The Senescence Process in Oat Leaves and its Regulation by Oxygen Concentration and Light Irradiance

The regulation of senescence by oxygen-concentration, lightirradiance and H₂O₂ has been studied in leaf segments of Avenasativa L. cv. Suregrain. The development of the components of the senescence process,for example chlorophyll breakdown, proteolysis (as soluble aminoacids), hydroperoxides (as malondi-aldehyde) and permeability(as conductivity) is accelerated in light as the O₂-tensionincreases. In darkness, 0.3% O₂ accelerates increases in hydroperoxides,permeability and proteolysis and delays the chlorophyll break-down,but 0.0005% O₂ delays all the components studied. In every casethe hydroperoxide content, permeability and proteolysis areclosely related. Any treatment inducing an increase in membranepermeability causes chlorophyll bleaching (photo-oxidation)if leaf segments are then treated with light in an atmospherecontaining oxygen. Light has a modulating effect on the senescenceprocess. An irradiance lower or higher than 40 W m^–2 hasan accelerating effect on the senescence process. (Received September 7, 1985; Accepted July 30, 1985)     

Hydroxyl Radicals and a Thylakoid-Bound Endopeptidase are Involved in Light- and Oxygen-Induced Proteolysis in Oat Chloroplasts

The hypothesis that light- and oxygen-induced proteolysis inchloroplasts is mediated by active oxygen species was examined.In order to determine whether or not H₂O₂ and/or ^{dot}OH radicalsare involved in these degradative processes we compared thedegradation of proteins in isolated oat chloroplasts exposedto white light at 80 W m^-2 with that in chloroplasts incubatedin darkness in the absence or presence of H₂O₂ or a ^{dot}OH-generatingsystem composed by ascorbic acid, FeCl₃ and H₂O₂ (Asc-Fe-H₂O₂).Light enhanced the rate of degradation of at least 18 polypeptides,while proteolysis was almost negligible in darkness in the abscenceof additives. H₂O₂ had a very small effect. However, Asc-Fe-H₂O₂-treatedchloroplasts in darkness showed a pattern of protein degradationalmost identical to that observed in the light. A thylakoid-boundendopeptidase (EP), the activity of which increased under photooxidativeenvironmental conditions and treatment with an ^{dot}OH-generatingsystem, was partially purified and characterized as a serinetypeprotease. Treatments with inhibitors of serine-type proteaseprevented both light- and Asc- Fe-H₂O₂-induced proteolysis.EP was more active against both soluble and membranous proteinsthat had been pretreated with Asc-Fe-H₂O₂ than against untreatedproteins. It is proposed that a high dose of light irradiationpromotes proteolysis by increasing the formation of ^{dot}OH,which may modify proteins such that they become more susceptibleto EP-catalyzed hydrolysis. ¹Fisiología Vegetal, Dept. de Biología Vegetal,Universidad de Alcalá de Henares, Present address: 28871Alcalá de Henares (Madrid), España.     

Nitric oxide increases toxicity of hydrogen peroxide against rat liver endothelial cells and hepatocytes by inhibition of hydrogen peroxide degradation

Nitric oxide (NO) and hydrogen peroxide (H₂O₂) show cooperativity in their cytotoxic action. The present study was performed to decipher the mechanisms underlying this phenomenon. In cultured liver endothelial cells and in cultured, glutathione-depleted hepatocytes, the combined exposure to NO (released by spermine NONOate, 1 mM) and H₂O₂ (released by glucose oxidase) induced cell injury that was far higher than the injury elicited by NO or H₂O₂ alone. In both cell types, the addition of the NO donor increased H₂O₂ steady-state levels, although with different kinetics: in hepatocytes, the increase in H₂O₂ levels was already evident at early time points while in liver endothelial cells it became evident after 2 h of incubation. NO exposure inhibited H₂O₂ degradation, assessed after addition of 50 µM, 200 µM, or 4 mM authentic H₂O₂, significantly in both cell types. However, again, early and delayed inhibition was observed. The late inhibition of H₂O₂ degradation in endothelial cells was paralleled by a decrease in glutathione peroxidase activity. Glutathione peroxidase inactivation was prevented by hypoxia or by ascorbate, suggesting inactivation by reactive nitrogen oxide species (NO_x). Early inhibition of H₂O₂ degradation by NO, in contrast, could be mimicked by the catalase inhibitor azide. Together, these results suggest that the cooperative effect of NO and H₂O₂ is due to inhibition of H₂O₂ degradation by NO, namely to inhibition of catalase by NO itself (predominant in hepatocytes) and/or to inhibition of glutathione peroxidase by NO_x (prevailing in endothelial cells). nitrogen monoxide; catalase; glutathione peroxidase     

