Accumulation of potassium by differentiating metaxylem elements of maize roots

The previous demonstration that the large late metaxylem vessels of field-grown maize ( Zea mays L. cv. Rosella) roots do not lose their crosswalls until they are 20–30 cm from the tip, and that the presence of a soil sheath outside the root was indicative of immature vessels within, greatly strengthened the hypothesis that ion accumulation into these roots was by uptake into living xylem element vacuoles. Proposals that salt movement into the xylem was by leakage or secretion into dead vessels became much less plausible. Potassium concentration in the vacuoles of late metaxylem elements was measured by X-ray microanalysis in unetched fracture faces of bulk, frozen-hydrated pieces of sheathed roots, and found to be in the range 150–400 m M . Potassium concentration in open vessels of bare roots, measured both with the microprobe and by spectrophotometry of aspirated sap, was in the range of 5 to 25 m M . It is concluded that uptake of potassium (and possibly other ions) is into living xylem elements, and that its release to the transpiration stream occurs by the breakdown of their crosswalls and the addition of their vacuoles to the solution in the vessels above.     

The involvement of high-energy phosphate in 2-deoxy-d-glucose transport in Kluyveromyces marxianus

Under aerobic conditions 2-deoxy-d-glucose was accumulated in Kluyveromyces marxianus mainly in a phosphorylated form. During sugar uptake both ATP, polyphosphate and orthophosphate levels decreased. Under anaerobic conditions considerably less sugar was taken up. The intracellular free sugar concentration did not exceed the medium concentration, whereas sugar phosphorylation leveled off at about 3 μmol/g yeast. In response to anaerobic 2-deoxy-d-glucose uptake only ATP and polyphosphate appeared to decrease. Within the experimental error sugar phosphorylation was counterbalanced by the polyphosphate decrease. Pulse labeling experiments revealed transport-associated phosphorylation under these anaerobic conditions, Further, kinetic studies on permeabilized cells showed that cytoplasmic ATP could not be the phosphoryl donor in this transport-associated phosphorylation. These results confirm and extend previous observations, indicating that polyphosphate plays a crucial role in 2-deoxy-d-glucose transport in Kluyveromyces marxianus.     

Protoporphyrin-induced photodynamic effects on band 3 protein of human erythrocyte membranes

In previous studies it has been shown that protoporphyrin-induced photodynamic effects on red blood cells are caused by photooxidation of amino acid residues in membrane proteins and by the subsequent covalent cross-linking of these proteins. Band 3, the anion transport protein of the red blood cell membrane, has a relatively low sensitivity to photodynamic cross-linking. This cannot be attributed to sterical factors inherent in the specific localization of band 3 in the membrane structure. Solubilized band 3, for instance, showed a similar low sensitivity to cross-linking. By extracellular chymotrypsin cleavage of band 3 into fragments of 60 000 and 35 000 daltons it could be shown that both fragments were about equally sensitive to photodynamic cross-linking. The 17 000 dalton transmembrane segment, on the other hand, was completely insensitive. Inhibition of band 3-mediated sulfate transport proceeded much faster than band 3 interpeptide cross-linking, presumably indicating that the inhibition of transport is caused by photooxidation of essential amino acid residues or intrapeptide cross-linking. A close parallel was observed between photodynamic inhibition of anion transport and decreased binding of 4,4′-diisothiocyanodihydrostilbene-2,2′-disulfonate (H₂DIDS), suggesting that a photooxidation in the immediate vicinity of the H₂DIDS binding site may be responsible for transport inhibition.     

