Showing papers in "Plant Physiology in 1984"

Biosynthesis of jasmonic Acid by several plant species.

Abstract: Six plant species metabolized (18)O-labeled 12-oxo-cis,cis-10,15-phytodienoic acid (12-oxo-PDA) to short chain cyclic fatty acids. The plant species were corn (Zea mays L.), eggplant (Solanum melongena L.), flax (Linum usitatissimum L.), oat (Avena sativa L.), sunflower (Helianthus annuus L.), and wheat (Triticum aestivum L.). Among the products was jasmonic acid, a natural plant constituent with growth-regulating properties. The pathway is the same as the one recently reported by us for jasmonic acid synthesis in Vicia faba L. pericarp. First, the ring double bond of 12-oxo-PDA is saturated; then beta-oxidation enzymes remove six carbons from the carboxyl side chain of the ring. Substrate specificity studies indicated that neither the stereochemistry of the side chain at carbon 13 of 12-oxo-PDA nor the presence of the double bond at carbon 15 was crucial for either enzyme step. The presence of enzymes which convert 12-oxo-PDA to jasmonic acid in several plant species indicates that this may be a general metabolic pathway in plants.

Site of action of chlorsulfuron: inhibition of valine and isoleucine biosynthesis in plants.

Abstract: The sulfonylurea herbicide chlorsulfuron blocks the biosynthesis of the amino acids valine and isoleucine in plants. Addition of these two amino acids to excised pea root (Pisum sativum L. var Alaska) cultures incubated in the presence of chlorsulfuron completely alleviates herbicide-induced growth inhibition. The site of action of chlorsulfuron is the enzyme acetolactate synthase which catalyzes the first step in the biosynthesis of valine and isoleucine. This enzyme is extremely sensitive to inhibition by chlorsulfuron having I(50) values ranging from 18 to 36 nanomolar. In addition, acetolactate synthase from a wide variety of tolerant and sensitive plants species is highly sensitive to inhibition by chlorsulfuron.

Imidazolinones: Potent Inhibitors of Acetohydroxyacid Synthase

Abstract: The imidazolinones, a new chemical class of herbicides, were shown to be uncompetitive inhibitors of acetohydroxyacid synthase from corn. This is the first common enzyme in the biosynthetic pathway for valine, leucine, and isoleucine. The Ki for the imidazolinones tested ranged from 2 to 12 micromolar. These results may explain the mechanism of action of these new herbicides.

Effects of Ultraviolet-B Irradiance on Soybean : V. The Dependence of Plant Sensitivity on the Photosynthetic Photon Flux Density during and after Leaf Expansion.

Abstract: Soybeans (Glycine max [L.] Merr. cv Essex) were grown in a green-house, and the first trifoliate leaf was either allowed to expand under a high photosynthetic photon flux density (PPFD) (1.4 millimoles per square meter per second) or a low PPFD (0.8 millimoles per square meter per second). After full leaf expansion, plants from each treatment were placed into a factorial design experiment with two levels of ultraviolet-B (UV-B) radiation (0 and 80 milliwatts per square meter biologically effective UV-B) and two levels of concomitant PPFD (0.8 and 1.4 millimoles per square meter per second) resulting in a total of eight treatments. Measurements of net photosynthesis and the associated diffusion conductances, ribulose-1,5-bisphosphate carboxylase activity, chlorophyll and flavonoid concentrations, and leaf anatomy were examined for all treatments. Leaves expanded in the high PPFD were unaffected by UV-B radiation while those expanded in the low PPFD were sensitive to UV-B-induced damage. Likewise, plants which were UV-B irradiated concomitantly with the high PPFD were resistant to UV-B damage, while plants irradiated under the low PPFD were sensitive. The results of this study indicate that both anatomical/morphological and physiological/biochemical factors contribute toward plant sensitivity to UV-B radiation.

Photosynthetic and stomatal responses of two mangrove species, Aegiceras corniculatum and Avicennia marina to long term salinity and humidity conditions.

Abstract: Gas exchange characteristics were studied in two mangrove species, Aegiceras corniculatum (L.) Blanco and Avicennia marina (Forstk.) Vierh. var australasica (Walp.) Moldenke, grown under a variety of salinity and humidity conditions. The assimilation rate was measured as a function of the intercellular CO2 concentration [A(ci) curve]. The photosynthetic capacity decreased with increase in salinity from 50 to 500 millimolar NaCl, as shown by decline in both the initial linear slope and the upper plateau of the A(ci) curve, with A. corniculatum being the more sensitive species. The decline in photosynthetic capacity was enhanced by increase in the leaf to air vapor pressure difference from 6 to 24 millibars, but this treatment caused a decrease in only the upper plateau of the A(ci) curve. Stomatal conductance was such that the intercellular CO2 concentration obtaining under normal atmospheric conditions occurred near the transition between the lower linear and upper plateau portions of the A(ci) curves. Thus, stomatal conductance and photosynthetic capacity together co-limited the assimilation rate, which declined with increasing salinity and decreasing humidity. The marginal water cost of carbon assimilation was similar in most treatments, despite variation in the water loss/carbon gain ratio.