Dormancy in Seeds of Charlock: IV. INTERRELATIONSHIPS OF GROWTH, OXYGEN SUPPLY AND CONCENTRATION OF INHIBITOR

Respiration measurements were made over a period of 24 h at25 °C on seeds and excised embryos maintained in Warburgflasks with partial pressures of oxygen ranging from 0 to 1atm. In the initial phase (0 to 4 h), the rate of oxygen uptake(Q_O2 of excised embryos increased linearly with external oxygenconcentration (C_O from 0 to 0.1 atm O₂ from 0.1 to 0.2 atm O₂the relation was curvilinear, and from 0.2 to 1.0 atm O₂ uptakewas independent of concentration. In later stages the relationbetween Q_O2 and C_o changed, and from 20 to 24 h the rate ofoxygen uptake increased with concentration to 1.0 atm O₂. Thechanges with time were associated with increase in rate of respiration,increase in cell size and cell number, and the oxidation offats. The decline in concentration of oxygen from the surfaceto the centre of embryos was calculated to be relatively smallat each external oxygen concentration. Althugh the rate of diffusionfailed to keep pace with consumption, the main parameters whichdetermined the internal oxygen status of the embryos were thesurface concentrations and the permeability of the seed coat.The resistance of the seed coat to diffusion of oxygen was foundto be very high, the coefficient of diffusion being about 10^–7mm² s^–1. The concentration of oxygen and in air were estimatedto be approximately 0.04 and 0.02 atm O₂, respectively. Sincea smaller concentration of oxygen (0.012 atm O₂) in the tissueswas found to be sufficient for growth, the dormancy of the seedswas not due to lack of oxygen. Dormancy appeared to be due tothe activity of growth-inhibiting substances, the concentrationof which increased with decrease in oxygen supply; below 0.1atm O₂ their rate of production increased with decrease in theoxygen concentration of the tissues. They accumulated withinthe testas of dormant seeds and prevented cell elongation. Extractsof the inhibitory substances were partially purifield by partitioningthe aqueous fraction with ether and separating chromatographically.The active principle(s) was not abscisic acid ((+)—AbscisinII, ‘Dormin’) or the mustard oil, allylisothiocyanate.     

外源NO对南方红豆杉幼苗光合色素及抗氧化酶的影响

以4年生南方红豆杉幼苗为实验材料,通过对南方红豆杉幼苗喷施不同浓度外源一氧化氮（NO）供体硝普钠溶液（0、0.01、0.1、0.5和1 mmol·L^-1SNP）,测定光合色素含量、抗氧化酶活性、丙二醛（MDA）含量和过氧化氢（H₂O₂）含量等生理指标,以探讨不同浓度外源NO对南方红豆杉叶片光合色素和抗氧化酶的影响。结果表明：喷施低浓度（0.01、0.1 mmol·L^-1）SNP可显著提高南方红豆杉叶片的叶绿素a、叶绿素b、类胡萝卜素和总叶绿素含量,增加叶绿素a/b的比值,而喷施高浓度（0.5、1 mmol·L^-1）SNP降低了叶片的光合色素含量。随着外源NO供体浓度的增加,叶片过氧化氢酶(CAT)活性显著增加,过氧化物酶(POD)活性先增加后降低。此外,处理前期,低浓度SNP处理明显提高了抗坏血酸过氧化物酶(APX)活性,而高浓度SNP处理显著降低了APX活性,处理后期APX活性随SNP浓度的增加而显著下降。喷施低浓度SNP可有效提高超氧化物歧化酶(SOD)活性和增加可溶性蛋白含量,降低MDA和H₂O₂的含量,而喷施高浓度SNP显著增加了MDA和H₂O₂的含量。因此,低浓度的SNP（<0.5 mmol·L^-1）处理南方红豆杉幼苗,可增加其叶绿素含量,提高抗氧化酶活性,降低MDA和H₂O₂的含量,而高浓度的SNP（≥0.5 mmol·L^-1）处理会降低叶绿素含量,提高H₂O₂含量,增加细胞膜质过氧化程度,从而对南方红豆杉幼苗造成一定伤害。     

