N-ethylmaleimide, quercetin, 2,4-dinitrophenol and sodium azide. The system had a distinct pH optimum (7.4) and optimum temperature (45°C). The low affinity system transported glucose, 2-deoxy-d-glucose (K_m = 7.5 mM), and 3-O-methylglucose (K_m = 1.5 mM). Accumulation again occurred against a concentration gradient. The low affinity system was inhibited by N-ethylmaleimide, quercetin and 2,4-dinitrophenol, but not by sodium azide. The rate of uptake by the low affinity system was constant over a wide temperature range (30–50°C) and was not much affected by pH; but as the pH of the medium was altered from 4.5 to 8.9 a co-ordinated increase in affinity for 2-deoxy-d-glucose (from 52.1 mM to 0.3 mM) and decrease in maximum velocity (by a factor of five) occurred. Both uptake systems were present in sporelings germinated in media containing sodium acetate as sole carbon source. Only the low affinity system could initially be demonstrated in glucose-grown tissue, although the high affinity system was restored by starvation in glucose-free medium. The half-time for restoration of high affinity activity was 3.5 min and the process was unaffected by cycloheximide. Addition of glucose to an acetate-grown culture inactivated the high affinity system with a half-life of 5–7.5 s. Addition of cycloheximide to an acetate-grown culture caused decay of the high affinity system with a half-life of 80 min. Regulation is thus thought to depend on modulation of protein activity rather than synthesis, and the kinetics of glucose, 2-deoxy-d-glucose and 3-O-methylglucose uptake would be consistent with there being a single carrier showing negative co-operativity.     

Kinetics of mitochondrial phosphate transport and rates of respiration and phosphorylation during greening of etiolated Avena leaves

Using the technique of silicone oil filtration of organelles and the inhibitor stop method, the kinetics of transport of inorganic phosphate across the inner mitochondrial membrane were tested in relation to different stages of greening (0 to 24 h) of etiolated laminae of Avena sativa L., and compared to the rates of oxygen consumption and ATP formation. The results demonstrate that there is a pronounced increase in phosphate transport after 3 h of greening, reaching values for V_max (about 17 mol mg protein^-1 h^-1) that are three times as high as those measured with mitochondria from etiolated tissue. This is also mirrored by the rates of respiration and oxidative phosphorylation. After 24 h of light treatment (4 Klx), respiration and ATP formation, as well as V decreased again to levels below those of the etiolated stage. In contrast to V, there was no change in the affinity between inorganic phosphate and the binding sites of the transporting systems involved, as indicated by a rather constant K_m (0.23 mM) for phosphate transport. Of the inhibitors of phosphate transport tested, mersalyl and methyl mercuric iodide were most efficient with identical characteristics of inhibition; but compared to animal mitochondria, the concentrations needed to result in similar amounts of inhibition, were more than ten times higher. The results are discussed with respect to plastid development.Abbreviations BSA bovine serum albumine - CH₃HgJ methyl mercuric iodide - Cyt cytochrome - HEPES N-2-hydroxyethylpiperazine-N-2-ethane-sulfonic acid - MDH malate dehydrogenase - NEM N-ethylmaleimide     

A systematic approach to the simulation of short-term processes in the plant-environment complex

Abstract. A conceptual framework is presented for modelling short-term processes in the plant and its environment as an integrated system. Flows of water, water vapour, heat, momentum, CO₂, soluble carbohydrate and phosphorus are all described by equations of the same general type, i.e. in terms of diffusivity-type parameters, capacities and potential gradients. A representative volume of the crop is divided horizontally into layers and vertically between crop and environment for treatment by a finite-difference method. Vertical flow occurs in the atmosphere, soil, stems and larger roots, andilateral flow between leaves and air, and between finer roots and soil. The interception of direct sunlight and the flux densities of downward and upward diffuse radiation within layers are calculated by a step-wise procedure.
The conversions of materials within the plant are treated as functions of appropriate state variables. Schemes for carbon and phosphorus provide for flow to and from the translocation system, and for photosynthesis, respiration and growth.
A model of a fully-established lucerne crop is described and the sensitivity of model performance to changes in a number of parameter values explored. Simulation runs under varying conditions indicate realistic prediction of diurnal trends.     