Acquisition of Thermotolerance in Soybean Seedlings : Synthesis and Accumulation of Heat Shock Proteins and their Cellular Localization.

Abstract: When soybean Glycine max var Wayne seedlings are shifted from a normal growth temperature of 28°C up to 40°C (heat shock or HS), there is a dramatic change in protein synthesis. A new set of proteins known as heat shock proteins (HSPs) is produced and normal protein synthesis is greatly reduced. A brief 10-minute exposure to 45°C followed by incubation at 28°C also results in the synthesis of HSPs. Prolonged incubation (e.g. 1-2 hours) at 45°C results in greatly impaired protein synthesis and seedling death. However, a pretreatment at 40°C or a brief (10-minute) pulse treatment at 45°C followed by a 28°C incubation provide protection (thermal tolerance) to a subsequent exposure at 45°C. Maximum thermoprotection is achieved by a 2-hour 40°C pretreatment or after 2 hours at 28°C with a prior 10-minute 45°C exposure. Arsenite treatment (50 micromolar for 3 hours) also induces the synthesis of HSP-like proteins, and also provides thermoprotection to a 45°C HS; thus, there is a strong positive correlation between the accumulation of HSPs and the acquisition of thermal tolerance under a range of conditions. During 40°C HS, some HSPs become localized and stably associated with purified organelle fractions ( e.g. nuclei, mitochondria, and ribosomes) while others do not. A chase at 28°C results in the gradual loss over a 4-hour period of the HSPs from the organelle fractions, but the HSPs remain selectively localized during a 40°C chase period. If the seedlings are subjected to a second HS after a 28°C chase, the HSPs rapidly (complete within 15 minute) relocalize in the organelle fractions. The relative amount of the HSPs which relocalize during a second HS increases with higher temperatures from 40°C to 45°C. Proteins induced by arsenite treatment are not selectively localized with organelle fractions at 28°C but become organelle-associated during a subsequent HS at 40°C.

Peroxidase Release Induced by Ozone in Sedum album Leaves Involvement of Ca2

Abstract: The effect of ozone was studied on the peroxidase activity from various compartments of Sedum album leaves (epidermis, intercellular fluid, residual cell material, and total cell material). The greatest increase following a 2-hour ozone exposure (0.4 microliters O3 per liter) was observed in extracellular peroxidases. Most of the main bands of peroxidase activity separated by isoelectric focusing exhibited an increase upon exposure to ozone. Incubation experiments with isolated peeled or unpeeled leaves showed that leaves from ozone-treated plants release much more peroxidases in the medium than untreated leaves. The withdrawal of Ca2+ ions reduced the level of extracellular peroxidase activity either in whole plants or in incubation experiments. This reduction and the activation obtained after addition of Ca2+ resulted from a direct requirement of Ca2+ by the enzyme and from an effect of Ca2+ on peroxidase secretion. The ionophore A23187 promoted an increase of extracellular peroxidase activity only in untreated plants. The release of peroxidases by untreated and ozone-treated leaves is considerably lowered by metabolic inhibitors (3-(3,4-dichlorophenyl)-1,1-dimethylurea and sodium azide) and by puromycin.

Ethylene: Symptom, Not Signal for the Induction of Chitinase and β-1,3-Glucanase in Pea Pods by Pathogens and Elicitors

Abstract: Infection of immature pea pods with Fusarium solani f.sp. phaseoli (a non-pathogen of peas) or f.sp. pisi (a pea pathogen) resulted in induction of chitinase and beta-1,3-glucanase. Within 30 hours, activities of the two enzymes increased 9-fold and 4-fold, respectively. Chitinase and beta-1,3-glucanase were also induced by autoclaved spores of the two F. solani strains and by the known elicitors of phytoalexins in pea pods, cadmium ions, actinomycin D, and chitosan. Furthermore, exogenously applied ethylene caused an increase of chitinase and beta-1,3-glucanase in uninfected pods. Fungal infection or treatment with elicitors strongly increased ethylene production by immature pea pods. Infected or elicitor-treated pea pods were incubated with aminoethoxyvinylglycine, a specific inhibitor of ethylene biosynthesis. This lowered stress ethylene production to or below the level of uninfected controls; however, chitinase and beta-1,3-glucanase were still strongly induced. It is concluded that ethylene and fungal infection or elicitors are separate, independent signals for the induction of chitinase and beta-1,3-glucanase.