外源H2O2处理对唐古特白刺愈伤组织脯氨酸代谢的影响

以唐古特白刺（Nitraria tangutorum Bobr.）愈伤组织为材料,研究外源H₂O₂（2和10 μmol·L^-1）处理下其脯氨酸含量及相关代谢酶活性的变化,试图从细胞水平揭示H₂O₂影响脯氨酸代谢的生理机制。结果显示,2和10 μmol·L^-1 H₂O₂处理24 h使唐古特白刺愈伤组织脯氨酸含量分别变为对照的112%和92%,而处理72 h后,脯氨酸含量增加为对照的141%和119%;与对照相比,外源H₂O₂处理诱导愈伤组织脯氨酸脱氢酶活性降低,而谷氨酸激酶活性升高,但鸟氨酸转氨酶活性无显著变化;此外,H₂O₂处理使唐古特白刺愈伤组织内源性H₂O₂含量升高。结果表明,外源H₂O₂诱导了唐古特白刺愈伤组织H₂O₂含量的增高和脯氨酸的积累,且H₂O₂处理下脯氨酸脱氢酶活性的降低及谷氨酸激酶的升高与愈伤组织脯氨酸的积累有关。     

镉胁迫对旱柳光合作用和内肽酶变化的影响

通过水培方法,添加不同浓度CdCl2(0、5、25、50 μmol·L^-1)处理14 d,测定叶绿素含量、核酮糖-1,5-二磷酸羧化酶/加氧酶(Rubisco)和磷酸烯醇式丙酮酸羧化酶(PEPC)活性、游离氨基酸含量及内肽酶活性,调查Cd对旱柳光合作用和内肽酶变化活性影响。结果发现：Cd处理降低了总叶绿素、叶绿素a、b含量; Rubisco活性随着介质中Cd浓度增加而降低;Cd抑制根和叶PEPC活性;同对照相比,根中游离氨基酸含量没有显著变化,而叶中游离氨基酸含量增加;不同浓度Cd处理降低根的内肽酶活性,高浓度Cd使叶内肽酶活力增加。这些结果表明,Cd通过降低叶绿素含量,促进叶内肽酶活性和抑制了CO₂羧化酶活性来影响旱柳光合作用。     

Lipid Breakdown in Smooth Microsomal Membranes from Bean Cotyledons Alters Membrane Proteins and Induces Proteolysis

The effects of lipid degradation on proteins of smooth microsomalmembranes isolated from young bean cotyledons have been examinedby three techniques, viz. fluorescence energy transfer fromtryptophan to cis-parinaric acid; protein spin-labelling with3-maleimido PROXYL; and SDS-PAGE. Lipid degradation was inducedin isolated membranes by activating phospholipase D and phosphatidicacid phosphatase through the addition of Ca²⁺, by treatmentwith exogenous phospholipase C to simulate the concerted actionsof phospholipase D and phosphatidic acid phosphatase or by treatmentwith exogenous phospholipase A₂ to generate endogenous substratefor lipoxygenase. All of the treatments induced time-dependentchanges in lipid-protein interaction and in protein conformation,and the treatment with phospholipase A₂ also engendered proteolysis.The effects of the Ca²⁺ and phospholipase C treatments on lipid-proteininteraction and protein conformation can presumably be partlyattributed to an accumulation of diacylglycerol in the membrane,whereas the induction of proteolysis by phospholipase A₂ appearsto be due to activated oxygen derived from the lipoxygenasereaction and ensuing lipid peroxidation. Lipid degradation inducedby these treatments simulates that which occurs during naturalsenescence of the cotyledons and hence these observations suggestthat loss of protein function and proteolysis in senescing membranesis facilitated by lipolytic and peroxidative activity withinthe lipid bilayer. Key words: Activated oxygen, lipids, membranes, proteins, senescence     

Ammonium release by nitrogen sufficient diatoms in response to rapid increases in irradiance