Fluorescence Transients as Indicator for the Existence of Regulatory Mechanisms in Photosynthetic Electron Transport

In green algae several characteristic differences in the slope of the fast 685 nm fluorescence transient indicate the existence of different mechanisms for the regulation of the photosynthetic electron transport in vivo with respect to the requirements for ATP and NADPH. Autotrophically cultivated Chlamydobotrys stellata exhibits a normal time curve of the fluorescence yield. Anaerobiosis and C0₂-deficiency raise the O-, I- and S-level, whereas the P- level is lowered and the I-D-decay disappears. The readdition of oxygen increases the fluorescence significantly. Supplementation of aerobic cells with CO₂ restores the normal fluorescence transients. The replacement of carbon dioxide by acetate as a carbon source in the light lowers the overall fluorescence emission and abolishes the D-P-increase and the P-S-decline. The presence of DCMU increases fluorescence only at high intensities of incedent light. Anaerobiosis in these photoheterotrophic algae lowers the fluorescence emission. In this case DCMU increases fluorescence even at low light intensities. In Gonium multicoccum, which shows a normal fluorescence transient when cultivated autotrophically, CO₂-deficiency abolishes the O-level and increases the I- and S-niveau. Additional anaerobiosis in CO₂-deficient cells raises the steady state emission. Readdition of oxygen to these cells raises the I- and S-level even more and prevents the build up of the P-level. In Gonium     

Energetics of galactose,proline, and glutamine transport in a cytochrome-deficient mutant of salmonella typhimurium

The effect of inhibitors and uncouplers on the osmotic shock-sensitive transport systems for glutamine and galactose (by the β-methyl galactoside permease) was compared to their effect on the osmotic shock-resistant proline and galactose permease systems in cytochrome-deficient cells of Salmonella typhimurium SASY28. Both osmotic shock-sensitive and -resistant systems were sensitive to uncouplers and to inhibitors of the membrane-bound Ca²⁺, Mg²⁺-activated adenosine triphosphatase. This suggests that uptake by both types of systems is energized in these cells by an electrochemical gradient of protons formed by ATP hydrolysis through the ATPase.     

Effects of amphotericin B on the electrical properties ofNecturus gallbladder: Intracellular microelectrode studies

Summary Intracellular microelectrode techniques were employed to study the mechanism by which amphotericin B induces a transient mucosa-negative transepithelial potential (V _ms) in the gallbladder ofNecturus. When the tissue was incubated in standard Na-Ringer's solution, the antibiotic reduced the apical membrane potential by about 40 mV, and the basolateral membrane potential by about 35 mV whereas the transepithelial potential increased by about 5 mV. The electrical resistance of the apical membrane fell by 83%, and that of the basolateral membrane by 40%; the paracellular resistance remained unchanged. Circuit analysis indicated that the equivalent electromotive forces of the apical and basolateral membranes fell by 35 and 11 mV, respectively. Changes in potentials and resistances produced by ionic substitutions in the mucosal bathing medium showed that amphotericin B produces a nonselective increase in apical membrane small monovalent cation conductance (K, Na, Li). In the presence of Na-Ringer's on the mucosal side, this resulted in a reduction of the K permselectivity of the membrane, and thus in a fall of its equivalent emf. During short term exposure to amphotericin B,P _Na/P _Cl across the paracellular pathway did not change significantly, whereasP _K/P _Na doubled. These results indicate that V _ms is due to an increase of g_Na across the luminal membranes of the epithelial cells (Cremaschiet al., 1977,J. Membrane Biol. 34:55); the data do not support the alternative hypothesis (Rose & Nahrwold, 1976.J. Membrane Biol. 29:1) that V _ms results from a reduction in shuntP _Na/P _Cl acting in combination with a rheogenic basolateral Na pump.     

Stereospecificity of peptide transport by germinating barley embryos

The stereospecific requirements for peptide transport in the scutellum of germinating barley (Hordeum vulgare) embryos are described. Replacement of an L-amino acid residue in a peptide by its D-stereoisomer decreases the affinity of the peptide for the transport site, leading to a reduction in transport. Substitution of a second D-residue reduces affinity still further. The extent to which transport is inhibited depends upon the position of the D-residue in the primary sequence, with D-residues at the C-terminus of the peptide having the greatest effect. Competition between D- and L-peptides indicates that they both enter via the same transport system. Although D-amino acids can be accumulated when presented as a peptide, these same D-residues are not transported when supplied as the free amino acids. L-Leu-D-leu is accumulated intact against a concentration gradient, indicating the operation of an active transport mechanism that can function without the involvement of peptidase activity.     

