Translocation of Organic Compounds in Sunflower IV. Effect of Temperature upon Translocation of Sugars

Apical halves of sunflower leaves were fed a solution of either¹⁴C-glucose or ^l4C-glucose and oligomycin at 5 and 25°C,respectively, and temperature, metabolism and translocationof ¹⁴C were measured at 10- to 30-min intervals. ¹⁴C-Labeledcompounds produced by control group leaves were compared tothose produced at 5°C and in the presence of oligomycin,at 25°C. In both experiments, similar small amounts of ¹⁴C-glucosemigrated into the basal half of the leaf. Sucrose translocationfrom the apical half was apparently affected by low temperatureand oligomycin. At least two types (affected and unaffectedby low temperature and oligomycin) of translocation seem tooccur simultaneously in sunflower. (Received July 4, 1980; Accepted November 27, 1980)     

Biosynthetic mechanism of glycolate in Chromatium II. Enzymic mechanism of glycolate formation by a transketolase system

The mechanism for the biosynthesis of glycolate in the photosyntheticbacterium Chromatium was studied. The enzymic nature of glycolatesynthesis from fructose-6-P and ribose-5-P by a cell-free bacterialextract, strictly dependent on the presence of ferricyanidein the assay mixture, was established. Removal of thiamine pyrophosphatefrom the assay mixture did not alter the magnitude of glycolateformation. By examining the reaction products from ¹⁴C-fructose-6-Pand ¹⁴C-ribose-5-P, the operating mechanism involved in glycolateformation was judged to be the oxidative breakdown of the glycolaldehyde-transketolaseaddition product by ferricyanide. The role of transketolasein the oxidative formation of glycolate in the photosynthesizingcells of Chromatium is discussed. ¹ This is paper XXXI in the series "Structure and Function ofChloroplast Proteins". Paper XXX is reference (5). The researchwas supported in part by grants from the Ministry of Educationof Japan (986009), the Toray Science Foundation (Tokyo), andthe Naito Science Foundation(Tokyo). (Received May 22, 1975; )     

Transport of shoot- and cotyledon-applied indole-3-acetic acid to Vicia faba root

To clarify the participation of indole-3-acetic acid (IAA) originatingfrom the shoot in root growth regulation and the mechanism ofIAA translocation from shoot to root, the movement of ¹⁴C-IAAwhich was applied to the epicotyl or the cotyledon of Viciafaba seedlings was investigated. The radioactivity of IAA appliedto the cotyledon moved faster to the root tip than that appliedto the epicotyl. On the basis of the effect of 2,3,5-triiodobenzoic acid on IAAmovement, a comparison with ¹⁴C-glucose movement and autoradiographicexamination, the nature of IAA movement was concluded to bepolar transport from the epicotyl to the basal part of the roots,while IAA movement from the epicotyl to the cotyledon, fromthe basal part of roots to the apical part, and from the cotyledonto the epicotyl and to the root took place in the phloem. Theradioactivity from ¹⁴C-IAA applied to the cotyledon accumulatedin lateral root primordia and vascular bundles. These factssuggest that IAA produced in cotyledons may participate in theregulation of Vicia root development. (Received December 21, 1979; )     

Ion Absorption and Allocation of Carbon Resources in Excised Pea Roots Grown in Liquid Medium in Absence or Presence of NaCl

Freshly excised pea roots (Pisum sativum cv. Alaska) when transferredto growth medium (130 mOsm) or growth medium containing salt( 370 mOsm) suffer an initial osmotic shock and lose water.Contol roots tended to accumulate potassium, particularly inthe apical zone, while those exposed to NaCl accumulated mainlysodium, potassium accumulation being depressed. Exposure tosalinity for 6 d caused increases in root protein, cellulose,uronic acid and lignin content per cell. In roots supplied with¹⁴C-glucose for 24 h immediately after excision there was littledifference in uptake of glucose and in its use in respirationbetween control and salt treated roots. However, there werenoticeable differences in incorporation of labelled carbon intoseveral cell fractions, and particularly into the cellulosefraction in the upper parts of the root. When roots were grownfor six days in culture before being supplied with [¹⁴C]glucose,uptake per root was greater in the 120 mM NaCl treatment, andthe fraction diverted to respiration was decreased by salinity.On a per cell basis incorporation into soluble starch, uronicacid and cellulose fractions was increased in the salt treatedroots. The data obtained are in accord with the previous findingsand are suggestive of increased synthesis of cell wall materials.No conclusion could be drawn as to whether the changes describedare of adaptive value. Pisum sativum L. cv. Alaska, root culture, salinity, osmoregulation, cell wall     