It has been hypothesized that nitrogen-replete diatoms, butnot flagellates, may release NO₂^–, NH₄⁺ or dissolved organicnitrogen (DON) following rapid increases in irradiance (andconsequently an increase in cellular electron energy), as mightbe expected to occur in a vertically well mixed estuarine system.Just as the increase in irradiance leads to an increase in cellularenergy, so too would a decrease in temperature, due to the temperaturedependency of biosynthetic enzymes. This hypothesis was testedby comparing the response of nitrogen-replete diatoms (Skeletomenacostatum, Thalassiosira weissflogii and Chaetoceros sp.) andflagellates (Dunaliella tertiolecta, Pavlova lutheri and Prorocentrumminimum) to rapid increases in irradiance and decreases in temperature.Short-term (<3 h) changes in extracellular NO₂^– andNH₄⁺ concentrations were measured in cultures following theseexperimental shifts, as well as in cultures retained at thegrowth irradiance. Net rates of NO₂^– and NH₄⁺ releasewere calculated from the time course of extracellular nitrogenconcentrations. As a fraction of NO₃^– uptake, NO₂^–release rates under the increased irradiance increased marginallyrelative to NO₂^– release rates under the growth irradiance.Release rates of NH₄⁺ under the increased irradiance increasednearly fivefold over release rates at the growth irradiance,and accounted for 84% of the NO₃^– uptake rate. In directcontrast to the diatom species, the flagellate species releasedNO₂^– under the higher experimental irradiance at ratesone half those of the release rates under the growth irradiance,and continued to take up NH₄⁺ under both irradiance conditions.Within the experimental boundaries, these findings have importantphysiological and ecological implications. The magnitude ofthe observed nitrogen release represents a significant physiologicalsink for electrons and, in fact, calculations suggest that upto 62% of the total electrons harvested could be consumed. Froman ecological perspective, these findings add to the body ofliterature which suggests that a significant fraction of thenitrogen that is taken up is ultimately released in dissolvedform. More importantly, these data suggest that DON is not theonly compound that phytoplankton may release in the aquaticenvironment.     

Factors Controlling Induction of Carbonic Anhydrase in Chlorella vulgaris 11h: Effects of CO2 and O2

Increase of carbonic anhydrase activity was enhanced by decreasingthe O₂ concentration when Chlorella vulgaris 11h cells grownunder 3% CO₂2 in ordinary air were transferred to low CO₂ conditions.The carbonic anhydrase activity finally attained under the steadystate was dependent on the CO₂ concentration, irrespective ofthe O₂ concentration used. (Received April 24, 1988; Accepted February 23, 1988)     

Hydrogen peroxide generated by copper amine oxidase is involved in abscisic acid-induced stomatal closure in Vicia faba

H₂O₂ is an essential signal in absicic acid (ABA)-induced stomatalclosure. It can be synthesized by several enzymes in plants.In this study, the roles of copper amine oxidase (CuAO) in H₂O₂production and stomatal closure were investigated. ExogenousABA stimulated apoplast CuAO activity, increased H₂O₂ productionand [Ca²⁺]_cyt levels in Vicia faba guard cells, and inducedstomatal closure. These processes were impaired by CuAO inhibitor(s).In the metabolized products of CuAO, only H₂O₂ could inducestomatal closure. By the analysis of enzyme kinetics and polyaminecontents in leaves, putrescine was regarded as a substrate ofCuAO. Putrescine showed similar effects with ABA on the regulationof H₂O₂ production, [Ca²⁺]_cyt levels, as well as stomatal closure.The results suggest that CuAO in V. faba guard cells is an essentialenzymatic source for H₂O₂ production in ABA-induced stomatalclosure via the degradation of putrescine. Calcium messengeris an important intermediate in this process. Key words: Abscisic acid, calcium, copper amine oxidase, hydrogen peroxide, putrescine, stomatal closure, Vicia faba Received 13 October 2007; Revised 16 December 2007 Accepted 20 December 2007     

