Oxygen Consumption Stimulated by δ-Aminolevulinic Acid in Darkness and during Irradiation of Dark Grown Wheat Leaves

Dark grown wheat leaves (Triticum aestivum L. cv. Starke II Weibull), treated with δ-aminolevulinic acid in darkness, showed an increased oxygen uptake as measured by a Warburg method. The production of CO₂ was also increased in darkness, giving an RQ ? 1. The increased respiration was dependent on the treatment time as well as on the concentration of the δ-aminolevulinic acid. Potassium cyanide suppressed both the normal and the increased respiration. The treatment with δ-aminolevulinic acid caused accumulation of high amounts of protochlorophyllide. Levulinic acid suppressed the increased oxygen uptake as well as the protochlorophyllide accumulation in δ-aminolevulinic acid treated leaves. Irradiation rapidly decreased the protochlorophyllide content with a simultaneous increase in oxygen uptake over the dark value. The peak value of the increase in oxygen uptake was reached after about 5 min. The light induced oxygen uptake was dependent on the amount of PChlide present at the onset of irradiation. Also the CO₂ production was increased during the first minutes of irradiation but soon fell under the buffer control value. Neither potassium cyanide nor heat denaturation affected the oxygen uptake in light in contrast to the effect on the CO₂ production, which was blocked by heat denaturation. The increased oxygen uptake in light initially seems to be a purely photochemical process leading to a release of CO₂, which release is probably an enzymatic process induced by the photo-oxidative decomposition of pigment.     

Endogenous light-on rhythm in respiration of a long-day duckweed, Lemna gibba G3 II. On basic and rhythmic components of the rhythm

Effects of respiratory substrates (glucose, malate, citrateand pyruvate) and inhibitors (fluoride, iodoacetate, azide andDNP) on the O₂-uptake rhythm in a long-day duckweed,Lemna gibbaG3 in continuous light period were examined. Rates of O₂-uptake at the starting point (6 hr after the beginningof a continuous light period) and at the time of the first peakof the rhythm (18 hr after the beginning of a continuous lightperiod) were equally increased by exogenous substrates. Sensitivityof respiration to fluoride or iodoacetate was almost the sameat the 6th and 18th hr. The O₂-uptake (at the 6th, 18th, 30thand 42nd hr) was increased by DNP by the same amount. Azideat lower concentrations than 5X10^–4 M did not affect O₂-uptakeat the 6th hr, but inhibited uptake at the 18th hr. In the presenceof 5 X 10^–4 M of azide the rates of O₂-uptake at the 18th,30th or 42nd hr were down to the rate at the 6th hr, which wasinsensitive to azide. These results suggest that the O₂-uptakerhythm consists of two components, i.e. the basic respirationwhich is promoted by exogenous substrate, sensitive to DNP andinsensitive to azide; and rhythmic respiration, which is sensitiveto azide, but is not influenced by exogenous substrate and DNP. (Received February 19, 1971; )     

Stomatal Movement and Sucrose Uptake by Guard Cell Protoplasts of Commelina benghalensis L.

Sucrose concentration in guard cells of epidermal strips ofCommelina benghalensis increased with stomatal opening. Sucroseuptake patterns were investigated using guard cell protoplastsof C. benghalensis. Sucrose (0.5 mM) uptake into these protoplastswas sensitive to pH, with an optimum at pH 6. Uptake of sucroseinto guard cell protoplasts was inhibited by 2,4-dinitrophenol(DNP), diethylstilbestrol (DES) and (ptrifluoromethoxy)carbonylcyanide phenylhydrozone (FCCP), while DCMU and o-phenanthrolinehad no effect on the uptake of sucrose. Fusicoccin (FC) stimulatedsucrose influx. The influence of pH and the effect of the metabolicinhibitors on the sucrose uptake into the guard cell protoplastsare consistent with an energy dependent membrane-function. (Received July 7, 1986; Accepted September 26, 1986)     

Separation of phenylalanine transport events by using selective inhibitors, and identification of a specific uncoupler activity in Yersinia pestis.