Suppression of Nodule Development of One Side of a Split-Root System of Soybeans Caused by Prior Inoculation of the Other Side

Abstract: In a split-root system of soybeans (Glycine max L. Merr), inoculation of one half-side suppressed subsequent development of nodules on the opposite side. At zero time, the first side of the split-root system of soybeans received Rhizobium japonicum strain USDA 138 as the primary inoculum. At selected time intervals, the second side was inoculated with the secondary inoculum, a mixture of R. japonicum strain USDA 138 and strain USDA 110. In a short-day season, nodulation by the secondary inoculum was inhibited 100% when inoculation was delayed 10 days. Nodulation on the second side was significantly suppressed when the secondary inoculum was delayed for only 96 hours. In a long-day season, nodule suppression on the second side was highly significant, but not always 100%. Nodule suppression on the second side was not related to the appearance of nodules or nitrogenase activity on the side of split-roots which were inoculated at zero time. When the experiments were done under different light intensities, nodule suppression was significantly more pronounced in the shaded treatments.

Fermentative Metabolism of Chlamydomonas reinhardtii: I. Analysis of Fermentative Products from Starch in Dark and Light

Abstract: The anaerobic starch breakdown into end-products in the green alga Chlamydomonas reinhardtii F-60 has been investigated in the dark and in the light. The effects of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and carbonyl cyanide- p -trifluoromethoxyphenyl hydrazone (FCCP) on the fermentation in the light have also been investigated. Anaerobic starch breakdown rate (13.1 ± 3.5 micromoles C per milligram chlorophyll per hour) is increased 2-fold by FCCP in the dark. Light (100 watts per square meter) decreases up to 4-fold the dark rate, an inhibition reversed by FCCP. Stimulation of starch breakdown by the proton ionophore FCCP points to a pH-controlled rate-limiting step in the dark, while inhibition by light, and its reversal by FCCP, indicates a control by energy charge in the light. In the dark, formate, acetate, and ethanol are formed in the ratios of 2.07:1.07:0.91, and account for roughly 100% of the C from the starch. H 2 production is 0.43 mole per mole glucose in the starch. Glycerol, d-lactate, and CO 2 have been detected in minor amounts. In the light, with DCMU and FCCP present, acetate is produced in a 1:1 ratio to formate, and H 2 evolution is 2.13 moles per mole glucose. When FCCP only is present, acetate production is lower, and CO 2 and H 2 evolution is 1.60 and 4.73 moles per mole glucose, respectively. When DCMU alone is present, CO 2 and H 2 photoevolution is higher than in the dark. Without DCMU, CO 2 and H 2 evolution is about 100% higher than in its presence. In both conditions, acetate is not formed. In all conditions in the light, ethanol is a minor product. Formate production is least affected by light. The stoichiometry in the dark indicates that starch is degraded via the glycolytic pathway, and pyruvate is broken down into acetyl-CoA and formate. Acetyl-CoA is further dissimilated into acetate and ethanol. In the light, acetate is produced only in the presence of FCCP and, when photophosphorylation is possible, it is used in unidentified reactions. Ethanol formation is inhibited by the light in all conditions.

Hydraulic Conductance as a Factor Limiting Leaf Expansion of Phosphorus-Deficient Cotton Plants

Abstract: Suboptimal levels of phosphorus (P) strongly inhibited leaf expansion in young cotton (Gossypium hirsutum L.) plants during the daytime, but had little effect at night. The effect of P was primarily on cell expansion. Compared to plants grown on high P, plants grown on low P had lower leaf water potentials and transpiration rates, and greater diurnal fluctuations in leaf water potential. Hydraulic conductances of excised root systems and of intact transpiring plants were determined from curves relating water flow rate per unit root length to the pressure differential across the roots. Both techniques showed that low P significantly decreased root hydraulic conductance. The effects of P nutrition on hydraulic conductance preceded effects on leaf area. Differences in total root length, shoot dry weight, and root dry weight all occurred well after the onset of differences in leaf expansion. The data strongly indicate that low P limits leaf expansion by decreasing the hydraulic conductance of the root system.