The Effect of Cycloheximide (Actidione) on Protein and Nucleic Acid Synthesis by Chlorella

Incorporation of ¹⁴C-phenylalanine, ¹⁴C-carbon dioxide, ¹⁴C-glucose,and ¹⁴C-glycine into the protein of Chlorella is inhibited bycycloheximide. A concentration of 2.5 µg per ml inhibitsincorporation by about 80 per cent; increasing the concentrationup to 10 µg per ml does not increase the degree of inhibition.The incorporation of ¹⁴C-adenine into ribonucleic acid (RNA)and deoxyribonucleic acid (DNA), and of ¹⁴C-glucose into polysaccharideis also inhibited. Unlike inhibition of protein synthesis, thatof nucleic acid and polysaccharide synthesis is observed onlyafter some delay. The delay is shortest for DNA synthesis andlongest for polysaccharide synthesis. Inhibition of ¹⁴C-glycineincorporation into DNA and RNA follows a similar pattern tothat obtained with ¹⁴C-adenine but the delay is much shorter.Cycloheximide also inhibits the formation of isocitrate lyasc(isocitrate-glyoxylate lyase, EC 4.1.3.1 [EC] ) when autotrophicallygrown cells are supplied with acetate.     

Glycine-serine transformation in the photosynthetic bacterium Chromatium vinosum

The metabolic transformation of glycine into serine in the photosyntheticbacterium Chromatium vinosum was accompanied by the evolutionof CO₂ due to decarboxylation of glycine. Isonicotinylhydrazideinhibited both ¹⁴CO₂ evolution and the formation of ¹⁴C-serinefrom ¹⁴C-glycine. The results indicate that a glycine-serinetransformation reaction takes place which is analogous to thatoccurring in green leaf tissues. Glycine may be metabolisedthrough serine by this reaction. The light stimulation of ¹⁴CO₂evolution and ¹⁴C-serine formation from 14C-glycine by the Chromatiumcells are judged to be results of the light-induced enhancementof ¹⁴C-glycine uptake by the bacterial cells. ¹This is paper 53 in the series "Structure and Function of ChloroplastProteins" and paper 7 of the series "Biosynthetic Mechanismof Glycolate in Chromatium". Paper 6 of the latter series isRef. 3 by Asami and Akazawa (1978). ²This study was aided by research grants from the Ministry ofEducation, Science and Culture of Japan and the Nissan ScienceFoundation (Tokyo). ³Postdoctoral Fellow (1980) of the Japan Society for the Promotionof Science. (Received May 20, 1980; )     

Suppression by 5-Bromo-2'-deoxyuridine of Transdifferentiation into Tracheary Elements of Isolated Mesophyll Cells of Zinnia elegans

5-Bromo-2'-deoxyuridine (BrdU), a thymidine analog, suppressedthe transdifferentiation into tracheary elements (TE) of isolatedmesophyll cells of Zinnia elegans without affecting cell division.Tracer experiments with [³H]BrdU indicated that 76% and 24%of the incorporated radioactivity was located in the DNA andthe RNA, respectively. Both thymidine and uridine counteractedthe inhibitory effect of BrdU on transdifferentiation but thymidinewas much more effective than uridine. These results suggestthat BrdU might interfere with transdifferentiation via itsincorporation into DNA. The timing of effective suppressionby BrdU was examined by monitoring the effects of the sequentialaddition of BrdU and thymidine with an interval of 12 h at varioustimes in culture. Transdifferentiation was most sensitive toBrdU between the 24th and the 36th hour of culture. This resultsuggests that this window of time is critical for DNA synthesisduring the transdifferentiation of isolated mesophyll cellsof Zinnia elegans into TE. ³Present address: Department of Chemical and Biological Sciences,Faculty of Science, Japan Women's University, Mejiro, Tokyo,112 Japan     