Phenylalanine transport in Yersinia pestis TJW was differentially inhibited by sulfhydryl blocking reagents, uncoupling agents, and respiratory inhibitors. Kinetic studies with potassium cyanide and sodium azide showed that these compounds have no immediate effect on the initial rate of phenylalanine transport, but have an immediate and severe inhibitory effect on the rate of oxygen uptake. Identical studies with p-chloromercuribenzoate (pCMB) and 2,4-dinitrophenol (DNP) showed that these compounds have an instantaneous and total inhibitory effect on phenylalanine transport. DNP stimulated oxygen uptake, and pCMB caused only a sluggish inhibiton of oxygen uptake. pCMB acted as a competitive inhibitor of phenylalanine transport, whereas DNP inhibitied noncompetitively. Arrenius plots of the initial rate of phenylalanine transport in pCMB- and DNP-treated cells showed that DNP alters the transition temperature of the phenylalanine transport system from 17 C for control cells to 12 C. DNP did not inhibit transport when cells were treated at temperatures of 2 to 10 C. PCMB did not alter the normal transition temperature and inhibited phenylalanine transport over a 2 to 30 C temperature range. Efflux induced by both pCMB and DNP were blocked by placing cells at low temperatures (2 to 20 C). Inhibition of adenosine 5'-triphosphate synthesis by DNP did not show any temperature sensitivity as did phenylalanine transport. These data indicate that: (i) respiration is not obligatory for active transport of phenylalanine in Y. pestis TJW; and (ii) pCMB inhibits transport activity by reacting with the sulfhydryl group(s) at the carrier binding site. The data show that the uncoupler, DNP, selectively alters a temperature-dependent property of phenylalanine transport, that is not related to uncoupling activity of DNP , and probably involves membrane lipid alterations.     

FACTORS AFFECTING THE PRODUCTION OF CHROMOSOME ABERRATIONS BY CHEMICALS

In the present paper, the results of a study on the influence of temperature, hydrogen ion concentration, and oxygen tension on the production of chromosome aberrations in Vicia root tips by maleic hydrazide (MH), di-(2, 3-epoxypropyl)ether (DEPE), and 8-ethoxycaffeine (EOC), are described. Variations in the hydrogen ion concentration of the treatment solutions did not significantly influence the effect of EOC and DEPE. In contrast, the MH effect was considerably diminished by raising the pH from 4.7 to 7.3. A marked increase in the frequencies of aberrations produced by DEPE and MH was obtained by raising the temperature from 3° to 25°C. The effect of EOC increased with rising temperature up to 12°C. With a further rise in temperature the effect of EOC decreased, so that at 25°C. it was of about the same magnitude as at 3°C. The effect of EOC was completely inhibited, and that of MH partly so, when during the treatment (1) oxygen was excluded from the solution, (2) respiration was inhibited by azide or cyanide, or (3) phosphorylation was uncoupled from respiration by 2, 4-dinitrophenol (DNP). Pretreatments with DNP had a similar effect, but posttreatments did not influence the frequencies of aberrations. The effect of DEPE was unchanged by anoxia. Pre- or posttreatments with DNP did not change the total number of aberrations produced by DEPE, but the appearance of the effect was considerably delayed. The results are discussed.     

Tissue Respiration and Mitochondrial Oxidative Phosphorylation of NaCl-Treated Pea Seedlings

The effect of sodium chloride added to root medium of pea seedlings on respiratory activity of tissue segments and on isolated mitochondria was studied. Salinization enhances the respiration of leaves about one-third on a fresh weight, dry weight or protein basis. Roots and stems show only 10 to 15% respiratory stimulation. The onset of respiratory increase in leaves roughly parallels the increase in NaCl content and the decrease in growth rate. At a later stage the elevated respiration is apparent in treated plants even though the concentration of NaCl reaches a plateau and osmotic adjustment is being reached. Stimulation of respiration was found in both etiolated and green plants. Experiments with DNP show that simple uncoupling by salt is not involved; the respiratory increase in control and treated tissue is proportionally the same.

In accordance with increased respiration rates observed in vivo, mitochondria from salt-treated plants show higher rates of oxygen uptake on several substrates. The effect of NaCl added during growth is long term and is distinct from the effect of NaCl added to mitochondria isolated from control plants. Since P/O ratios are not affected by NaCl, the potential for oxidative phosphorylation in salt-affected tissue appears to increase. It is postulated that this increase may lead to changes in ADP and ATP content, and in turn, affect regulation of metabolic pathways.

