Tangential Movement of 14C-labelled Assimilates in Stems of Willow

It was shown that, when ¹⁴CO₂, was fed to a single leafy shootof a rooted willow cutting, activity appeared within 12 hrs.in honeydew collected from a colony of the aphid Tuberolachnussalignus (Gmelin), feeding on the opposite side of the stem20 cm. below the base of the leafy shoot. This was shown tobe due to a tangential movement of labelled materials throughthe bark. This tangential movement was found only in cuttings with limitedroot growth. Where an active root sink was present, extensivetangential movement was absent. Using two aphid colonies on either side of the stem, determinationsof the speed of tangential movement have been made. The highestvalue obtained was 10.6 mm./hr.     

Studies on Translocation of Metabolites in the Xylem of Grapevine Shoots

Fluid extracted from grapevine shoots by suction contained upto 0.2 per cent (w/v) of sugar. The total sugar concentrationand the relative concentrations of sucrose, glucose, and fructosechanged along the length of a shoot. Whereas sucrose is absentfrom or present only in traces in fluid bleeding from cut stumps,it was found to be a major constituent in fluid extracted fromsections of wood by suction. ¹⁴C-glucose administered to thewood of an excised shoot moved up in the transpiration stream,moved laterally, into the bark, and was partly converted tosucrose and fructose. When ¹⁴CO₂ was supplied to a leaf of anintact vine, radioactive sugars were found in the fluid afterwardsextracted from the wood by suction. When a ring of bark wasremoved from the stem at a point above the ¹⁴CO₂-treated leafmost of the ¹⁴C in the bark, wood, and extracted fluid belowthe ring was in the form of sugar but ¹⁴C in tissues above thering was mainly in organic acids and amino acids. It is suggestedthat a barrier within the wood prevents diffusion of sugarsinto the transpiration stream, and that the fluid extractedfrom the wood by suction cannot be regarded as ascending sapsince it contains considerable amounts of non-moving material.     

The Movement of Sugars into the Sieve Elements of Bark Strips of Willow: I. METABOLISM DURING TRANSPORT

¹⁴C-labelled sugars (sucrose, glucose, and fructose) were suppliedto the cambial surface of bark strips of willow, and in conjunctionwith the aphid stylet technique (Weatherley, Peel, and Hill,1959), the movement into and distribution of activity in thesieve tube sap was examined. As well as sucrose, free hexosesand sugar phosphates were found to contain the ¹⁴C-label, andrapid interconversions occurred between these compounds somewherealong the entry route. Sucrose entry into the sieve elementswas accompanied by at least a partial breakdown into its componenthexoses, and the involvement of a sugar phosphate pool was alsodemonstrated. Possible transformation sites on the entry routebetween the solution bathing the cambial surface and the sieveelement vacuole are discussed.     

Effect of Temperature on Starch Degradation in Chlorella vulgaris 11h Cells

Analysis of products formed in Chlorella vulgaris 11 h cellsduring photosynthesis in air containing 3,000 ppm ¹⁴CO₂ at varioustemperatures revealed that the level of ¹⁴C-starch was maximumaround 20–24?C and decreased with further rise in temperatureuntil 40?C, while ¹⁴C-sucrose greatly increased at temperaturesabove about 28?C. Elevating the temperature from 20 to 38?Cduring photosynthetic ¹⁴CO₂ fixation resulted in a remarkabledecrease in ¹⁴C in starch and a concomitant increase in ¹⁴Cin sucrose. This conversion of starch to sucrose when shiftingthe temperature from 20 to 38?C proceeded even in the dark.Hydrolysis of sucrose by rß-fructosidase showed that,irrespective of the experimental conditions, the radioactivitiesin sucrose were equally distributed between glucose and fructose.The enhancement of starch degradation with temperature risewas more remarkable than that of the activity of ribulose bisphosphatecarboxylase from the same cells. When Chlorella cells whichhad been preloaded with ¹⁴C-starch after photosynthesis for30 min at 20?C were incubated in the dark for an additional30 min at 20?C, ¹⁴C-starch was degraded by only about 4%. However,the values after 30-min dark incubation at 28, 32, 36 and 40?Cwere increased by about 10, 19, 36 and 50%, respectively. Duringthe temperature-dependent conversion of starch to sucrose, nosignificant amount of radioactivity accumulated in free glucoseand maltose. (Received October 27, 1981; Accepted January 9, 1982)     

Photosynthetic Assimilation of 14CO2 by Mature Brown Stems of the Tea Plant (Camellia sinensis L.)