Effect of Salinity on Nodule Formation by Soybean

Abstract: A split-root growth system was employed to evaluate the effect of NaCl on nodule formation by soybean (Glycine max L. Merr. cv Davis). By applying the salinity stress and rhizobial inoculum to only one-half the root system, the effects of salinity on shoot growth were eliminated in the nodulation process. Rhizobium colonization of inoculated root surfaces was not affected by the salt treatments (0.0, 26.6, 53.2, and 79.9 millimolar NaCl). While shoot dry weight remained unaffected by the treatments, total shoot N declined from 1.26 grams N per pot at 0.0 millimolar NaCl to 0.44 grams N per pot at 79.9 millimolar NaCl. The concentration of N in the shoot decreased from 3.75% N (0.0 millimolar NaCl) to 1.26% N at 79.9 millimolar NaCl. The decrease in shoot N was attributed to a sharp reduction in nodule number and dry weight. Nodule number and weight were reduced by approximately 50% at 26.6 millimolar NaCl, and by more than 90% at 53.2 and 79.9 millimolar NaCl. Nodule development, as evidenced by the average weight of a nodule, was not as greatly affected by salt as was nodule number. Total nitrogenase activity (C2H2 reduction) decreased proportionally in relation to nodule number and dry weight. Specific nitrogenase activity, however, was less affected by salinity and was not depressed significantly until 79.9 millimolar NaCl. In a second experiment, isolates of Rhizobium japonicum from nodules formed at 79.9 millimolar NaCl did not increase nodulation of roots under salt stress compared to nodule isolates from normal media (0.0 millimolar NaCl). Salt was applied (53.2 millimolar NaCl) to half root systems at 0, 4, 12, and 96 hours from inoculation in a third experiment. By delaying the application of salt for 12 hours, an increase in nodule number, nodule weight, and shoot N was observed. Nodule formation in the 12- and 96-hour treatments was, however, lower than the control. The early steps in nodule initiation are, therefore, extremely sensitive to even low concentrations of NaCl. The sensitivity is not related to rhizobial survival and is probably due to the salt sensitivity of root infection sites.

Measurement of Subcellular Metabolite Levels in Leaves by Fractionation of Freeze-Stopped Material in Nonaqueous Media

Abstract: This paper describes a technique for measuring the in vivo metabolite levels in the chloroplast stroma, the cytosol, and the vacuole of spinach (Spinacia oleracea U.S.A. hybrid 424) leaves. Spinach leaves were freeze stopped and the frozen tissue was ground and lyophilized. The dry material was homogenized by sonication in a mixture of carbon tetrachloride and heptane, and fractionated by density gradient centrifugation. Measurements of the activity of marker enzymes in various subcellular compartments show the chloroplastic material mainly appearing in the lightest fractions and the cytosolic material in the middle of the gradient, whereas most of the vacuolar material is found in the heaviest fraction. Using the measured distributions of metabolites and of marker enzymes in each fraction of the gradient, the subcellular distribution of the metabolite can be calculated.As a first application, the new fractionation technique was used to investigate the subcellular contents of malate and sucrose in spinach leaves. The results show striking diurnal changes of sucrose and malate, with both substances primarily located in the vacuolar compartment. About three times more malate is present at the end of the day than at the end of the night. The sucrose content in the vacuole falls from a maximum of 45 millimolars at the end of the day to an almost undetectable value of approximately 1 millimolar at the end of the night.

Pea xyloglucan and cellulose : I. Macromolecular organization.

Abstract: A macromolecular complex composed of xyloglucan and cellulose was obtained from elongating regions of etiolated pea (Pisum sativum L. var. Alaska) stems. Xyloglucan could be solubilized by extraction of this complex with 24% KOH-0.1% NaBH(4) or by extended treatment with endo-1,4-beta-glucanase. The polysaccharide was homogeneous by ultracentrifugal analysis and gel filtration on Sepharose CL-6B, molecular weight 330,000. The structure of pea xyloglucan was examined by fragmentation analysis of enzymic hydrolysates, methylation analysis, and precipitation tests with fucose- or galactose-binding lectins. The polysaccharide was composed of equal amounts of two subunits, a nonasaccharide (glucose/xylose/galactose/fucose, 4:3:1:1) and a heptasaccharide (glucose/xylose, 4:3), which appeared to be distributed at random, but primarily in alternating sequence. The xyloglucan:cellulose complex was examined by light microscopy using iodine staining, by radioautography after labeling with [(3)H]fucose, by fluorescence microscopy using a fluorescein-lectin (fucose-binding) as probe, and by electron microscopy after shadowing. The techniques all demonstrated that the macromolecule was present in files of cell shapes, referred to here as cell-wall ;ghosts,' in which xyloglucan was localized both on and between the cellulose microfibrils. Since the average chain length of pea xyloglucan was many times the diameter of cellulose microfibrils, it could introduce cross-links by binding to adjacent fibrils and thereby contribute rigidity to the wall.

Rapid Accumulation of γ-Aminobutyric Acid and Alanine in Soybean Leaves in Response to an Abrupt Transfer to Lower Temperature, Darkness, or Mechanical Manipulation