     

Examination of the Possibility of the Involvement of the Alternative Respiration in Seed Germination

The possibility that the alternative respiration (AR) is involvedin seed germination was followed during and after imbibitionby noting the changes in the sensitivity to the respiratoryinhibitors, KCN and salicylhydroxamic acid (SHAM). This wascarried out by measuring oxygen uptake of mitochondrial preparationsfrom storage cotyledons and of the whole axis in a selectionof legumes (Vigna mungo, V. radiata, V. angularis, V. sinensisPhaseolus vulgaris, Pisum sativum and Dolichos lablab). Theextent of participation of AR in the respiration of cotyledonarymitochondria was variable according to species and time afterimbibition, and varied even within the same genus ( Vigna).In some species, AR was not observed. As to the axis, SHAM treatmentby itself had little effect on the rate of respiration, but,when applied with KCN, reduced oxygen uptake to an extent greaterthan KCN treatment alone. It was suggested that AR was not necessarilyimportant for seed germination. Key words: Alternative respiration, mitochondria, seed germination     

Effects of Deltamethrin on Field Grown Potato Plants:Biochemical and Ultrastructural Aspects

The effects of repeated spraying of field grown plants of Solanumtuberosum L. with deltamethrin 24, 41 and 55 d after emergence,on chlorophyll level, oxygen uptake, RuBisCo activity and leafultrastructure, were studied. Chlorophyll amount increased in treated plants after the secondand third spraying, whereas RuBisCo activity showed a highervalue than in the controls only after the last treatment. Incontrast, the respiratory activity was stimulated by the firstspraying and no significant differences were found with theensuing treatments. The general ultrastructure of mesophyllcells was not altered by deltamethrin, but morphometric analysisof electron micrographs showed that the fractional volume ofchloroplasts and starch in the treated leaves were consistentlylower than in controls. The biochemical and ultrastructural data suggested that thevegetative cycle of the sprayed plants had been extended.Copyright1993, 1999 Academic Press Solanum tuberosum L., potato plant, deltamethrin     

Rapid Shedding of Roots from Azolla filiculoides Plants in Response to Inhibitors of Respiration

Chemicals that are known to be inhibitors of respiration, namely,sodium azide, sodium cyanide, DNP, CCCP and DCCD, caused sheddingof roots of Azolla filiculoides plants. Complete shedding ofroots of more than 10 mm in length occurred when Azolla plantswere treated with sodium azide, DNP or CCCP at concentrationsabove 50µM, 30µM and 20µM, respectively. Theshedding in response to sodium azide, DNP and CCCP was veryrapid and was complete within 5–20 min. Microscopic studies revealed the presence of large cells atthe outer surface of the base of roots that were about to beshed. The tested-chemicals caused the expansion, rounding upand separation of these cells, probably via the rapid absorptionof water, with resultant shedding of roots. When detached roots were immersed in a solution of sodium azide,DNP or CCCP, the large cells expanded and rounded up. Thesecells were gradually separated from the roots. However, theseparation of the cells caused by DNP was inhibited by the presenceof various buffers at acidic pH. By contrast, buffers at neutralpH greatly facilitated the separation of cells irrespectiveof whether DNP was present or absent. The results suggest that the separation of cells involves anincrease in the pH of the external solution in the vicinityof the large cells. A change in ion fluxes of the large cells,which accompanies an increase in pH of the external solution,may cause the rapid absorption of water by the cells and resultin the expansion and separation of the cells. (Received June 15, 1993; Accepted October 25, 1993)     

EFFECT OF AQUEOUS FLUORIDE SOLUTIONS ON RESPIRATION OF INTACT BUSH BEAN SEEDLINGS

Applegate , Howard G., Donald F. Adams , and Roy C. Carriker . (Washington State U., Pullman.) Effect of aqueous fluoride solutions on respiration of intact bush bean seedlings. I. Inhibition and stimulation of oxygen uptake. Amer. Jour. Bot. 47(5): 339—345. Illus. 1960.–Intact bush bean seedlings were infiltrated with 1 × 10^–1, 1 × 10^–2, 1 × 10^–3, and 1 × 10^–4 M fluoride. A linear relationship was found to exist between the fluoride concentrations of the infiltrating solutions and tissue fluoride. The higher fluoride concentrations inhibited oxygen uptake, whereas lower fluoride concentrations accelerated oxygen uptake. Both the inhibition and acceleration of oxygen uptake by fluoride were modified by light, CO₂, and presence or absence of chlorophyll.     