Pinitol occurrence in soybean plants as affected by temperature and plant growth regulators

Unsuitable temperatures are frequently encountered by soybean(Glycine max L. Merr.) plants grown in the field. Certain polyolshave been reported to protect plants from high temperature orfrost damage. Controlled environment studies were conductedto investigate the effect of stressful temperature regimes onthe content of pinitol (3-O-methyl-D-chiro-inositol) in soybeanplants. Hydroponically-grown soybean plants were subjected tohigh (35/30 C) or low (15/10 C) day/night temperature stresses,and pinitol content in different plant parts was determinedusing high performance liquid chromatography (HPLC). A syntheticplant growth regulator, PGR-IV, was foliarly applied to theplants to evaluate its effect on pinitol content in differentplant components. Uniformly-labelled ¹⁴C-glucose was fed intothe leaves via the transpiration stream, and the effects ofhigh temperature and EXP-S1089, another synthetic plant growthregulator, on the incorporation of ¹⁴C-glucose into pinitolwas evaluated using HPLC separation and scintillation spectrometry.High-temperature stress significantly increased plant pinitolcontent and the incorporation of ¹⁴C-glucose into pinitol, butdecreased the content of sucrose, glucose and fructose. Underlow-temperature stress, there was hardly any change in pinitolcontent, but a drastic increase in soluble sugars. PGR-IV enhancedpinitol translocation from leaves to stems and roots, whileEXP-S1089 increased pinitol/sucrose ratio. Accumulation of pinitolmay be an adjustment mechanism of the plant to reduce high-temperaturedamage, but not low-temperature injuries. Key words: Pinitol, soybean, temperature, plant growth regulator     

Biosynthesis of alicyclic acids from 14C-labeled glucose and erythrose by isolated cells from Vigna angularis leaves

Mesophyll cells isolated enzymatically from Vigna angularisleaves were fed ¹⁴Cglucose or ¹⁴C-erythrose and the time-courseof ¹⁴C incorporation into shikimic and quinic acids was examined.When ¹⁴C-glucose was fed to the cells, the highest radioactivityin quinic acid was observed after 10 hr of incubation, whilethat in shikimic acid was after 14 hr. In the experiment with¹⁴C-erythrose, the radioactivity in shikimic acid rose strikinglyup to the 3rd hour, but ¹⁴C in quinic acid increased graduallyduring the incubation. The incorporation of ¹⁴C into shikimicacid was enhanced when unlabeled shikimic or quinic acid wassupplied to the cells simultaneously with either ¹⁴C-glucoseor ¹⁴G-erythrose, whereas that into quinic acid was not significantlyincreased by these alicyclic acids. The difference in incorporationrate of ¹⁴C into quinic acid from that into shikimic acid wasmore conspicuous in the isolated mesophyll cells than in theepicotyls of V. angularis seedlings. ¹ Present address: Department of Biology, Faculty of Science,Kumamoto University, Kumamoto 860, Japan. (Received September 22, 1978; )     

Correlative Inhibition of Lateral Bud Growth in Phaseolus vulgaris L. Ethylene and the Physical Restriction of Apical Growth

Restriction of apical growth in Phaseolus by enclosing the upperpart of the shoot in sealed or ventilated tubes induced developmentof axillary buds beneath the enclosure. Enclosed parts of shootsshowed a reduction of leaf growth and, in experiments wherethe tubes were sealed, of internode extension. Enclosure ofthe shoots in large vessels that did not restrict leaf expansion,but which contained 0?5 vols 10^–6 ethylene, similarlyinduced axillary bud growth. Analysis of the gaseous extractof physically restricted shoots showed a 2?5-fold increase inethylene concentration. The results suggest involvement of ethylenein the release of correlative inhibition brought about by physicalrestriction of apical growth.     