Electrophysiological properties of onion guard cells

Intracellular electrical recordings in onion (Allium cepa L.) guard cells show that they maintain a membrane potential difference (MPD), inside negative. The MPD of intact cells averaged -72±29 mV (n=45); MPD of cells partially digested with a cellulolytic enzyme, -39±7 mV (n=65). Evidence indicates that the guard cells have two electrically distinct compartments, presumably delimited by the plasmalemma and tonoplast. Epidermal cells in partially digested preparations also showed MPD that could be either positive (+15±7 mV; n=23) or negative (-15 ±8 mV; n=13). Guard cells exposed to light-dark cycles hyperpolarized in the light and depolarized in the dark. The largest observed voltage changes reached 52 mV during hyperpolarizations and 60 mV during depolarizations. The light responses saturated with roughly exponential kinetics, with the depolarizations exhibiting a slower second phase that might be related to the contracting movements of the guard cells. Initial rates of the responses averaged about 14 mV min^-1 in the dark and about 8 mV min^-1 in the light. The results can be interpreted as electrical correlates of fluctuations in intracellular potassium concentration, as light-induced changes in membrane permeability, or as the photoactivation of an electrogenic proton pump. The last possibility seems to be the simplest interpretation of the data that also provides us with a mechanism driving the ion fluxes associated with stomatal function.     

Oxidation and reduction of plastoquinone by photosynthetic and respiratory electron transport in a cyanobacterium Synechococcus sp.

The I-D dip, an early transient of the fluorescence induction, was examined as a means to monitor redox changes of plastoquinone in cells of a cyanobacterium, Synechococcus sp. That the occurrence of the dip depends upon the reduced state of the plastoquinone pool was indicated by observations that 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone and 3-(3,4-dichlorophenyl)-1,1-dimethylurea did not affect the initial rise to I but abolished the subsequent decline from I to D and that illumination of the cells with light 1, prior to fluorescence measurements, eliminated the transient. The I-D dip was prominent in freshly harvested cells containing abundant endogenous substrates, disappeared slowly as the cells were starved by aeration but reappeared on addition of fructose to the starved cells in the dark. The dip that had been induced by a brief illumination of the starved cells with light 2 was rapidly diminished in the dark and KCN inhibited the dark decay of the transient. The results indicate that plastoquinone is reduced with endogenous as well as exogenous substrates and oxidized by a KCN-sensitive oxidase in the dark, thus providing strong support for the view that plastoquinone of photosynthetic electron transport also functions in respiration. In addition, the occurrence of a cyclic pathway of electrons from Photosystem I to plastoquinone, possibly via ferredoxin or NADP, was suggested. Several lines of evidence indicate that, under a strong light 2, Photosystem I-dependent oxidation of plastoquinone predominates over Photosystem II-dependent reduction of the quinone in the cyanobacterium which contains Photosystem I more abundantly than Photosystem II.     

Biphasic uptake of iron-transferrin complex by L1210 murine leukemia cells and rat reticulocytes

The kinetics of the cellular uptake of iron-transferrin complex was studied in L1210 murine leukemia cells and rat reticulocytes using ¹²⁵I-transferrin. Saturation of transferrin with iron was necessary for optimal uptake. Following the incubation of cells with the radiolabeled complex a biphasic pattern of uptake was observed. The initial phase was rapid and relatively temperature-independent and was not altered by ethylamine, an inhibitor of transglutaminase activity which is necessary for receptor-mediated endocytosis. This phase was considered to result from receptor-ligand interaction which could be reversed to a great degree by replacement with unlabeled transferrin. A plateau was then reached, indicating a saturation of receptors. After 30 min a second phase of uptake was indicated by the second rise in the curve. This phase was slow, relatively temperature-dependent and could be abolished by ethylamine. It was interpreted as evidence of internalization of the ligand. Analysis of the data from competition studies with unlabeled transferrin indicated that the first phase might itself comprise a reversible and an irreversible step with a ratio of 5 to 1.4 for bound transferrin. Thus, the cellular uptake of iron-transferrin complex may consist of a reversible ligand-receptor interaction. Conformational changes may render this interaction irreversible and the internalization of the ligand may then follow.