Effect of Metabolic Inhibitors on Pinocytosis of Uranyl lons by Radish Root Cells: Probable Mechanisms of Pinocytosis

Pinocytotic uptake of uranyl ions by root cells of the radishRaphanus sativus L. cv. Red Giant and effects of monoiodacetate(MIA), 2,4-dinitrophenol (DNP), 2-mercaptoethanol (ME) and cytochalasinB (CB) on this process were studied. The number of invaginationsand pinocytotic vesicles in cells incubated with 0.15 mM uranylacctate (UA) is 15–20 times greater than in control cells,as estimated by counting the structures (plasmalemma derivatives)in serial sections. Monoiodacetate (0.05 mM) slightly stimulatedand ME (1.5 mM) completely inhibited pinocytosis. DNP (0.1 mM)inhibits UA pinocytosis by 45 per cent: DNP combined with MIAdiminishes pinocytotic activity by nearly 70 per cent. In thepresence of CB (4 µM) and UA quite large invaginations(exceeding 1 µm) have been observed. Cells treated withUA and UA $ MIA contain considerably more secretory vesicles.The results provide evidence for two types of pinocytosis inradish root cells; one independent of metabolic energy (I) andthe other essentially dependent on energy from respiration (II).Experiments with liposomes which show a pinocytosis-like behaviourin the presence of the pinocytosis inductors (including UA)indicate that type I is due solely to the effect exerted bythe inductor on the membrane. Type II is probably directly dependenton metabolism and may be regarded as a combination of uptakeand of sui generis removal of any excess plasmalemma increasedby exocytotic secretion. Raphanus sativus, pinocytosis, exocytosis, metabolic inhibitors, liposomes, electrostatic interaction, lipid peroxidation     

Participation of the cyanide-resistant pathway in respiration of winter rape leaves as affected by plant cold acclimation

Leaf slices sampled from winter rape plants ( Brassica napus L., var. oleifera L., cv. GórczaánAski), grown in cold (5°C), showed an increase in the dark respiration rate (measured at 25°C) as compared to slices cut from control plants (grown at 20/15°C). The effect of low temperature was most pronounced after 4 days of plant growth in the cold. Oxygen uptake by control slices was 60% inhibited by 1 m M KCN and was insensitive to 2.5 m M salicylhydroxamic acid (SHAM). On the contrary, respiration of leaf slices from cold-pretreated plants was more resistant to cyanide (35% inhibition after 4 days of cold treatment) and was 30% inhibited by SHAM. The patterns of cold-induced changes in total respiratory activity and in the estimated activity of alternative pathway were similar. It seems that in leaf slices from plants grown in the cold, the cyanide-resistant, alternative pathway participates in oxygen uptake. Cold treatment of plants also brought about a 4-fold increase in the level of soluble sugars, which reached a maximum on day 4 of exposure to cold. Addition of sucrose to the incubation medium resulted in an immediate increase in oxygen uptake by slices with low endogenous sugar level. The respiration stimulated by sucrose addition was more resistant to cyanide than the basal respiration and it was inhibited by SHAM. It is concluded that the operation of the alternative pathway is responsible for the increased oxygen uptake by the cold-grown winter rape leaves and it may be induced by an increased sugar supply for respiratory processes.     

Ethylene Production and Action during Foliage Senescence in Hedera helix L.

Detached leaves of Hedera helix remained green in the dark for20 d. Exogeneous ethylene increased respiration, endogeneousethylene biosynthesis and non-protein ninhydrin-positive compounds,while promoting losses of chlorophyll and sucrose. The sensitivityof the leaves to exogenous ethylene (1–100 mm³ dm^-3) variedgreatly with time of year. Ethylene treatment increased ADP,ATP, UDP and GTP appreciably but had no effect on other nucleotides.Leaves senescing naturally on the parent plant increased theirrates of ethylene evolution and oxygen uptake. These resultssupport the view that endogenous ethylene plays an integralpart in the senescence of detached and attached ivy leaves. Key words: Ethylene, leaves, senescence, nucleotides     

