Biosynthesis and Metabolism of Indol-3yl-acetic Acid: I. THE NATIVE INDOLES OF BARLEY AND TOMATO SHOOTS

Barley and tomato shoots were examined quantitatively for naturally-occurringindole compounds. Both tissues were found to contain detectablelevels of indol-3yl-acetic acid (IAA), indol-3yl-aldehyde, tryptamine,5-hydroxytryptamine and malonyltryptophan. Tomato shoots alsocontained small amounts of indol-3yl-lactic acid and tryptophol.These two compounds were absent in barley, but this tissue containedsignificant quantities of 3 -aminomethylindole, 3-methylaminomethylindole,gramine, N-methyltryptamine and 5-hydroxy-N-methyltryptamine.The possible importance of these compounds in the biosynthesisof IAA, or in the formation of alkaloids, is discussed.     

Biosynthesis and Metabolism of Indol-3yl-acetic Acid: III. PARTIAL PURIFICATION AND PROPERTIES OF A TRYPTAMINE-FORMING L-TRYPTOPHAN DBCARBOXYLASE FROM TOMATO SHOOTS

A tryptamine-forming, L-tryptophan decarboxylase (E.C. 4.1.1.27 [EC] )from tomato shoots, has been partially purified and characterized.The properties of the enzyme were compared with those of tryptophantransaminase isolated from the same tissue, and separation ofthese two enzymes by ammonium sulphate fractionation clearlydemonstrated that tryptamine formation was due to the activityof the decarboxylase enzyme. Tryptophan decarboxylase was foundto be pyridoxal phosphate dependent and appeared to have substrateaffinities different from those of 5-hydroxytryptophan decarboxylase(E.C. 4.1.1.28 [EC] ) found in animal tissue. The importance of tryptophandecarboxylase in the biosynthesis of indol-3yl-acetic acid isdiscussed.     

Levels of Indol-3yl-acetic acid (IAA), IAA-oxidase and Peroxidase in Developing Cucumber Seedlings

The level of indol-3yl-acetic acid (IAA) in the cotyledons of cucumber seedlings increases in the period 4 to 11 days after germination. In hypocotyls and roots the IAA level decreases. IAA-oxidase activity of homogenates of cotyledons, hypocotyls and roots decreases with age. The soluble fraction of all three organs contains an IAA-oxidase, which may be allosteric. It is suggested that this IAA oxidase limits the upper level of IAA in the cell. The cell wall fraction of the three organs contains IAA-oxidases with conventional kinetics. The function of this IAA oxidase is probably to degrade exogenous IAA.     

The Response of Root Meristems to Colchicine and Indol-3yl-acetic acid in Vicia faba L.

The effects of colchicine and IAA treatments on mitotic activityin various root proliferating tissues have been determined.Lateral root primordia were not affected by IAA, though 24 hfollowing treatment mitotic activity was severely inhibitedin the apical meristems of 1-cm-long attached lateral rootsand primary roots. Primordia were also less sensitive to colchicinetreatment than root apical meristems. Thus telophase figureswere present in the former meristems 3 h following treatment,but not in the latter. Primordia and apical meristems respondedto the same extent, however, to the colchicine-induced increasein number of cells in metaphase, anaphase, and telophase, 3h after treatment began. The apparent difference between largeprimordia and root apical meristems in this respect was dueto the failure of colchicine to penetrate the cells of the formerproliferating tissues as rapidly as the latter. IAA was foundto prevent the increased MI found 24 h following colchicinetreatment only in those meristems where IAA inhibited mitoticactivity at this time. IAA treatments, either alone or withcolchicine, were also found to maintain mitotic activity in1-cm-long lateral roots which were excised from the primaryroots 24 h previously. In such laterals which were not treatedwith IAA, MI was zero at 24 h. It is concluded from the datareported in this paper that, during the development of rootapical meristems, changes take place in the response of cellsto factors affecting mitotic activity.     