Nutrient Dilution by Starch in CO2-enriched Chrysanthemum

Increasing growth irradiance and CO₂ generally decreases foliarnutrient concentration on a dry weight basis and increases foliarstarch concentration. However, the extent to which starch concentrations‘dilute’ foliar nutrient concentrations when thelatter are expressed on a dry weight basis is not known. Todetermine the importance of differential starch accumulationin calculating nutrient concentrations on a dry weight basis,leaf nutrient and starch concentrations were measured in Chrysanthemum? morifolium ‘Fiesta’ (Ramat.) cuttings grown atthree irradiance levels and two CO₂ levels for eight weeks inboth winter and spring. On a dry weight basis, foliar concentrationsof most nutrients were lower in both seasons as a result ofthe elevated CO₂ and irradiance levels, and total dry weightswere higher. Per cent starch was greater at the high CO₂ levelin both seasons but was only greater at higher irradiances inthe winter experiment. When starch was subtracted from the leafdry weights, the differences between CO₂ and irradiance treatmentsdisappeared with respect to N, P, K, Ca, Mg, S, and B but notfor Fe, Mn, Zn, and Cu. Key words: CO₂ enrichment, starch, nutrients, irradiance     

H2O2-mediated permeability II: importance of tyrosine phosphatase and kinase activity

We previously reportedthat exposure of endothelial cells to H₂O₂results in a loss of cell-cell apposition and increased endothelialsolute permeability. The purpose of this study was to determine howtyrosine phosphorylation and tyrosine phosphatases contribute tooxidant-mediated disorganization of endothelial cell junctions. Wefound that H₂O₂ caused a rapid decrease in total cellular phosphatase activity that facilitates a compensatory increase in cellular phosphotyrosine residues.H₂O₂ exposure also results in increasedendothelial monolayer permeability, which was attenuated by pp60, aninhibitor of src kinase. Inhibition of protein tyrosinephosphatase activity by phenylarsine oxide (PAO) demonstrated a similarpermeability profile compared with H₂O₂,suggesting that tyrosine phosphatase activity is important inmaintaining a normal endothelial solute barrier. Immunofluorescence shows that H₂O₂ exposure caused a loss ofpan-reactive cadherin and -catenin from cell junctions that was notblocked by the src kinase inhibitor PP1.H₂O₂ also caused -catenin to dissociate fromthe endothelial cytoskeleton, which was not prevented by PP1. Finally,we determined that PP1 did not prevent cadherin internalization. Thesedata suggest that oxidants like H₂O₂ produce biological effects through protein phosphotyrosine modifications bydecreasing total cellular phosphatase activity combined with increasedsrc kinase activity, resulting in increased endothelial solute permeability.

     

Effects of Oxidative Stress Caused by Oxygen and Hydrogen Peroxide on Energy Metabolism and Senescence in Oat Leaves

In attached oat leaves the levels of adenine nucleotides decreasedduring leaf development and senescence. However, the energycharge (EC) only decreased from 0.90 in 4-cm leaves to 0.80in senescent leaves. In detached leaves the levels of adeninenucleotides increased for 48 h, in association with an increasein RNase activity and a decrease in levels of RNA. The EC remainedhigh until late senescence when levels of adenine nucleotidesfell to about 30% of initial values. A decrease in energy parametersinduced by transfer from light to darkness and from high (21%)to low (0.5% and anoxia) concentrations of oxygen resulted inan increase in membrane permeability. Oxidative stress (above 0.5% O₂ induced an increase in levelsof malondialdehyde (MDA) and then in permeability, associatedwith a decrease in levels of adenine nucleotides. Oxidativestress provoked by 0.05 and 0.10 M H₂O₂ caused a more rapiddecrease in energy parameters than O₂. Under oxidative stress(above 0.5% O₂) there is, first of all, an increase in membranepermeability and then a decrease in energy parameters, whichin turn are involved with senescence via increases in oxidationof membranes and degradation of energy-producing systems. (Received October 6, 1987; Accepted October 19, 1988)     

Effect of Hydrogen Peroxide on Degradation of Cell Wall Associated Proteins in Growing Bean Hypocotyls