Influences of Light Quality, Growth Regulators and Inhibitors of Protein Synthesis on Respiration of Protoplasts from Meristematic Cells in Barley Seedlings

The influences of light of different wavelengths and plant growthregulators on the respiration of protoplasts isolated from tissue0 to 5 mm above the basal intercalary meristem of barley (Hordeumvulgare L. cv. Patty) leaves were studied. Respiration was measuredusing oxygen electrodes and a Cartesian-diver technique. Red,far-red and blue light all stimulated respiration in the protoplastsbut not in mitochondria isolated from them. Gibberellic acid stimulated respiration in protoplasts but abscisicacid had the opposite effect. Physiological concentrations ofindole-3-acetic acid and kinetin had no influence in eitherdirection. Combinations of gibberellic acid with light of anywavelength always increased respiration. Red or far-red light treatments in the presence of abscisicacid decreased dark respiration and only blue light significantlyreversed the inhibitory effect of abscisic acid. Cycloheximidemarkedly increased dark respiratory activity; chloramphenicolwas without effect. These results indicate that mitochondrialactivity in the leaf basal intercalary meristem was partiallycontrolled through phytochrome and a blue light receptor, andby gibberellic and abscisic acids. Changes in cytosolic proteinsynthesis were important for the initiation of enhanced mitochondrialactivity in meristems. Hordeum vulgare L., barley, abscisic acid, Cartesian-diver microrespirometry, gibberellic acid, meristematic respiration, protoplasts     

Effects of 2,4-dinitrophenol on membrane lipids of roots

Previous work has shown that the undissociated form of 2,4-dinitrophenol (DNP) increases the permeability of barley (Hordeum vulgare var. trebi) roots to ions. The present studies were undertaken to determine whether the effects of undissociated DNP were directly on membrane lipids. Relative amounts of the principal fatty acids from the lipids of barley root membranes were assayed as a function of DNP concentration, pH, and time of treatment under conditions similar to the previous studies of DNP effects on permeability. Undissociated DNP increases the proportions of palmitic and oleic acids and decreases linoleic and linolenic acids with no changes in the amounts of total fatty acids. The effects are immediate, as are the effects on permeability. Only the undissociated DNP is effective. Anionic DNP has no effect, although it is the major species taken up by the roots both at pH 5 and pH 7. DNP has no effect on respiration at either pH, indicating that undissociated DNP effects are on the membranes and not a general metabolic effect. The close parallelism between the effects of DNP on the composition of membrane lipids and on permeability suggests that the increase in permeability produced by undissociated DNP is due to a direct effect on the root membranes.     

The Action of Nitrophenols on Respiration and on Glucose Assimilation in Yeast

The consumption of 2·25 mg. glucose by a resting suspensionof baker's yeast in the presence of finely graded concentrationsof 3:5 -dinitro-o-cresol was followed at pH 2·4 and 5·0by measuring oxygen uptake and carbon-dioxide output; at pH8·5 oxygen uptake alone was measured. At each pH certainconcentrations of dinitrocresol stimulated respiration and increasedthe ratio of glucose oxidized to glucose assimilated; at higherconcentrations, assimilation was reduced to zero, but respirationbecame inhibited and the whole of the glucose was then metabolizedby aerobic fermentation. Dinitrocresol is believed to be actingin these experiments as an uncoupling agent, lowering the netrate of formation of energy-rich phosphate and so allowing respirationto proceed faster while assimilation, an endergonic process,is inhibited. The maximum respiratory stimulation obtainable at pH 8·5,325 per cent, of the control, is greater than can be obtainedat pH 2·6 or 5·0, which may be due to the slowerrespiration rate of controls at pH 8's. The slow endogenousrespiration can also be stimulated more than the exogenous.With o- and p-nitrophenols similar relations are obtained, butphenol does not stimulate respiration, although it inhibitsassimilation in lower concentrations than respiration. Undernitrogen, a stimulation of fermentation can be obtained withdinitrocresol at pH 5·0 but not at 2·6. At bothpH levels, assimilation is more easily suppressed than fermentationrate.     