Abstract: Soybean (Glycine max [L.] Merr) leaves contain a low level (0.05 micromole per gram fresh weight) of gamma-aminobutyric acid (Gaba) but the concentration of this non-protein amino acid increased to 1 to 2 micromoles per gram fresh weight within 5 minutes after transfer of plants or detached leaves from 33 degrees C to 22 degrees C or lower temperatures. A parallel decrease occurred in the concentration of glutamate. Accumulation of Gaba was also triggered by mechanical damage to the soybean leaves, but in plants subjected to a gradual reduction in temperature (2 degrees C per minute) only a small increase in Gaba occurred. A rapid increase in the concentration of alanine and decrease in glycine occurred upon transfer of the soybean plants to darkness and was not influenced by temperature. When plants were returned to normal growing conditions, all changes in amino acid concentrations were fully reversed in 1 hour.In soybean leaf discs incubated with [(14)C]glutamate, a rapid accumulation of [(14)C]Gaba was detected, and glutamate decarboxylase activity of the soybean leaf considerably exceeded (>30-fold) that of Gaba pyruvate transaminase. Part of the transaminase was localized in the mitochondria, but glutamate decarboxylase was not associated with any organelle or membrane component of the leaf cell. We consider that Gaba accumulation results from some change in intracellular compartmentation of the cell triggered by low temperature shock or mechanical damage. The accumulation of alanine due to a light-dark transition could be accounted for by transamination. [(14)C]Alanine formation was demonstrated when soybean leaf extracts were incubated with glutamate, aspartate, or serine and [(14)C]pyruvate.The changes in amino acid concentrations described for soybean leaves were demonstrated for all the vegetative tissues of the soybean plant and at variable rates in the leaves of a range of plant species. The response in detached tomato (Lycopersicon esculentum Mill.) leaves was of a similar magnitude to soybean. Thus, precautions are necessary to minimize changes in amino acid composition induced by manipulation and extraction of plant material.

Inhibition of 2,4-dichlorophenoxyacetic Acid-stimulated elongation of pea stem segments by a xyloglucan oligosaccharide.

Abstract: Xyloglucan, isolated from the soluble extracellular polysaccharides of suspension-cultured sycamore (Acer pseudoplatanus) cells, was digested with an endo-β-1,4-glucanase purified from the culture fluid of Trichoderma viride. A nonasaccharide-rich Bio-Gel P-2 fraction of this digest inhibited 2,4-dichlorophenoxyacetic-acid-stimulated elongation of etiolated pea stem segments. The inhibitory activity of this oligosaccharide fraction exhibited a well-defined concentration optimum between 10−2 and 10−1 micrograms per milliliter. Another fraction of the same xyloglucan digest, rich in a structurally related heptasaccharide, did not, at similar concentrations, significantly inhibit the elongation.

Thermolysin is a suitable protease for probing the surface of intact pea chloroplasts.

Abstract: Several proteases, i.e., pronase, a mixture of trypsin and chymotrypsin, and thermolysin were screened as potential surface probes of isolated intact pea (Pisum sativum var Laxton's Progress No. 9) chloroplasts. Of these, only thermolysin met the criteria of a suitable probe. Thermolysin destroyed outer envelope polypeptides, but did not affect inner envelope polypeptides, envelope permeability properties or such chloroplast activities as metabolite transport and O(2) evolution.

Analysis of the Complexity and Diversity of mRNAs from Pollen and Shoots of Tradescantia.

Abstract: The mRNAs of the mature pollen grain of Tradescantia paludosa at anesthesia and of vegetative shoots have been compared by analyzing the kinetics of hybridization between homologous and heterologous reactions of cDNA to poly(A)RNA in excess. The mRNAs in pollen can be divided into three abundance classes with complexities of 5.2 × 104, 1.6 × 106, and 2.1 × 107 nucleotides. The three classes are made up of sequences that constitute 15, 60, and 24% of the mRNAs and each sequence is present on an average at 26,000, 3,400, and 100 copies, respectively, per pollen grain. About 20,000 different genes are expressed in pollen as compared to about 30,000 in vegetative shoots. Estimates have been made of pollen mRNA sequences shared with those of shoot tissue and of shoot sequences common to those in pollen.

Physiological Responses to Salinity in Selected Lines of Wheat

Abstract: Two selections of bread wheat, Triticum aestivum L., differing in their relative salt resistance, were grown in salinized solution culture, and relative growth rates, osmotic adjustment, ion accumulation, and photosynthesis were monitored to study the responses of the plants to salinity. Differences in water relations were minimal and were only apparent for 3 days following salinization. The lines differed substantially in their relative growth rates and photosynthetic responses for several weeks following salinization, despite full osmotic adjustment. Concentrations of major cations and Cl − in the plant organs were remarkably similar in both lines, indicative of minimal differences in gross ion absorption and translocation. The authors interpret these results to suggest that the major difference between these two lines of wheat was their response to specific ion effects, at the level of the organ, tissue, cell, and subcellular entities. Superior compartmentation of toxic ions by the more salt-tolerant line, presumably in the vacuole, might have enabled it to maintain its cytoplasmic metabolic apparatus in a stabler and more nearly normal state than the sensitive line was able to do; a measure of true cytoplasmic toleration of salt may also be a factor.

Kok effect and the quantum yield of photosynthesis : light partially inhibits dark respiration.