The fixation of ¹⁴CO₂ by mature brown stems of the tea plantwas studied by supplying ¹⁴CO₂ to selected stems of pruned andintact plants for 24 h under field conditions. Utilization of¹⁴C assimilates for the production of new shoots was also examined.The photosynthetic nature of the fixation of ¹⁴CO₂ is demonstrated.The efficiency of this fixation was very low compared with thattaking place in leaves. The movement of labelled assimilatesfrom stem bark to the roots was inappreciable, whereas newlyemerged shoots on the pruned frame drew labelled assimilatesfrom stem bark.     

The Effects of Leaf Age and Senescence on the Distribution of Carbon in Lolium temulentum

Assimilate distribution in leaves of Lolium temulentum was establishedby root absorption of [¹⁴C]sucrose and after exposure to ¹⁴CO₂.Age determined the amount of carbon assimilated, with more labelbeing incorporated during expansion than at maturity. Duringsenescence ¹⁴C assimilation was much lower. Ethanol-solubleextracts from various tissues of root-labelled plants containedmost of the radioactivity chiefly in basic and acidic compounds.The neutral fraction was composed predominantly of sucrose. Sucrose was comparably labelled in leaves from plants fed equalamounts of either [¹⁴C]sucrose, glucose, or fructose and onlytraces of labelled monosaccharides appeared in extracts. Radioactive sucrose was translocated rapidly from mature leaveswhereas, in the expanding leaf, carbon incorporation was directedtowards growth and the greater proportion of label present atligule formation was in ethanol-insoluble material. Induced senescence, of a mature leaf fed during expansion, produceda rapid loss from the pool of insoluble ¹⁴C. This was accompaniedby a reduction in the contents of chlorophyll and soluble proteinand an accumulation of amino acids. The onset of senescencecaused changes in leaf sugar levels which were correlated withincreased rates of respiration.     

Metabolism of Radioactive Sugars by Tobacco leaf Disks

Destarched tobacco-leaf disks were floated on per cent. (w/v)solutions of sucrose uniformly labelled with ¹⁴C in either theglucose or fructose moiety, and on invert sugar in which onehexose only was so labelled. The experiments were carried outin an atmosphere of oxygen at 25° C. Seventy-five per cent,of the sugar lost from the external solutions was recoveredas starch, sucrose, fructose, glucose, and CO₂. With sucroseas the substrate, 30 per cent, of the material was recoveredas CO₂ and 17 per cent. each as starch, sucrose, fructose, andglucose. With invert sugar as the substrate, 30 per cent, wasagain recovered as CO₂ only 20 per cent. as the three sugarstogether, and 50 per cent. as starch. Whichever hexose was initiallylabelled and whether the sugar was supplied as sucrose or hexose,the relative specific activities of starch and sucrose in theleaf disks and of the CO₂ evolved were equal or nearly equalto that of the sugar supplied. With sucrose as the substratethe sucrose in the disks retained its asymmetry of label, andfree hexoses produced were similarly asymmetrically labelled.When invert sugar was the substrate the sucrose synthesizedwas strongly labelled in both moieties, as also were the freehexosea. It is concluded that fructose and glucose free or combinedin sucrose were equally available for starch synthesis and CO₂,formation, and that there can be no question of preferentialutilization of one or other hexose. Starch and CO₂ must arisefrom a common source in which readily formed derivatives ofthe hexoses are rapidly equilibrated. Free hexose cannot participatedirectly in either sucrose or starch synthesis. Accumulationof sugar not immediately metabolized and inversion of sucrosetake place at a site remote from the common pool. A scheme toaccommodate the results is discussed.     

Investigation of Bidirectional Movement of Tracers in Sieve Tubes of Salix Viminalis L.