A THEORY OF INJURY AND RECOVERY : II. EXPERIMENTS WITH MIXTURES.

1. The equations which serve to predict the injury of tissue in 0.52 M NaCl and in 0.278 M CaCl₂ and its subsequent recovery (when it is replaced in sea water) also enable us to predict the behavior of tissue in mixtures of these solutions, as well as its recovery in sea water after exposure to mixtures. 2. The reactions which are assumed in order to account for the behavior of the tissue proceed as if they were inhibited by a salt compound formed by the union of NaCl and CaCl₂ with some constituent of the protoplasm (certain of these reactions are accelerated by CaCl₂). 3. In this and preceding papers a quantitative theory is developed in order to explain: (a) the toxicity of NaCl and CaCl₂; (b) the antagonism between these substances; (c) the fact that recovery (in sea water) may be partial or complete, depending on the length of exposure to the toxic solution.     

Regulation of Ethylene Biosynthesis in Virus-Infected Tobacco Leaves : II. TIME COURSE OF LEVELS OF INTERMEDIATES AND IN VIVO CONVERSION RATES

Ethylene production was stimulated severalfold during the hypersensitive reaction of Samsun NN tobacco to tobacco mosaic virus (TMV). Exogenous methionine or S-adenosylmethionine (SAM) did not increase ethylene evolution from healthy or TMV-infected leaf discs, although both precursors were directly available for ethylene production. This indicates that ethylene production is not controlled at the level of methionine concentration or availability, nor at the level of SAM production or concentration. In contrast, 1-aminocyclopropane-1-carboxylic acid (ACC) stimulated ethylene production considerably. Thus, ethylene production is primarily limited at the level of ACC production.     

The Site of the Inhibition of the Shikimate Pathway by Glyphosate: II. INTERFERENCE OF GLYPHOSATE WITH CHORISMATE FORMATION IN VIVO AND IN VITRO

In the presence of the nonselective herbicide glyphosate (N-[phosphonomethyl]glycine), buckwheat (Fagopyrum esculentum Moench) hypocotyls and cultured cells of Galium mollugo L. accumulate an organic acid, which was identified as shikimate by mass-spectroscopy of its methyl ester. After growth in 0.5 millimolar glyphosate for 10 days, G. mollugo cells contained shikimate in amounts of up to 10% of their dry weight. Synthesis of chorismate-derived anthraquinones in G. mollugo was blocked by glyphosate. Chorismate and o-succinylbenzoate (an anthraquinone precursor) alleviated the inhibition. The conclusion drawn from these experiments, that glyphosate inhibits a step in the biosynthetic sequence from shikimate to chorismate, was substantiated by the finding that glyphosate is a powerful inhibitor of the conversion of shikimate to chorismate in cell-free extracts from Aerobacter aerogenes 62-1.     

VIRUS DISEASES OF CACAO IN WEST AFRICAL: II. CROSS-IMMUNITY EXPERIMENTS WITH VIRUSES 1A, 1B AND 1C

Experiments on cross-immunity reactions between three viruses attacking Theobroma cacao L. on the Gold Coast are described. A field trial involving 3 acres of graft-inoculated trees revealed some degree of protection afforded by Theobroma virus 1B against infection with virus 1A. The protection appeared to be more effective against insect inoculation than against graft transmission, being only temporary in the latter. Virus 1C (probably unrelated but not to be called Theobroma virus 2 until more evidence is available) conferred no protection against virus 1A.
The latent periods for these viruses were calculated from this experiment, which also provided data on their effects on yield. Virus 1A reduced yield by 50% in the first year after inoculation and killed the trees in the second. Virus 1B had no appreciable effect on yield; virus 1C reduced yield by 50 % in the third year after inoculation but there was no further decline in the fourth.
The rates of spread of these viruses were compared and significant differences demonstrated.     