Abstract: The linear response of photosynthesis to light at low photon flux densities is known to change abruptly in the vicinity of the light compensation point so that the quantum yield seems to decrease as radiation increases. We studied this ;Kok effect' in attached sunflower (Helianthus annuus L. cv IS894) leaves using gas exchange techniques. The effect was present even though respiration was constant in the dark. It was observed at a similar photon flux density (7 to 11 micromole photons per square meter per second absorbed photosynthetically active radiation) despite a wide range of light compensation points as well as rates of photosynthesis. The effect was not apparent when photorespiration was inhibited at low pO(2) (1 kilopascal), but this result was complicated because dark respiration was quite O(2)-sensitive and was partially suppressed under these conditions. The Kok effect was observed at saturating pCO(2) and, therefore, could not be explained by a change in photorespiration. Instead, the magnitude of the effect varied as dark respiration varied in a single leaf, and was minimized when dark respiration was minimized, indicating that a partial suppression of dark respiration by light is responsible. Quantum yields measured at photon flux densities between 0 and 7 to 11 micromole photons per square meter per second, therefore, represent the combined yields of photosynthesis and of the suppression of a component of dark respiration by light. This leads to an overestimate of the quantum yield of photosynthesis. In view of these results, quantum yields of photosynthesis must be measured (a) when respiration is constant in the dark, and (b) when dark respiration has been inhibited either at low pO(2) to eliminate most of the light-induced suppression of dark respiration or at photon flux densities above that required to saturate the light-induced suppression of dark respiration. Significant errors in quantum yields of photosynthesis can result in leaves exhibiting this respiratory behavior if these principles are not followed.

Host-Pathogen Interactions XXV. Endopolygalacturonic Acid Lyase from Erwinia carotovora Elicits Phytoalexin Accumulation by Releasing Plant Cell Wall Fragments

Abstract: Heat-labile elicitors of phytoalexin accumulation in soybeans (Glycine max L. Merr. cv Wayne) were detected in culture filtrates of Erwinia carotovora grown on a defined medium containing citrus pectin as the sole carbon source. The heat-labile elicitors were highly purified by cation-exchange chromatography on a CM-Sephadex (C-50) column, followed by agarose-affinity chromatography on a Bio-Gel A-0.5m gel filtration column. The heat-labile elicitor activity co-purified with two α-1,4-endopolygalacturonic acid lyases (EC 4·2·2·2). Endopolygalacturonic acid lyase activity appeared to be necessary for elicitor activity because heat-inactivated enzyme preparations did not elicit phytoalexins. The purified endopolygalacturonic acid lyases elicited pterocarpan phytoalexins at microbial-inhibitory concentrations in the soybean-cotyledon bioassay when applied at a concentration of 55 nanograms per milliliter (1 × 10−9 molar). One of these lyases released heat-stable elicitors from soybean cell walls, citrus pectin, and sodium polypectate. The heat-stable elicitor-active material solubilized from soybean cell walls by the lyase was composed of at least 90% (w/v) uronosyl residues. These results demonstrate that endopolygalacturonic acid lyase elicits phytoalexin accumulation by releasing fragments from pectic polysaccharides in plant cell walls.

Localization and capacity of proton pumps in roots of intact sunflower plants.

Abstract: Proton extrusion by roots of intact sunflower plants ( Helianthus annuus L.) was studied in nutrient solutions or in agar media with a pH indicator. Proton extrusion was enhanced by either iron deficiency, addition of fusicoccin, or single salt solutions of ammonium or potassium salts. The three types of proton extrusion differ in both localization along the roots and capacity. From their sensitivity to ATPase inhibitors it seems justified to characterize them as proton pumps driven by plasma membrane APTases. Enhanced proton extrusion induced by preferential cation uptake from (NH 4 ) 2 SO 4 or K 2 SO 4 was uniformly distributed over the whole root system. In contrast, the enhancement effect of fusicoccin was confined to the basal root zones and that of iron deficiency to the apical root zones. Also the rates of proton extrusion per unit of root fresh weight differed remarkably and increased in the order: Fusicoccin ≪ K 2 SO 4 4 ) 2 SO 4 Under iron deficiency the average values of proton extrusion for the whole root system are 5.6 micromoles H + per gram fresh weight per hour; however, for the apical root zones values of about 28 micromoles H + can be calculated. This high capacity is most probably related to the iron deficiency-induced formation of rhizodermal transfer cells in the apical root zones. It can be assumed that the various types of root-induced acidification of the rhizosphere are of considerable ecological importance for the plant-soil relationships in general and for mobilization of mineral nutrients from sparingly soluble sources in particular.

Proline and glycine betaine influence protein solvation.