Sieve-tube sap was collected as honeydew from the aphid Tuberolachnussalignus (Gmelin) feeding on a young shoot or a mature stemof willow (Salix viminalis L.). Two radioactive tracers (either³H-glucose and ¹⁴C-sucrose, or ¹⁴CO₂, Na₂ ¹⁴CO₃, and ³²P-orthophosphate)were applied to leaves or to bark abrasions at each end of thestem. In some experiments the tracers were applied after thestart of honeydew production, whilst in others the tracers wereapplied before the aphids were sited. In most cases aphids feeding between the tracer applicationpoints produced doubly-labelled honeydew during a 24–48-hperiod from tracer application. In some instances the firstdrops of honeydew contained one tracer, followed several hourslater by the other tracer, whilst some aphids produced doubly-labelledhoneydew from the first drop. Singly-labelled or inactive honeydewwas found in only a few cases. It was demonstrated that when ³H-glucose and ¹⁴C-sucrose wereapplied at opposite ends of a stem that the tracers moved inopposite directions in the form of sucrose. The data suggest that a simultaneous bidirectional movementof two tracers may occur in the same sieve tube. However, thepossibility that the doubly-labelled honeydew could be producedby lateral movement from one sieve tube carrying one tracerto an adjacent punctured sieve tube carrying the other tracercannot be discounted.     

Glucose, pyruvate, and acetate metabolism by developing soybean seeds

Developing soybean cotyledons were incubated with glucose-¹⁴C,pyruvate-¹⁴C, and acetate-¹⁴C. Glucose was metabolized by boththe Embden-Meyerhof-Parnas pathway and the pentose phosphatepathway. Developing soybean cotyledons also have the capacityto synthesize sucrose since ¹⁴C was found in fructose and sucrosefrom glucose incubations. Complete analysis showed that thecarbons from glucose were directed into CO₂, lipid, and solids.Pyruvate was metabolized to a C-2 unit which is presumably acetylCoA. After conversion to the C-2 unit, the carbons of pyruvatewere metabolized in the same manner as acetate. Both pyruvateand acetate carbons were directed predominately into lipids. (Received January 6, 1976; )     

Preparation of Radioactive Starch, Glucose and Fructose from C14O2

Radioactive starch, glucose and fructose have been preparedfrom tobacco leaves after assimilation of C¹⁴O₂. The apparatusused for photosynthesis consisted of a shallow Perspex leafchamber connected to a closed gas system, in which C¹⁴O₂ wasgenerated from BaC¹⁴O₂. Six leaves, area 14 to 18 sq. dm. whenexposed to bright sunlight with an initial CO₂ concentrationof 8 to 10 per cent., assimilated 3.35 g. of C¹⁴O₂ in 8 to 10hours. At least 80 per cent. of the C¹⁴O₂ supplied appearedin the leaves as starch and sugar and over 80 per cent. of theradioactivity was accounted for in these carbohydrates. Thespecific activity per m. atom of carbon of the isolated productswas 85 to 90 per cent. of that of the C¹⁴O₂. Small amounts ofradioactive carbon were also incorporated in the leaf proteinand in the celluose, hemicellulose and polyuronides.     

Effects of Carbon Dioxide on Metabolism by the Glycollate Pathway in Leaves

When solutions of [¹⁴C]glycollate, glycine, serine, glycerate,or glucose were supplied to segments of wheat leaves throughtheir cut bases in the light, most of the ¹⁴C was incorporatedinto sucrose in air but in CO₂-free air less sucrose was made.The synthesis of sucrose was decreased because metabolism ofserine was partly blocked. Sucrose synthesis from glucose andglycerate in CO₂-free air was decreased but to a smaller extent;relatively more CO₂ was evolved and serine accumulated. Theeffects of DCMU and light of different wavelengths on metabolismby leaves of L-[U-¹⁴C]serine confirmed that simultaneous photosyntheticassimilation of carbon was necessary for the conversion of serineto sucrose. Of various products of photosynthesis fed exogenouslyto the leaves -keto acids were the most effective in promotingphotosynthesis of sucrose and release of ¹⁴CO₂ from ¹⁴C-labelledserine. This suggests that in CO₂-free air the metabolism ofserine may be limited by a shortage of -keto acid acceptorsfor the amino group. In CO₂-free air added glucose stimulatedproduction of CO₂ and sucrose from D-[U-¹⁴C]- glycerate andno competitive effects were evident even though glucose is convertedrapidly to sucrose under these conditions. In addition to asupply of keto acid, photosynthesis may also provide substratesthat can be degraded and provide energy in the cytoplasm forthe conversion of glycerate to sugar and phosphates and sucrose.     