Uptake, Translocation, and Metabolism of IAA in the Olive (Olea europea): I. UPTAKE AND TRANSLOCATION OF [1-14C] IAA IN DETACHED 'MANZANILLA' OLIVE LEAVES

Absorption, translocation, and decarboxylation of [1–¹⁴C]IAAby excised mature and young olive leaves were studied. The decarboxylationwas considerably more intense in mature leaves than in youngones, while the opposite was true for absorption. The rate ofdecarboxylation was dependent on the presence of peltate scalesof the leaves. The amount of non-biological decarboxylationand the possible effect of bacterial contamination on the systemwere studied and the rate of their involvement is discussed.     

The Translocation of Antibiotics in Higher Plants: II. THE MOVEMENT OF GRISEOFULVIN IN BROAD BEAN AND TOMATO

The uptake and translocation of griseofulvin from water cultureby broad bean and tomato has been studied; observations werealso made on its decay in broad bean. In most cases griseofulvinwas determined by bioassay. Where possible the bioassay waschecked by chemical estimations and was found to be adequate. The amount of griseofulvin taken up by the broad bean was proportionalto the volume of water transpired for any single concentrationof the treating solution and the decay was exponentially relatedto the time of exposure in the tissues. The accumulation ofgriseofulvin in the tissue had, therefore, an exponential componentbut within the limits of error there was constant relationshipbetween accumulation and transpiration over the time periodsused in the trials. The rate of accumulation in tomato was alsoconstant. The amount of griseofulvin accumulated by both beanand tomato after a definite time was a linear function of theconcentration of the treating solution. There were two processes involved in the uptake of griseofulvinby the broad bean: (a) an initial rapid entry into the rootswhich was inhibited by sodium azide and dinitrophenol at concentrationswhich did not reduce transpiration; and (b) a prolonged uptakelinearly related to transpiration which was not affected bythese concentrations of the inhibitors.     

Effects of Iron and Oxygen on Chlorophyll Biosynthesis : I. IN VIVO OBSERVATIONS ON IRON AND OXYGEN-DEFICIENT PLANTS

Corn (Zea mays, L.), bean (Phaseolus vulgaris L.), barley (Hordeum vulgare L.), spinach (Spinacia oleracea L.), and sugarbeet (Beta vulgaris L.) grown under iron deficiency, and Potamogeton pectinatus L, and Potamogeton nodosus Poir. grown under oxygen deficiency, contained less chlorophyll than the controls, but accumulated Mg-protoporphyrin IX and/or Mg-protoporphyrin IX monomethyl ester. No significant accumulation of these intermediates was detected in the controls or in the tissue of plants stressed by S, Mg, N deficiency, or by prolonged dark treatment. Treatment of normal plant tissue with δ-aminolevulinic acid in the dark resulted in the accumulation of protochlorophyllide. If this treatment was carried out under conditions of iron or oxygen deficiency, less protochlorophyllide was formed, but a significant amount of Mg-protoporphyrin IX and Mg-protoporphyrin IX monomethyl ester accumulated.     

Dormancy in Cereal Seeds: II. THE NATURE OF THE GASEOUS EXCHANGE IN IMBIBED BARLEY AND RICE SEEDS