Partitioning of 14C Labelled Assimilates in Arctium tomentosum

Plants of the biennial Arctium tomentosum were grown from seedto seed-set in an open field under three different treatments:control plants receiving full light intensity, plants with aleaf area reduced by 45 per cent, and shaded plants receivingonly 20 per cent of natural illumination. At various stagesof development the youngest fully expanded leaf of one plantin each treatment was exposed to ¹⁴CO₂ for half an hour. Subsequentdistribution of labelled assimilates in various plant partswas determined after eight hours. In the first year, the mostdominant sink was the tap root irrespective of variation inassimilate supply. During the production of new vegetative growthin the second season, a larger amount of radioactive photosynthatewas recovered from above ground parts, especially during formationof lateral branches. Seed filling consumed 80–90 per centof labelled carbon exported from the exposed leaf. In the secondyear, the most pronounced difference between treatments wasin the degree of apical dominance, being highest in shaded plantsand lowest in the plants with cut leaves. Results from ¹⁴C experimentsagreed fairly well with a ‘partitioning coefficient’derived from a growth analysis of plants grown independentlyunder the same experimental conditions. Reasons for discrepanciesbetween the ¹⁴C results and the partitioning coefficient arediscussed. Arctium tomentosum, burdock, variation in assimilate supply, assimilate distribution, ¹⁴CO₂, labelling, growth analysis     

Biosynthetic mechanism of glycolate in Chromatium III. Effects of {alpha}-hydroxy-2-pyridinemethanesulfonate, glycidate and cyanide on glycolate excretion

Effects of -hydroxy-2-pyridinemethanesulfonate (-HPMS), 2,3-epoxypropionate(glycidate), and cyanide on the photosynthetic activities ofChromatiumwere studied. -HPMS stimulated photosynthetic CO₂fixation in the bacterial cells in both N₂ and O₂ environments.The formation and subsequent excretion of both glycolate andglycine in the O₂ atmosphere were markedly enhanced by -HPMS.In contrast to a recent report by Zelitch [Arch. Biochem. Biophys.163: 367–377 (1974) ] that glycidate specifically inhibitsglycolate formation in tobacco leaf disks, we found that ithad no influence on CO₂ fixation by Chromatium in either N₂or O₂ atmosphere, and that the synthesis and extracellular excretionof glycolate were markedly stimulated by glycidate treatment.Cyanide (0.01–1 mM) exerted a marked inhibitory effecton photosynthetic CO₂ fixation in N². In O₂ atmosphere, photosynthesiswas stimulated by 0.01 mM cyanide, and inhibited by it abovethis level. Both the incorporation of ¹⁴CO₂ into glycolate andthe total synthesis of glycolate in the light were also enhancedby 0.01 mM cyanide, and strongly inhibited above that concentration. ¹This is paper XXXVI in the series "Structure and Function ofChloroplast Proteins," and the research supported in part bygrants from the Ministry of Education of Japan (No. 111912),the Toray Science Foundation (Tokyo) and the Naito Science Foundation(Tokyo). (Received May 31, 1976; )     

Synergistic Function of rolB, rolC, ORF13 and ORF14 of TL-DNA of Agrobacterium rhizogenes in Hairy Root Induction in Nicotiana tabacum

Comparison of the frequency of rooting in the tobacco leaf segmentsinoculated with Agrobacterium tumefaciens harboring variouscombinations of rolB, rolC, ORF13 and ORF14 of TL-DNA of Riplasmid (pRiHRI) revealed that the genes differ in their functionto stimulate adventitious root induction. A single gene rolBinduced roots, while rolC, ORF13 and ORF14 independently promotedthe root induction by the rolB gene. The effects of these geneson the rolB-mediated rooting were in the order of ORF13>rolCORF14. Present address: Laboratory of Phylogenetic Botany, Departmentof Biology, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba,263-8522 Japan. ² Present address: Department of Chemical and Biological Sciences,Faculty of Science, Japan Women's University, 2-8-1 Mejirodai,Bunkyo-ku, Tokyo, 112-8681 Japan.     