Characteristics of Nectar Secretion by the Extrafloral Nectaries of Ricinus communis

The characteristics of nectar secretion by excised extrafloralnectaries of Ricinus have been examined. Secreted nectar wasfound to contain three sugars: sucrose, glucose and fructose,with glucose and fructose occurring in a 1: 1 ratio. All threesugars supported secretion when used in the culture medium andthe yield of nectar sugar was found to be concentration-dependent.Other sugar sources failed to support secretion. Experimentsusing ¹⁴C-sugars and ¹⁴CO₂ fed to intact plants allowed themovement of sugars through the nectary to be examined. Sucrosesynthesis occurs when excised glands are fed glucose and thisoccurs very early in the transport through the nectary. Themain sugar transported was sucrose, with little hydrolysis occurringuntil the final step of secretion. There was no evidence thatsucrose hydrolysis occurs either by invertase in the nectaror by a microbial flora. Inhibitors of respiration were foundto inhibit secretion as did anaerobiosis. Temperature also hada marked effect, with a temperature coefficient of 1.8. However,secretion of sucrose was not affected by anaerobic conditions,low temperatures or inhibitors of respiration as markedly asthat of glucose and fructose. Electron microscopy revealed the presence of a thickened andheavily stained wall at the inner border of the secretory epidermallayer. This wall contained numerous plasmodesmata at a frequencyof 14 per µm² and may represent an apoplastic barrier.Light microscope cytochemistry revealed that acid phosphataseis primarily located in the nectiferous tissue, while ATPaseis concentrated in the epidermis. The possibility that the nectarycontains two pathways for sucrose secretion, both apoplasticand symplastic, is discussed. Key words: Invertase, nectary, plasmodesmata, Ricinus communis, sucrose     

Ammonia Induces Starch Degradation in Chlorella Cells

When ammonia was added to cells of Chlorella which had fixed¹⁴CO₂ photo synthetically, ¹⁴C which had been incorporated intostarch was greatly decreased. A similar effect was observedwhen potassium nitrate and sodium nitrite were added. The ammonia-induceddecrease in ¹⁴C-starch was observed in all species of Chlorellatested. With cells of C. vulgaris 11h, most of the radioactivityin starch was recovered in sucrose, indicating that ammoniainduces the conversion of starch into sucrose. The percent of¹⁴C recovered in sucrose differed from species to species andpractically no recovery in sucrose was observed in C. pyrenoidosa.In most species tested, the enhancing effects of blue lightand ammonia on O₂ uptake as well as the ammonia effect on starchdegradation were greater in cells which had been starved inphosphate medium in the dark than in non-starved cells. In contrast,the enhancing effect of ammonia on dark CO₂ fixation was muchgreater in non-starved cells. C. pyrenoidosa was unique in thatblue light did not show any effect on its O₂ uptake. (Received August 15, 1984; Accepted November 16, 1984)     

Uptake and Utilization of Fructose by Anabaena variabilis ATCC 29413. Effect on Respiration and Photosynthesis

Anabaena variabilis ATCC 29413 showed a constitutive mechanismfor fructose uptake which was further enhanced by growing thecells with fructose. The uptake process was energydependentas indicated the inhibitory effect of the uncoupler carbonylcyanidem-chlorophenylhydrazone (CCCP) and the reduction induced byincubating the cells in the dark or in the light with DCMU.Cells adapted to growth on fructose showed increased rates ofrespiration both in the dark and in the light. The rate of ¹⁴CO₂evolved from radiolabelled fructose was lower in the light thanin the dark or in the presence of DCMU. This fact can be partiallyexplained by the photosynthetic reutilization of respiratory¹⁴CO₂. Modifications in photosynthesis were observed in fructose-growncells. PS I and PS II activity measured in spheroplasts obtainedby lysozyme treatment were enhanced by fructose probably asa way to compensate the lower concentration of chorophyll showedby fructose-grown cells. The photosynthetic affinity for externalCO₂ and the rate of photosynthesis dependent on external inorganiccarbon were reduced by fructose. (Received September 24, 1991; Accepted February 14, 1992)     