When barley seeds imbibe water, the O₂ uptake of non-dormantseeds is considerably less than that of dormant seeds for atleast the first 6 h, irrespective of the rate at which the seedshad previously lost dormancy. During the initial 6 h of imbibition, the CO₂ output of dormantbarley seeds is usually only slightly greater than and sometimesno different from that of nondormant seeds. The CO₂ output ofdormant seeds is reduced by about 66 percent by millimolar KCN,whereas that of non-dormant seeds is decreased by about 12–13per cent only. The CO₂ output of dormant barley in nitrogenis considerably less than the CO₂ output of non-dormant seedsunder the same conditions. Dormant rice seeds also show a higher initial O₂ uptake thannon-dormant seeds, though this is not generally as marked asin barley. Similarly, the initial CO₂ output of dormant seedsis distinctly greater than that of non-dormant seeds, but inmillimolar KCN it is depressed to a greater extent than in non-dormantseeds. In nitrogen, the CO₂ outputs of dormant and non-dormantseeds were found to be the same. Consequently, unlike barley,dormant rice seeds appear to be as capable of carrying out alcoholicfermentation under anaerobic conditions as nondormant seeds. In barley, increasing the O₂ tension from 21 per cent to 100per cent increased the oxygen uptake of dormant seeds more thanthat of non-dormant seeds (an increase of 53 per cent as against20–23 Per cent). In dormant seeds there was a concomitantincrease in CO₂ output (about 50 per cent), but the CO₂ outputof non-dormant seeds was hardly affected. High concentrations of CO₂ are inhibitory to the germinationof both dormant and non-dormant barley seeds. At a concentrationof 10 per cent, however, CO₂ is inhibitory only to dormant seeds,although at 2.5–5 per cent it is sometimes stimulatoryto the germination of dormant seeds. A 24–h treatmentwith appropriate concentrations of ethanol, lactic acid, oracetaldehyde is also stimulatory to the germination of dormantbarley seeds. Histochemical investigations in barley indicated the presenceof peroxidase, cytochrome oxidase, and -glycero-phosphate dehydrogenasein the embryo, aleurone layer, and in a layer associated withthe testa. A number of other redox enzymes were detected inthe embryo and aleurone layer only. No differences in distributionor intensity of activity were detected between dormant and nondormantseeds.     

EFFECT OF CHROMIUM(VI) ON PHOTOSYSTEM II ACTIVITY AND HETEROGENEITY OF SYNECHOCYSTIS SP. (CYANOPHYTA): STUDIED WITH IN VIVO CHLOROPHYLL FLUORESCENCE TESTS1

The inhibitory effect of Cr(VI) on the PSII of Synechocystis sp. was studied. Cr(VI) reduced O₂ evolution and inhibited the water‐splitting system in PSII. S‐states test and flash induction test showed that Cr(VI) exposure increased the proportion of inactivated PSII (PSII_X) and PSII_β reaction centers, which increased the fluxes of dissipated energy. JIP test and Q_A^? reoxidation test demonstrated that Cr(VI) treatment induces inhibition of electron transport from Q_A^? to Q_B/Q_B^? and accumulation of P₆₈₀⁺. More Q_A^? had to be oxidized through S₂(Q_AQ_B)^? charge recombination and oxidation by PQ9 molecules in PSII under Cr(VI) stress. These changes finally decreased the index of photosynthesis performance.     

Uptake, Translocation, and Metabolism of IAA in the Olive (Olea europea): II. THE FATE OF EXOGENOUSLY APPLIED [2-14C]IAA TO DETACHED 'MANZANILLA' OLIVE LEAVES

The uptake, translocation, and metabolic pathway of IAA degradationin detached ‘Manzanilla’ olive leaves were studiedusing [2–¹⁴C] IAA. The uptake of radioactivity was veryrapid. With scraped leaves devoid of their peltate scales, itreached 80 percent of total applied radioactivity after 6 h.Translocation from the site of application in mature leaveswas negligible. Limited transport was apparent only in youngleaves. The absorbed label was initially ethanol-extractablebut later half of the radioactivity could not be extracted inethanol and had to be extracted with 1 N KOH. The first andmajor metabolite of the IAA in the olive leaves was identifiedas indolecarboxylic acid (ICA). Minor amounts of indoleacetylasparticacid (IAsp) and indolealdehyde (IAld) were also detected. Theidentity of the KOH-extractable label was not determined.     