Inhibition of respiration by sodium fluoride and monoiodoacetic acid in wheat leaves at different oxygen tensions and in the presence of methylene blue and L-cysteine

Six-day leaves of wheat (Triticum vulgare) were used to test the effect of oxygen, nitrogen and different mixtures of these gases, methylene blue and L-cysteine on the inhibition of respiration by sodium fluoride and monoiodoacetic acid using the Warburg manometric method. In an atmosphere of 100% oxygen total respiration was increased by an average of 45%, 10% oxygen led to a decrease in respiration by 20%. However, no simultaneous changes were found in the degree of inhibition by these enzyme poisons. On decreasing the oxygen content to 5% inhibition of respiration was almost doubled while at the same time respiration fell by an average of 23%. In nitrogen, the respiration rate varied around 10% and inhibition was almost 100%. The results showed that the action of enzyme poisons is only influenced by different oxygen content at low oxygen tension and is evidently associated with the dominance of a given energy system (glycolysis). On using methylene blue and L-cystein in concentrations of 10-3M a decrease in the relative inhibition of respiration by enzyme poisons was also found. The total respiration rate, however, was only raised by 6–8%. Since the change in the degree of inhibition on using these substances is caused by an increase in the participation of respiratory processes resistant to glycolytic inhibitors it can, in this case be assumed that there is an increase in the activity of the pentose cycle in such tissue.     

STUDIES ON RESPIRATION IN DEVELOPING POPPY SEEDS

The excised ovules of Papaver somniferum have been used to investigatethe endogenous uptake of oxygen during their maturation intoseeds. Effect of some activators and inhibitors and the activityof succinic dehydrogenase have been studied. There appears tobe an intimate correlation between respiration and the anatomicalchanges in the ovule. There is an increased oxygen uptake duringor immediately preceding the events of (a) pollination, (b)divisions of the endosperm nuclei, (c) cell wall formation inthe endosperm, and (d) elongation of cotyledons. The activityof succinic dehydrogenase follows a curve essentially similarto that of oxygen uptake. The low intensity of respiration duringcertain period of development appears not to be due to exhaustionof endogenous substrates. Endogenous substrates do, however,appear to be limiting on the days when maximum respiration occurssince added sugar does significantly enhance oxygen uptake.The results with two inhibitors, iodoacetate and fluoroacetate,suggest the operation of glycolysis and tricarboxylic acid cycleduring the earlier phase of seed development when the rate ofgrowth is highest. (Received September 1, 1964; )     

THE EFFECT OF INFECTION WITH TOBACCO MOSAIC VIRUS ON THE RESPIRATION OF TOBACCO LEAVES OF VARYING AGES IN THE PERIOD BETWEEN INOCULATION AND SYSTEMIC INFECTION

Leaves of tobacco plants inoculated with tobacco mosaic virus were divided into three groups: ( a ) inoculated leaves; ( b ) younger non-inoculated leaves present at the time of inoculation; ( c ) leaves formed since inoculation. The respiration rate of each group was compared with that of similar leaves from healthy plants. The respiration rate of inoculated leaves was increased by a constant amount for 3 weeks after inoculation, when it decreased. The respiration rate of group ( b ) leaves was not affected at any time, and that of group ( c ) leaves was decreased by 10% when they showed symptoms. The increased respiration in the inoculated leaves occurred too soon to reflect virus formation, and it is suggested that it reflects an initial change in infected cells preparatory to virus synthesis. The subsequent decrease in respiration may be due to the accumulation of virus which does not contribute to the total leaf respiration.     

Differences between Effects of Undissociated and Anionic 2,4-Dinitrophenol on Permeability of Barley Roots

Effects of 2,4-dinitrophenol (DNP) and several other substituted phenols on permeability of barley roots (Hordeum vulgare var. Trebi) to ions were assayed as a function of pH and phenol concentration. Solutions containing 0.1 micromolar undissociated DNP increase the permeability of barley root cells to small ions such as K⁺, Na⁺, Ca²⁺, and Cl⁻ with no inhibition of respiration. Undissociated forms of the other phenols increase permeability also, but they are less effective than DNP. Only the undissociated DNP is effective. Anionic DNP does not increase permeability or inhibit ion uptake, although it is the major species accumulated by the roots, both at pH 5 and pH 7. At pH 7, in contrast to pH 5, 10 micromolar DNP has no effect on ion permeability of barley roots yet it uncouples oxidative phosphorylation of barley root mitochondria. This indicates that the all too common use of DNP as a test for active transport or involvement of ATP synthesis can be misleading.