Protoporphyrin-induced photodynamic effects on transport processes across the membrane of human erythrocytes

Previous studies have shown that illumination of erythrocytes with visible light in the presence of protoporphyrin results in cross-linking of membrane proteins and deterioration of several membrane functions, e.g. active transport of K⁺ and Na⁺.In the present study it is shown that carrier-mediated transport of glucose, l-leucine, sulphate and glycerol is also inhibited by the photodynamic process, whereas non-specific permeability of glycerol and thiourea is increased.It is shown that these effects are not caused by lipid peroxidation, but by photooxidation of membrane proteins. The inhibition of carrier-mediated transport is caused either by photodynamic oxidation of susceptible essential amino acid residues of the carrier molecules, or by an aspecific perturbation of the membrane structure, leading to inhibition of carrier functions.     

Effects of salinity and phosphate on ion distribution in lupin leaflets

Lupin ( Lupinus luteus L. cv. Weiko III) were grown in nutrient solution over a range of inorganic phosphate (P_i) concentrations, with or without 50 m M NaCl. Plants with high P_i (2 m M ) and salt showed progressive leaf necrosis and had higher concentrations of total phosphate than plants grown with high P_i alone. Most of the extra total phosphate in salt treated plants was in the P_i form. P_i supply did not influence Na⁺, K⁺ or Cl⁻ concentrations in epidermal vacuoles or mesophyll cells. However, epidermal vacuoles accumulated more monovalent cations (Na⁺ and K⁺) than Cl⁻, and in vacuoles of plants grown with 0.1 m M P_i additional P_i was accumulated, possibly to maintain charge balance. Plants grown with 2 m M P_i did not accumulate additional P_i in epidermal vacuoles, but showed higher phosphorus levels in cell walls. It is suggested that at moderate phosphorus concentrations P_i plays a role in epidermal osmotic adjustment, possibly explaining the beneficial role of additional phosphorus on salt stressed plants. At high P_i supply with salt, P_i does not contribute to osmotic adjustment and instead accumulates in cell walls. However, the cause of leaf damage under conditions of high phosphorus supply and salinity is still not entirely clear.     

Kinetic analysis of l-lactate transport in human erythrocytes via the monocarboxylate-specific carrier system

Three parallel pathways of l-lactate transport across the membrane of human red blood cells can be discriminated: (a) by nonionic diffusion; (b) via the band 3 anion exchange protein; and (c) via a specific monocarboxylate carrier system. Influx of lactate via the latter system leads to alkalinization of the medium, suggesting lactate-proton symport. Kinetic analysis of initial lactate influx via the monocarboxylate carrier indicates a symport system with ordered binding of the two ligands, in the sense that a proton binds first to the translocator, followed by lactate binding to the protonated carrier. The influence of varying trans-pH under conditions of net (zero-trans) flux with constant cis-pH indicates that the monocarboxylate translocator should be considered as a mobile carrier, with the ligand-binding sites exposed alternately to the outside and the inside of the membrane.     

Localization of polyphosphates in Saccharomyces fragilis, as revealed by 4′,6-diamidino-2-phenylindole fluorescence

The fluorescent dye 4′,6-diamidino-2-phenylindole has its emission maximum at 456 nm. Fluorescence intensity at this wavelength is significantly increased by various negatively-charged polyelectrolytes. Among several polyelectrolytes tested, polyphosphates appeared to be unique in the sense that they shifted the emission maximum from 456 to 526 nm. Addition of Saccharomyces fragilis cells to a diamidinophenylindole solution caused an immediate shift of the emission maximum to 526 nm, followed by a gradual increase of fluorescence at 456 nm. The 526 nm, but not the 456 nm fluorescence was instantly quenched by non-penetrating cations, like UO²⁺₂. These results suggest a momentary interaction of diamidinophenylindole with polyphosphate, localized outside the plasma membrane, followed by a slow penetration of the dye into the cells, yielding increased fluorescence at 456 nm by interaction of the dye with e.g., nucleic acids. This was confirmed by fluorescence microscopy. After addition of diamidinophenylindole the yeast cells exhibited an immediate green-yellow fluorescence of the membrane, that was suppressed by UO²⁺₂. After longer incubation times the cytoplasm and nucleus developed a blue fluorescence.     