Biosynthetic mechanism of glycolate in Chromatium IV. Glycolate-glycine transformation

The metabolic transformation of glycolate to glycine occurringin photosynthesizing cells of Chromatium was investigated bythe radioisotopic technique and by amino acid analysis. By analyzingthe distribution of radiocarbon upon feeding [1-¹⁴C] glycolate,[2-¹⁴C] glyoxylate and [1-¹⁴C] glycine to bacterial cells, itwas demonstrated that glycolate is converted to glycinc viaglyoxylate, and both glycolate and glycine are excreted extracellularly.Although the formation of serine was barely detected by theabove two techniques in both N₂ and O₂ atmospheres, it was foundthat ¹⁴CO₂ is evolved quite markedly from both [1-¹⁴C] glycolateand [1-¹⁴C] glycine fed to the Chromatium cells. Analyticalresults of transient changes in amino acid compositions underatmospheric changes of N₂O₂ and by the addition of exogenousglycolate in N₂ confirm the notion that glycolate is convertedto glycine. Acidic amino acids (glutamic acid and aspartic acid)appear to take part in glycine formation as amino donors. Theformation of glycine from glycolate in a N₂ atmosphere suggeststhat an unknown glycolate dehydrogenation reaction may operatein the overall process. ¹ This is paper XXXVII in the series ‘Structure and Functionof Chloroplast Proteins’. Paper XXXVI is ref. (5). Theresearch was supported in part by grants from the Ministry ofEducation of Japan (No. 111912), the Toray Science Foundation(Tokyo) and the Naito Science Foundation (Tokyo). (Received July 14, 1976; )     

An Examination of the Value of 14C-urea as a Source of 14CO2 for Studies of Assimilate Distribution

Twenty-four hours after leaf 3 of a plant of Lolium multiflorumLam, was supplied with a droplet of ¹⁴C-urea and the plant enclosedin a polyethylene bag with an untreated plant, there were significantamounts of radiocarbon recovered from the untreated plant. Theleaf treated with ¹⁴C-urea was the major source of ¹⁴C leakagebut significant losses were also recorded from other parts ofthe plant. Reducing the humidity within the bag decreased theamount of ¹⁴CO₂ which escaped. Losses of radiocarbon from CO₂-treated plants were very low compared with those from urea-treatedplants but the pattern of assimilate distribution within thetwo types of plants was very similar. The possible causes ofthese effects are considered and the usefulness of ¹⁴C-ureaas a source of ¹⁴CO₂ discussed.     

Axillary Bud Formation in Two Isogenic Lines of Tomato Showing Different Degrees of Apical Dominance

Grafting experiments have been carried out in which rootstocksof the cultivar Craigella were paired with scions of an isogenicline Craigella Lateral Suppressor (ls ls) and vice versa, andthe levels of hormones in the roots and shoots of the graftedplants examined. The roots of Craigella plants differed from those of LateralSuppressor in that they contained a higher proportion of a cytokininthat co-chromatographed with N⁶ - (²—isopentenyl) adenosine.Reciprocal grafts did not lead to any qualitative or quantitativechanges in the cytokinins in the roots of either line. GraftingLateral Suppressor scions on Craigella rootstocks led to anincrease in the IAA content of the apical region and the ABAcontent of the stem tissue immediately below it, but when Craigellascions were grafted on Lateral Suppressor rootstocks there wereno changes in the level of either hormone. Cytokinins applied to the leaf axils of Lateral Suppressor plantsresulted in lateral bud initiation in the axils above the pointof treatment but not if the plants were also given a short periodof far-red light at the end of the photoperiod. Cytokinins wereineffective in initiating lateral buds in grafted Lateral Suppressorscions. It is suggested that root-produced cytokinins influence lateralbud outgrowth indirectly by way of their effect on the levelsof IAA and ABA in the shoot. Lycopersicon esculentum Mill., tomato, apical dominance, growth regulation, indol-3yl acetic acid, abscisic acid, cytokinins     

Purification and Properties of UDP-glucuronate Pyrophosphorylase from Pollen of Typha latifolia Linne