Synthesis and Dissolution of Starch labelled with 14Carbon in Tobacco Leaf Tissue

Tobacco leaves depleted of starch, were detached and allowedto assimilate equal amounts of ¹⁴CO₂ and ¹²CO₂in succession,and vice versa. Distribution of radioactivity in starch, andsugars was determined after assimilation and after disks cutfrom the leaves had been kept in darkness for times up to 40hours. The amount and activity of the CO₂ was also determined.¹⁴C and ¹²C were incorporated in equal amounts into starch independentlyof the order in which they were supplied. In contrast sucrosehad high activity ¹⁴C was given last, and hexose a low one.The reverse was true when ¹²C was given last. Activity of respiratoryCO₂ was slightly higher when ¹⁴C was assimilated last as comparedwith ¹²C. In the dark only ¹⁴C or ¹²C was at first lost fromstarch, in accordance or removal of discrete layers. Analyticalresults show that starch is the main respiratory substrate andto account for the redistribution of radioactivity in passageto CO₂ it is concluded that sucrose occurs at two sites separatedinter-or intra-cellularly, one of which is in equilibrium withthe system intrconverting starch and CO₂ and at the other hexosesare produced by inversion. A starch-like polysaccharide is formedduring assimilation which persists in the dark and there isa significant contribution to respiration of carbon from non-carbohydratesources when leaf disks are kept on the dark.     

Growth of Ryegrass (Lolium perenne L.) Exposed to SO2: II. CHANGES IN THE DISTRIBUTION OF PHOTO ASSIMILATED 14C

Perennial ryegrass (Lolium perenne L.) cv. S23 was exposed to0, 50, and400 µg m– ³ SO₂ for a 29 d period, harvested,and then exposed under the same regime for a further 22 d periodof regrowth. Leaves from plants representing each exposure concentrationwere photosynthetically fed ¹⁴CO₂ for 5 min at the end of eachperiod. A significant increase in photoassimilation of ¹⁴CO₂and retention of ^I4C, concomitant with significant decreasesin [¹⁴C]glycine and [¹⁴C]serine with increasing SO₂ concentration,implied that there was an inhibition of the photorespiratorypathway. At the second harvest, leaves from plants exposed to400 µg m– ³ SO₂ also exhibited significant increasesin [¹⁴C]sucrose and [¹⁴C]fructose.     

Characterization of Hexose Transporter for Facilitated Diffusion of the Tonoplast Vesicles from Pear Fruit

Tonoplast vesicles were prepared from the flesh tissue of maturepear fruit. Sugar uptakes into the vesicles determined by twodifferent methods, the membrane and the gel filtration methods,were quite similar. The uptake was highest for glucose and subsequently,in order, for fructose, sucrose and sorbitol. It was not stimulatedby addition of ATP, although the vesicles could create a protongradient. However, the uptakes were significantly inhibitedby p-chloromercuribenzene sulphonate (PCMBS, SH-reagent andinhibitor of sugar transporter). Further, the PCMBS-sensitiveuptakes of glucose and fructose saturated with their increasedconcentrations. Thus, these PCMBS-sensitive uptakes are mediatedby the transporter of facilitated diffusion. The uptakes ofglucose or fructose each had two K_m values. K_m values for glucosewere 0.35 and 18 mM, and those for fructose were 1.6 and 25raM. The uptake of 0.2 mM glucose was inhibited by 2 mM fructoseand that of 2 mM fructose was inhibited by 2 mM glucose, butneither was inhibited by sucrose or sorbitol. O-methyl-glucose(OMG) also inhibited both the glucose and fructose uptakes.Therefore, the same transporter may mediate both glucose andfructose uptakes at lower concentrations; this hexose transportsystem differed from the sucrose and sorbitol transport systems. ¹Research Fellow of the Japan Society for the Promotion of Science. ²Present address: Faculty of Agriculture, Tohoku University,1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, 981 Japan.     