Inhibition of lipid peroxidation by disulfiram and diethyldithiocarbamate does not prevent hepatotoxin-induced cell death in isolated rat hepatocytes. A study with allyl alcohol, tert-butyl hydroperoxide, diethyl maleate, bromoisovalerylurea and carbon tetrachloride

The relationship between lipid peroxidation and cell death, induced by a number of hepatotoxins, was studied in isolated rat hepatocytes. Disulfiram (DSF) and diethyldithiocarbamate (DDC) completely prevented lipid peroxidation, induced by allyl alcohol, tert-butyl hydroperoxide (t-BHP), diethyl maleate (DEM), bromoisovalerylurea (BIU) and carbon tetrachloride (CCl4). Lipid peroxidation was measured by the formation of both thiobarbituric acid positive material and conjugated dienes. However, DSF and DDC did not protect against cell death, induced by these hepatotoxins. In the presence of DSF or DDC, cell death occurred even earlier in time. We conclude that cell death can occur in the absence of lipid peroxidation. Therefore, lipid peroxidation is not a requisite for the induction of cell death.     

Xanthine oxidase-catalysed oxidation of paracetamol.

[3H]Arachidonic acid is released after stimulation of rabbit neutrophils with fMet-Leu-Phe or platelet-activating factor (PAF). The release is rapid and dose-dependent, and is inhibited in phorbol 12-myristate 13-acetate (PMA)-treated rabbit neutrophils. The protein kinase C (PKC) inhibitor 1-(5-isoquinoline-sulphonyl)-2-methylpiperazine (H-7) prevents this inhibition. In addition, PMA increases arachidonic acid release in H-7-treated cells stimulated with fMet-Leu-Phe. [3H]Arachidonic acid release, but not the rise in the concentration of intracellular Ca2+, is inhibited in pertussis-toxin-treated neutrophils stimulated with PAF. The diacylglycerol kinase inhibitor R59022 increases the concentration of diacylglycerol and potentiates [3H]arachidonic acid release in neutrophils stimulated with fMet-Leu-Phe. This potentiation is not inhibited by H-7. These results suggest several points. (1) A rise in the intracellular concentration of free Ca2+ is not sufficient for arachidonic acid release in rabbit neutrophils stimulated by physiological stimuli. (2) A functional pertussis-toxin-sensitive guanine nucleotide regulatory protein and/or one or more of the changes produced by phospholipase C activation are necessary for arachidonic acid release produced by physiological stimuli. (3) Agents that stimulate PKC potentiate arachidonic acid release, and this potentiation is not inhibited by H-7. These agents produce their actions in part by direct membrane perturbation.     

The Role of Polyphosphates in the Transport Mechanism of Glucose in Yeast Cells

Several cations inhibit anaerobic fermentation of glucose by intact yeast cells. Some ions (e.g. Hg⁺⁺) penetrate into the cytoplasm and cause an irreversible inhibition of fermentation. Other ions (e.g. UO₂⁺⁺, Ni⁺⁺, and Co⁺⁺) are reversibly bound to a substance at the outside of the yeast cell identified as polyphosphate. Although the cations are bound to exactly the same extent, their influences on fermentation differ greatly. Thorium ions are bound not only to the polyphosphates, but in addition, to phosphatides in the cell membrane. Under circumstances in which glucose is transported into the cell, the amount of polyphosphate in the outer face of the membrane decreases considerably. If yeast is poisoned with monoiodoacetate, the number of glucose molecules that can still be taken up equals the original number of cation-binding sites at the outer surface of the membrane. These data suggest that one molecule of glucose is taken up in connection with the disappearance of one polyphosphate monomer. The hypothesis is framed that the uptake of glucose into the yeast cell is associated with an enzymic phosphorylation (possibly of the carrier), with polyphosphate as phosphate donor. The inhibition of glucose uptake caused by certain metal ions may be the consequence of induced changes in the spatial arrangement of polyphosphate chains; the greater the change in configuration, the larger is the inhibition.