Abstract: Glutamine synthetase from barley ( Hordeum distichum L.) is precipitated by polyethylene glycol (PEG). Proline, in a concentration-dependent manner, reduces the amount of enzyme precipitated by PEG, although the effect of the imino acid can be counteracted by raising the level of PEG. The effect of PEG is a function of mer number and concentration and the influence of both elements can be ameliorated by proline. PEG-induced enzyme precipitation is a function of pH, as is its interaction with both proline and betaine in the reaction. The lack of effect of amount of enzyme on the proline and PEG effects supports the conclusion that, in this system, proline and PEG do not function through interaction with the protein. Other compounds, such as glycine, glucose, and sucrose, can decrease the PEG-induced precipitation of the enzyme, although glycerol was not active under the conditions employed. The results are consistent with the proposition that a protein-containing system in which high concentrations of proline and/or betaine are present, is better `protected9 against the biologically unfavorable consequences of dehydration-induced thermodynamic perturbation.

Nickel in higher plants: further evidence for an essential role.

Abstract: Soybeans (Glycine max [L.] Merr.) grown in Ni-deficient nutrient solutions accumulated toxic urea concentrations which resulted in necrosis of their leaflet tips, a characteristic of Ni deficiency. Estimates of the Ni requirement of a plant were made by using seeds produced with different initial Ni contents. When compared to soybeans grown from seeds containing 2.5 nanograms Ni, plants grown from seeds containing 13 nanograms Ni had a significantly reduced incidence of leaflet tip necrosis. Plants grown from seeds containing 160 nanograms Ni produced leaves with almost no leaflet tip necrosis symptoms. Neither Al, Cd, Sn, nor V were able to substitute for Ni.In other experiments, a small excess of EDTA was included in the nutrient solution in addition to that needed to chelate micronutrient metals. Under these conditions, nodulated nitrogen-fixing soybeans had a high incidence of leaflet tip necrosis, even when 1 micromolar NiEDTA was supplied. However, in nutrient solutions containing inorganic sources of N, 1 micromolar NiEDTA almost completely prevented leaflet tip necrosis, although no significant increase in leaf urease activity was observed. Cowpeas (Vigna unguiculata [L.] Walp) grown in Ni-deficient nutrient solutions containing NO(3) and NH(4) also developed leaflet tip necrosis, which was analogous to that produced in soybeans, and 1 micromolar NiEDTA additions prevented these symptoms.These findings further support our contention that Ni is an essential element for higher plants.

Influence of Osmotic Adjustment on Leaf Rolling and Tissue Death in Rice (Oryza sativa L.)

Abstract: Osmotic adjustment, measured by the lowering of the osmotic potential at full turgor, and its influence on leaf rolling and leaf death was assessed in the lowland rice (Oryza sativa L.) cultivar IR36 in both the greenhouse and field. The degree of osmotic adjustment varied with the degree and duration of stress, but was usually 0.5 to 0.6 megapascal (maximally 0.8 to 0.9 megapascal) under severe stress conditions. In leaves in which osmotic adjustment was 0.5 to 0.6 megapascal, leaf rolling and leaf death occurred at lower leaf water potentials in adjusted than in nonadjusted leaves. We conclude that osmotic adjustment aids in the drought resistance of rice by delaying leaf rolling, thereby maintaining gas exchange, and by delaying leaf death.

Role of gibberellin in the growth response of submerged deep water rice.

Abstract: We have shown previously that ethylene, which accumulates in the air spaces of submerged stem sections of rice (Oryza sativa L. cv "Habiganj Aman II"), is involved in regulating the growth response caused by submergence. The role of gibberellins in the submergence response was studied using tetcyclacis (TCY), a new plant growth retardant, which inhibits gibberellin biosynthesis. Stem sections excised from plants that had been watered with a solution of 1 micromolar TCY for 7 to 10 days did not elongate when submerged in the same solution or when exposed to 1 microliter per liter ethylene in air. Gibberellic acid (GA(3)) at 0.3 micromolar overcame the effect of TCY and restored the rapid internodal elongation in submerged and ethylene-treated sections to the levels observed in control sections that had not been treated with TCY. The effect of 0.01 to 0.2 micromolar GA(3) on internodal elongation was enhanced two- to eight-fold when 1 microliter per liter ethylene was added to the air passing through the chamber in which the sections were incubated. GA(3) and ethylene caused a similar increase in cell division and cell elongation in rice internodes. Thus, ethylene may cause internodal elongation in rice by increasing the activity of endogenous GAs. In internodes from which the leaf sheath had been peeled off, growth in response to submergence, ethylene and GA(3) was severely inhibited by light.