The possible involvement of active oxygen species and an apoplasticendopeptidase (EP) in the digestion of cell wall proteins wasstudied in extracellular fluid (EF) from hypocotyls of Phaseolusvulgaris at different stages of elongation. EF proteins underwentsignificant changes in polypeptide pattern during hypocotylgrowth, which were characterized by increases in 35, 39, 40and 50 kDa peptides and appearance of 61, 70 and 75 kDa peptidesat the exponential growth phase. EFs also contain endopeptidase[Gómez et al. (1994) Agriscientia 11:3]. Autolysis experimentswithout or with purified EP revealed that many cell wall polypeptidesare liable to degradation by the protease. Besides, EF polypeptidesincreased their susceptibility to EP during hypocotyl elongation.The 50 and 40 kDa polypeptydes were poorly degraded when extractedfrom hypocotyls in active growth, but greatly hydrolyzed whenextracted from fully elongated tissues, suggesting that in thecourse of growth proteins underwent modifications that renderedthem more prone to proteolytic attack. These modifications seemedto involve active oxygen species, as indicated by: (a) H₂O₂level rised when protein susceptibility to EP increased; and(b) EF proteins from growing hypocotyls (comparatively lesssusceptible to EP) treated with H₂O₂ were rapidly degraded bythe protease. (Received April 27, 1995; Accepted July 31, 1995)     

Regeneration of Ribulose 1,5-bisphosphate and Ribulose 1,5-bisphosphate carboxylase/oxygenase Activity Associated with Lack of Oxygen Inhibition of Photosynthesis at Low Temperature

The nature of the lack of oxygen inhibition of C₃-photosynthesisat low temperature was investigated in white clover (Trifoliumrepens L.). Detached leaves were brought to steady-state photosynthesisin air (34 Pa p(CO²), 21 kPa p(O₂), balance N₂) at temperaturesof 20°C and 8°C, respectively. Net photosynthesis, ribulose1,5-bisphosphate (RuBP) and ATP contents, and ribulose 1,5-bisphosphatecarboxylase/oxygenase (RuBPCO) activities were followed beforeand after changing to 2·0 kPa p(O₂). At 20°C, lowering p(O₂) increased net photosynthesis by37%. This increase corresponded closely with the increase expectedfrom the effect on the kinetic properties of RuBPCO. Conversely,at 8°C net photosynthesis rapidly decreased following adecrease in p(O₂) and then increased again reaching a steady-statelevel which was only 7% higher than at 21 kPa p(O₂). The steady-staterates of RuBP and associated ATP consumption were both estimatedto have decreased. ATP and RuBP contents decreased by 18% and33% respectively, immediately after the change in p(O₂) suggestingthat RuBP regeneration was reduced at low p(O₂) due to reducedphotophosphorylation. Subsequently, RuBP content increased again.Steady-state RuBP content at 2·0 kPa p(O₂) was 24% higherthan at 21 kPa p(O₂). RuBPCO activity decreased by 22%, indicatingcontrol of steady-state RuBP consumption by RuBPCO activity. It is suggested that lack of oxygen inhibition of photosynthesisat low temperature is due to decreased photophosphorylationat low temperature and low p(O₂). This may be due to assimilateaccumulation within the chloroplasts. Decreased photophosphorylationseems to decrease RuBP synthesis and RuBPCO activity, possiblydue to an acidification of the chloroplast stroma. Key words: Oxygen inhibition, photosynthesis, ribulose bisphosphate carboxylase/oxygenase     

大气臭氧浓度升高对水稻叶片膜脂过氧化及保护酶活性的影响

采用开顶式气室（open top chambers,OTCs）装置,以水稻品种“3694繁”(Oryza sativa L.)为材料,研究3种处理：过滤大气（CF,O₃浓度约为20 nl·L^-1）、环境大气（NF,O₃浓度约为40 nl·L^-1）和高浓度O₃（EO,O₃浓度约为75 nl·L^-1）下叶片可溶性蛋白质含量、膜脂过氧化程度与主要保护酶活性的变化.结果表明：过滤大气与环境大气处理之间各个指标无显著差异.与CF处理相比,高浓度O₃处理条件下水稻叶片中可溶性蛋白含量显著下降,而过氧化氢（H₂O₂）和抗坏血酸（ASA）含量显著增加;超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、过氧化物酶（POD）活性升高,最大升高幅度分别为93.7%、39.9%和312.4%;抗坏血酸过氧化物酶（APX）活性呈先升高后下降趋势; 不同O₃处理条件下叶片中丙二醛（MDA)含量无显著变化.表明抗氧化系统可有效抑制水稻叶片中的O₃胁迫引起的膜质过氧化作用,说明水稻品种“3694繁”对O₃胁迫有一定抗性.     