UDP-glucuronate pyrophosphorylase from the pollen of Typha latifoliaLinne was purified about 600-fold by protamine sulfate treatment,ammonium sulfate fractionation, gel filtration, chromatofocusing,affinity chromatography, and isoelectric focusing. The purificationwas carried out using buffer containing 20% sucrose which helpedto prevent enzyme inactivation. This enzyme required equimolarlevels of Mg²⁺ to PP_i or UTP for maximum velocity of enzymecatalysis. Results of experiments on product inhibition andthe initial velocity of the enzyme catalysis reaction suggesteda Theorell-Chance mechanism. ¹ Present address: Japan Spectroscopic Co., 2967-5, Ishikawa-Cho,Hachioji-City, Tokyo 192, Japan. (Received April 5, 1983; Accepted September 26, 1983)     

STUDIES ON THE RE-ASSIMILATION OF RESPIRATORY CO2 IN ILLUMINATED LEAVES

Experiments were performed, using rice, barley and Hydrangealeaves, to examine the re-assimilation of respiratory ¹⁴CO₂while photosynthesis is going on in an open air flow system. It was found that the leaves which had assimilated ¹⁴CO₂ beforehandevolved, when kept under photosynthesizing conditions, threeto four tenths (variable according to plant species and externalconditions) of the amount of ¹⁴CO₂ to be produced in the dark.Such an incomplete re-utilization of ¹⁴CO₂ was observed alsoin spinach leaf homogenate as well as in the leaves which hadpreviously absorbed ¹⁴C-glucose. The ¹⁴CO₂ output in rice leaves was found to be acceleratedby the light of high intensity. A possibility of light stimulationon the respiration was suggested. (Received October 7, 1961; )     

Further studies on the metabolism of glucose in the process of "glucose-bleaching" of Chlorella protothecoides

Previous studies have demonstrated that when cells of Chlorellaprotothecoides are incubated in a medium containing glucosebut no nitrogen source, they are profoundly bleached with degenerationof chloroplast structure and photosynthetic activity. When anitrogen source (urea) is added to the glucose medium, bleachingof algal cells is greatly suppressed. In this work the metabolismof glucose in the process of glucose-induced bleaching was studiedusing ¹⁴C-glucose as tracer. Changes in algal cell activityfor ¹⁴CO₂-evolution and ¹⁴C-incorporation into various cellularsubstances from ¹⁴C-glucose were followed. Most conspicuouswere increases in cellular activities for assimilating ¹⁴C-glucoseinto lipids (fatty acids) and glucose polymer. When urea wasadded to the glucose medium, the incorporation of ¹⁴C by algalcells into fatty acids was greatly reduced, while the assimilationof ¹⁴C into glucose polymer was increased. These and previous observations suggest that the formation oflarge amounts of lipids (fatty acids) probably is causally relatedto the induction of algal cell bleaching. (Received March 5, 1969; )     

The Translocation of 14C-Photoassimilate from Normal and Mutant Leaves to the Pods of Pisum sativum L.

In experiments using radioactive carbon dioxide (¹⁴CO₂) a comparisonwas made of the ¹⁴C-photoassimilate translocation potentialsof two normal leaved (genotype AfAfTlTl) and two mutant formsof Pisum sativum (pea). A ¹⁴CO₂ administration method is describedthat permitted ¹⁴C-translocation studies to be conducted underfield conditions. One of the mutants available produced tendrils in place of leaves(afafTlTl). The other mutant studied was without tendrils buthad a much branched petiole with numerous relatively minuteleaflets (afaftltl). These mutants and the normal-leaved cultivarswith which they were compared were not isogenic lines. Lengthybackcrossing would be required before full assessment couldbe made of the possible agronomic value of such mutations. An interim evaluation of these mutants was based on ¹⁴C-distributionassays that were conducted 48 h after feeding ¹⁴CO₂, to specifiedleaves. The indication was that in translocation terms the leafand pod had a well defined respective source and sink relationshipthat was independent of leaf morphology. In each case the podswhich constituted the major ¹⁴C sinks depended on which leafhad been fed ¹⁴CO₂. With regard to sink specific activity asdefined by the quantity of ¹⁴C incorporated per unit dry weightof pod, the mutants were not significantly different from normal. The implication of these findings was that fundamental changesin pea leaf morphology could be made genetically without a markedeffect on the photoassimilate export potential of the leaf.