Short-Term Nitrate Effects on Hydroponically-Grown Soybean cv. Bragg and its Supernodulating Mutant: II. DISTRIBUTION AND RESPIRATION OF RECENTLY-FIXED 13C-LABELLED PHOTOSYNTHATE

Fully symbiotic or nitrate treated (3 d, 4·0 mol m^–3)soybean (Glycine max [L.] Merr.) cv. Bragg and a nitrate tolerantsupernodulating soybean mutant nts 1007 were exposed to ¹³Cenriched CO₂ for a period of 10 h. During this period and forthe subsequent 24 h, continuous measurements of ¹³CO₂ and ¹²CO₂evolution of their root systems were undertaken. Three harvestsduring the experiment allowed determinations of the distributionof recently fixed carbon in different plant organs. These measurementsindicated higher dependence of N₂ fixation in nts 1007 on recentlyfixed carbon (RFC) by showing elevated RFC concentrations innodules as well as their augmented respiration. Root respirationof both genotypes was generally more reliant on stored carbon. Nitrate induced in all measured parameters a clear responsein the mutant analogous to the wild type, but quantitative differencesremained throughout. Nodule respiratory activity, the relativespecific activity (RSA), and the utilization of RFC were substantiallyreduced, but remained higher in nts 1007 than in Bragg, whilethe demand of roots for RFC increased in Bragg more than inthe supernodulator. The elevated carbon requirement of the nodulecomplement of the mutant and a high dependence on recently fixedcarbon could be attributed to higher nodule growth and maintenancecosts of the supernodulating genotype and were not associatedwith augmented nitrogen fixation activity. This less efficientutilization of carbon and the associated almost parasitic characterof the nodule complement of nts 1007 is considered to be thecause of reduced growth of the mutant. No evidence was foundfor a physiologically based nitrate tolerance in terms of nitrogenfixation. Key words: Glycine max, nitrate, N₂fixation, respiration, carbon partitioning, steady-state labelling     

THE INFLUENCE OF ELECTROLYTES ON THE CATAPHORETIC CHARGE OF COLLOIDAL PARTICLES AND THE STABILITY OF THEIR SUSPENSIONS : II. EXPERIMENTS WITH PARTICLES OF GELATIN,CASEIN, AND DENATURED EGG ALBUMIN.

1. This paper gives measurements of the influence of various electrolytes on the cataphoretic P.D. of particles of collodion coated with gelatin, of particles of casein, and of particles of boiled egg albumin in water at different pH. The influence of the same electrolyte was about the same in all three proteins. 2. It was found that the salts can be divided into two groups according to their effect on the P.D. at the isoelectric point. The salts of the first group including salts of the type of NaCl, CaCl₂, and Na₂SO₄ affect the P.D. of proteins at the isoelectric point but little; the second group includes salts with a trivalent or tetravalent ion such as LaCl₃ or Na₄Fe(CN)₆. These latter salts produce a high P.D. on the isoelectric particles, LaCl₃ making them positively and Na₄Fe(CN)₆ making them negatively charged. This difference in the action of the two groups of salts agrees with the observations on the effect of the same salts on the anomalous osmosis through collodion membranes coated with gelatin. 3. At pH 4.0 the three proteins have a positive cataphoretic charge which is increased by LaCl₃ but not by NaCl or CaCl₂, and which is reversed by Na₄Fe(CN)₆, the latter salt making the cataphoretic charge of the particles strongly negative. 4. At pH 5.8 the protein particles have a negative cataphoretic charge which is strongly increased by Na₄Fe(CN)₆ but practically not at all by Na₂SO₄ or NaCl, and which is reversed by LaCl₃. the latter salt making the cataphoretic charge of the particles strongly positive. 5. The fact that electrolytes affect the cataphoretic P.D. of protein particles in the same way, no matter whether the protein is denatured egg albumin or a genuine protein like gelatin, furnishes proof that the solutions of genuine proteins such as crystalline egg albumin or gelatin are not diaphasic systems, since we shall show in a subsequent paper that proteins insoluble in water, e.g. denatured egg albumin, are precipitated when the cataphoretic P.D. falls below a certain critical value, while water-soluble proteins, e.g. genuine crystalline egg albumin or gelatin, stay in solution even if the P.D. of the particles falls below the critical P.D.