H+-ATPase Activity from Storage Tissue of Beta vulgaris: I. Identification and Characterization of an Anion-Sensitive H+-ATPase

Abstract: Microsomal membranes isolated from red beet ( Beta vulgaris L.) storage tissue were found to contain high levels of ionophore-stimulated ATPase activity. The distribution of this ATPase activity on a continuous sucrose gradient showed a low density peak (1.09 grams per cubic centimeter) that was stimulated over 400% by gramicidin and coincided with a peak of NO 3 − -sensitive ATPase activity. At higher densities (1.16-1.18 grams per cubic centimeter) a shoulder of gramicidin-stimulated ATPase that coincided with a peak of vanadate-sensitive ATPase was apparent. A discontinuous sucrose gradient of 16/26/34/40% sucrose (w/w) was effective in routinely separating the NO 3 − -sensitive ATPase (16/26% interface) from the vanadate-sensitive ATPase (34/40% interface). Both membrane fractions were shown to catalyze ATP-dependent H + transport, with the transport process showing the same differential sensitivity to NO 3 − and vanadate as the ATPase activity. Characterization of the lower density ATPase (16/26% interface) indicated that it was highly stimulated by gramicidin, inhibited by KNO 3 , stimulated by anions (Cl − > Br − > acetate > HCO 3 − > SO 4 2− ), and largely insensitive to monovalent cations. These characteristics are very similar to those reported for tonoplast ATPase activity and a tonoplast origin for the low density membrane vesicles was supported by comparison with isolated red beet vacuoles. The membranes isolated from the vacuole preparation were found to possess an ATPase with characteristics identical to those of the low density membrane vesicles, and were shown to have a peak density of 1.09 grams per cubic centimeter. Furthermore, following osmotic lysis the vacuolar membranes apparently resealed and ATP-dependent H + transport could be demonstrated in these vacuole-derived membrane vesicles. This report, thus, strongly supports a tonoplast origin for the low density, anion-sensitive H + -ATPase and further indicates the presence of a higher density, vanadate-sensitive, H + -ATPase in the red beet microsomal membrane fraction, which is presumably of plasma membrane origin.

Cell Surfaces in Plant-Microorganism Interactions IV. Fungal Glycopeptides Which Elicit the Synthesis of Ethylene in Plants

Abstract: The production of ethylene by melon ( Cucumis melo cv Cantaloup charentais) tissues is stimulated during incubation in the presence of fungal glycopeptides extracted from Colletotrichum lagenarium , a pathogen of melon. These glycopeptides, called elicitors of ethylene, are found in the mycelium, the cell wall, and the culture filtrate. Elicitation of ethylene is a relatively early phenomenon and lasts for several hours. Upon purification of the crude elicitor extract by gel filtration and ion exchange chromatography, three elicitors were isolated. The three elicitors contained amino acid, sugar, and phosphate residues, and they have a decreased activity after partial chemical degradation of their sugar moiety. Elicitation of ethylene is not fungal species specific. Elicitors of phytoalexins, obtained from three Phytophtora species, enhanced ethylene biosynthesis in melon tissues.

Induction of Heat Shock Protein Messenger RNA in Maize Mesocotyls by Water Stress, Abscisic Acid, and Wounding

Abstract: Exposure of the excised growing region of the mesocotyl of young corn seedlings to heat shock stimulated the production of specific heat shock proteins and the intensification of synthesis of two proteins with a molecular weight of approximately 70,000. Water stress and abscisic acid also stimulated synthesis of these 70,000-dalton proteins, and other unique proteins distinct from those induced by heat shock. Growing tissues of intact corn mesocotyls exposed to heat shock, water stress, or abscisic acid accumulated mRNA species homologous to a cloned genomic probe of the 5' end of the 70,000-dalton Drosophila heat shock protein gene. Since cut segments of the mesocotyl under unstressed conditions produced a similar mRNA, we suggest that the hsp 70 gene is activated in corn by a variety of diverse stresses. Production of the mRNA is rapid, but transient, being induced within 3 hours of the imposition of the stress, but declining after reaching a maximum at 9 hours.

A Debranching Enzyme Deficiency in Endosperms of the Sugary-1 Mutants of Maize

Abstract: Many of the sugary-1 mutants of maize (Zea mays L.) have the highly branched water-soluble polysaccharide, phytoglycogen, in quantities equal to or greater than starch as an endosperm storage product in mature seeds. We find that all sugary mutants investigated are deficient in debranching enzyme [alpha-(1, 6)-glucosidase] activity in endosperm tissue 23 days postpollination and suggest that this deficiency is the primary biochemical lesion leading to phytoglycogen accumulation in sugary endosperms. This would indicate that the amylopectin component of starch depends on an equilibrium between the activities of branching enzymes introducing alpha-1,6 branch points into the linear alpha-1,4 glucans and debranching enzymes. The debranching enzyme activities from nonsugary endosperms can be separated into three peaks on a hydroxyapatite column. The sugary endosperm extracts lack one of these peaks of activity while the other two fractions have much reduced activity. The embryos of developing seeds (23 days after pollination) from both sugary and nonsugary genotypes have equivalent debranching activity. The debranching enzyme activity of developing endosperms is proportional to the number of copies (0 to 3) of the nonmutant (Su) allele present suggesting that the Su allele may be the structural gene for this debranching enzyme, although this is not definitive. This identification of debranching enzyme activity as being the biochemical lesion in sugary endosperms is consistent with several previous observations on the mutant.