Tetcyclacis and Abscisic Acid-Sensitive Reduction of Extracellular Ferricyanide by Mesophyll Cells of Valerianella locusta and Lemna gibba

Electron transport across the plasma membrane of Valerianellalocusta mesophyll cells and intact fronds of Lemna gibba, inducedby 10^–3 M ferricyanide, was inhibited by tetcyclacis,an inhibitor regarded to be specific for cytochrome P-450 mono-oxygenases.The effect was dependent on the concentration of tetcyclacisand the duration of preincubation. The apparent rate of trans-membraneelectron transport increased in the presence of catalase, indicatingtetcyclacis-induced H₂O₂-production or additional tetcyclacis-independentH₂O₂ release. The findings suggest an interaction of cytochromeP-450 with the plasma membrane-located electron transport chain.This redox-chain could be involved in the degradation of abscisicacid, being located at the plasma membrane. This assumptionis supported by the finding that ABA inhibits extracellularferricyanide reduction. Key words: Abscisic acid, cytochrome P-450 mono-oxygenase, plasma membrane, tetcyclacis     

Glucosamine inhibits angiotensin II-induced cytoplasmic Ca2+ elevation in neonatal cardiomyocytes via protein-associated O-linked N-acetylglucosamine

We previously reported that glucosamine and hyperglycemia attenuate the response of cardiomyocytes to inositol 1,4,5-trisphosphate-generating agonists such as ANG II. This appears to be related to an increase in flux through the hexosamine biosynthesis pathway (HBP) and decreased Ca²⁺ entry into the cells; however, a direct link between HBP and intracellular Ca²⁺ homeostasis has not been established. Therefore, using neonatal rat ventricular myocytes, we investigated the relationship between glucosamine treatment; the concentration of UDP-N-acetylglucosamine (UDP-GlcNAc), an end product of the HBP; and the level of protein O-linked N-acetylglucosamine (O-GlcNAc) on ANG II-mediated changes in intracellular free Ca²⁺ concentration ([Ca²⁺]_i). We found that glucosamine blocked ANG II-induced [Ca²⁺]_i increase and that this phenomenon was associated with a significant increase in UDP-GlcNAc and O-GlcNAc levels. O-(2-acetamido-2-deoxy-D-glucopyranosylidene)-amino-N-phenylcarbamate, an inhibitor of O-GlcNAcase that increased O-GlcNAc levels without changing UDP-GlcNAc concentrations, mimicked the effect of glucosamine on the ANG II-induced increase in [Ca²⁺]_i. An inhibitor of O-GlcNAc-transferase, alloxan, prevented the glucosamine-induced increase in O-GlcNAc but not the increase in UDP-GlcNAc; however, alloxan abrogated the inhibition of the ANG II-induced increase in [Ca²⁺]_i. These data support the notion that changes in O-GlcNAc levels mediated via increased HBP flux may be involved in the regulation of [Ca²⁺]_i homeostasis in the heart. hypertrophy; left ventricle; calcium channels; calcium signaling     

H(2)O(2) and ethanol act synergistically to gate ryanodine receptor/calcium-release channel

We examined theeffect of low concentrations of H₂O₂ on theCa²⁺-release channel/ryanodine receptor (RyR) to determineif H₂O₂ plays a physiological role in skeletalmuscle function. Sarcoplasmic reticulum vesicles from frog skeletalmuscle and type 1 RyRs (RyR1) purified from rabbit skeletal muscle wereincorporated into lipid bilayers. Channel activity of the frog RyR wasnot affected by application of 4.4 mM (0.02%) ethanol. Openprobability (P_o) of such ethanol-treated RyRchannels was markedly increased on subsequent addition of 10 µMH₂O₂. Increase of H₂O₂to 100 µM caused a further increase in channel activity. Applicationof 4.4 mM ethanol to 10 µM H₂O₂-treated RyRsactivated channel activity. Exposure to 10 or 100 µMH₂O₂ alone, however, failed to increaseP_o. Synergistic action of ethanol andH₂O₂ was also observed on the purified RyR1 channel, which was free from FK506 binding protein (FKBP12).H₂O₂ at 100-500 µM had no effect onpurified channel activity. Application of FKBP12 to the purified RyR1drastically decreased channel activity but did not alter the effects ofethanol and H₂O₂. These results suggest thatH₂O₂ may play a pathophysiological, butprobably not a physiological, role by directly acting on skeletalmuscle RyRs in the presence of ethanol.