Impairment of sucrose utilization for cell wall formation in the roots of aluminum-damaged cotton seedlings

Shoots of normal cotton seedlings rapidly fixed a pulse of ¹⁴CO₂from the ambient atmosphere and translocated some of the resultinglabeled sucrose to the roots. Roots of these plants assimilatedmost of the radioactivity from a 10-min labeling pulse intoinsoluble cell wall materials and other stable metabolites within4 to 6 hr after the pulse. However, roots of cotton seedlingswhich had been exposed to 1 ppm of Al^3$ for 24 hr before labelingtended to accumulate the ¹⁴C-label as free sucrose. Histologicand microautoradiographic evidence suggested that Al^3$ impairedthe root's capacity to utilize sucrose in further metabolicproducts so that ¹⁴C-labeled sucrose was not polymerized intocell wall materials as it was in the roots of control plants. (Received July 7, 1971; )     

Carbon dioxide assimilation and photosystem II deficiency in bundle sheath strands isolated from C4 plants

The activities of Hill reaction and photosynthetic ¹⁴CO₂ fixationin bundle sheath strands enzymatically isolated from millet(Panicum miliaceum) were 3–15 times as high as those observedin corn (Zea mays). In both preparations, 3-phosphoglyceratewas the initial ¹⁴CO₂ fixation product and the radioactivitywas incorporated into sucrose and insoluble compounds (glucose-polymers)during the later period. After 20 sec of photosynthetic ¹⁴CO₂fixation, the percent of ¹⁴C incorporated into sugar phosphatesin millet was about 3 times as high as that in corn, while incorn, the percent of ¹⁴C in 3-phosphoglycerate was higher thanthat observed in millet throughout the experimental period.When ¹⁴C-phosphoglycerate was added to the isolated bundle sheathstrands, the rates of transfer of the radioactivity to dihydroxyacetonephosphate and sugar diphosphates in millet were significantlyhigher than those in corn. These results indicate that in thebundle sheath strands isolated from corn in which photosystemII activity is deficient, the reductive pentose cycle is impairedat the reduction step of 3-phosphoglycerate to glyceraldehydephosphate due to the limited supply of NADPH through the photoelectrontransport system. In contrast, the bundle sheath strands isolatedfrom millet which have adequate photosystem II activity cancarry out normal photosynthetic CO₂ fixation. (Received January 23, 1975; )     

Photorespiratory Amino Donors, Sucrose Synthesis and the Induction of CO2 Fixation in Barley Deficient in Glutamine Synthetase and/or Glutamate Synthase

Murray, A. J. S., Black well, R. D., Lea, P. J. and Joy, K.W. 1988. Photorespiratory amino donors, sucrose synthesis andthe induction of CO₂ fixation in barley deficient in glutaminesynthetase and/or glutamate synthase.—J. exp. Bot. 39:845–858. A number of mutants of barley have been produced which lackboth chloroplastic glutamine synthetase and ferredoxin-dependentglutamate synthase activities. The plants accumulated ammoniato the same extent as mutants deficient in only glutamine synthetasebut shared the gas-exchange characteristics of the glutamatesynthase deficient parent. These mutants have been used to demonstratedirectly the ability of alanine to ameliorate the dramatic dropin fixation rate normally exhibited by glutamate synthase deficientmutants on transfer to photorespiratory conditions. Immediatelyafter transfer to air, the mutants deficient in glutamate synthaseactivity demonstrated a reduced ability to incorporate ¹⁴C derivedfrom ¹⁴CO₂ into sucrose. This effect was, however, dependenton the previous induction of CO₂ fixation. Use of ¹⁴CO₂ revealedthat the induction phase of CO₂ fixation was altered in allthree mutants. Neither of the parents nor the double mutantaccumulated sucrose in air under conditions which promote sucroseaccumulation by the wild type. The implications of these resultsfor photosynthesis and the control of sucrose synthesis arediscussed. Key words: Photorespiratory barley mutant, amino donors, sucrose, GS, glutamate synthase.