Pathway of phloem loading in the C3 tropical orchid hybrid Oncidium Goldiana

The apoplast of mature leaves of the tropical orchid OncidiumGoldiana was perfused with 0.5 mM p-chloromercuribenzenesulphonicacid (PCMBS) via the transpiration stream in order to test themode of phloem loading. The efficacy of introducing PCMBS byperfusion was shown by saffranin O dye movement in the veinsand leaf apoplast in control experiments. Photoassimilate exportas the result of phloem loading was measured by collection of¹⁴CO₂-derived photoassimilates from the basal cut-ends of intactleaves. Phloem loading and translocation of photoassimilates was inhibitedby 89% in leaves perfused with PCMBS for 1 h. The effect ofPCMBS on leaf photosynthesis was minimal. The amount of radiocarbonfixed by PCMBS-treated leaves averaged 89% of control leavesperfused with distilled water. A negative correlation betweenthe total amount of photoassimilate exuded and the calculatedconcentration of PCMBS in the leaf apoplast was also observed.The results indicate that phloem loading in Oncidium Goldianaoccurs via the apoplastic pathway. Key words: Phloem loading, apoplast, PCMBS, tropical orchid     

Characteristics of the phloem path: analysis and distribution of carbohydrates in the petiole of Cyclamen

Compartmentation fluxes of carbohydrates along the phloem path were analysed in the petiole of Cyclamen persicum (L.) Mill. Sucrose represented the dominant fraction (58-75% of soluble carbohydrates in the vascular symplast). Planteose (12-22%), glucose (3-8%) and fructose (3-13%) occurred in lower amounts (data from liquid chromatography, percentages of the total peak area). Starch was not detectable. Upon feeding leaves with ¹⁴CO2, 98% and 90% of radiolabel was recovered as sucrose in the vascular symplast after 3 h and 24 h, respectively. Thus, sucrose appeared to be the exclusive transport sugar in Cyclamen. Experiments with asymmetrically labelled sucrose revealed that there was no metabolism of translocated sucrose. Analysis of six consecutive petiole segments (each 2 cm in length) showed a homogeneous longitudinal distribution of these sugars differed markedly. On average, the sucrose concentration amounted to 4.7 and 0.4 mg g^-1 FM in the vascular apoplast and petiole parenchyma, respectively. Sucrose was unloaded with out hydrolysis and stored in the periphery of the phloem path. Planteose was identified as another storage saccharide. Sucrose synthesis by sucrose phosphate synthase occurred when isolated vascular bundles were incubated with [¹⁴C]glucose or [¹⁴C]fructose. These data suggest that the phloem path is characterized by both source and sink like activity.     

Sucrose Transport and Phloem Unloading in Stem of Vicia faba: Possible Involvement of a Sucrose Carrier and Osmotic Regulation

Stems of Vicia faba plants were used to study phloem unloading because they are hollow and have a simple anatomical structure that facilitates access to the unloading site. After pulse labeling of a source leaf with ¹⁴CO₂, stem sections were cut and the efflux characteristics of ¹⁴C-labeled sugars into various buffered solutions were determined. Radiolabeled sucrose was shown to remain localized in the phloem and adjacent phloem parenchyma tissues after a 2-hour chase. Therefore, sucrose leakage from stem segments prepared following a 75-minute chase period was assumed to be characteristic of phloem unloading. The efflux of ¹⁴C assimilates from the phloem was enhanced by 1 millimolar p-chloromercuribenzene sulfonic acid (PCMBS) and by 5 micromolar carbonyl cyanide m-chlorophenly hydrazone (CCCP). However, PCMBS inhibited and CCCP enhanced general leakage of nonradioactive sugars from the stem segments. Sucrose at concentrations of 50 millimolar in the free space increased efflux of [¹⁴C]sucrose, presumably through an exchange mechanism. This exchange was inhibited by PCMBS and abolished by 0.2 molar mannitol. Increasing the osmotic concentration of the efflux medium with mannitol reduced [¹⁴C]sucrose efflux. However, this inhibition seems not to be specific to sucrose unloading since leakage of total sugars, nonlabeled sucrose, glucose, and amino acids from the bulk of the tissue was reduced in a similar manner. The data suggest that phloem unloading in cut stem segments is consistent with passive efflux of sucrose from the phloem to the apoplast and that sucrose exchange via a membrane carrier may be involved. This is consistent with the known conductive function of the stem tissues, and contrasts with the apparent nature and function of unloading in developing seeds.     

The Cellular Pathway of Radial Transfer of Photosynthates in Stems of Phaseolus vulgaris L.: Effects of Cellular Plasmolysis and p-Chloromercuribenzene Sulphonic Acid

Cellular plasmolysis with l M solutions of mannitol appearedto sever plasmodesmatal interconnections between all cells ofthe stems of Phaseolus vulgaris plants except the sieve element-companioncell (se—cc) complexes. Phloem loading and uptake of [¹⁴C]sucroseby the storage cells of the stems was unimpaired by cellularplasmolysis followed by rehydration of the stem tissues. Accumulationof phloem-transported ¹⁴C-photosynthates of the treated stemswas inhibited in summer-grown plants and unaffected in winter-grownplants indicating that phloem unloading follows a symplasticand a free-space route respectively depending on growth season.At a concentration that did not interfere with cellular metabolism,p-chloromercuribenzene sulphonic acid (PCMBS) applied to thestems blocked [¹⁴C]sucrose loading into the phloem and storagecells of the stem, but had no effect on the pool size of free-spacesugars. This latter response is consistent with a facilitatedmechanism of sugar unloading to the stem free-space. Accumulationof phloem-transported ¹⁴C-photosynthates was stimulated by PCMBSand this effect was most pronounced in winter-grown plants.Cellular plasmolysis followed by rehydration abolished the PCMBSaction on ¹⁴C-photosynthate accumulation. This effect is consistentwith a PCMBS induction of phloem unloading through the stemsymplast. It is proposed that phloem unloading in bean stemsmay follow either a free-space or symplastic route and thatthe latter route is entrained under sink-limited conditions. Phaseolus vulgaris, french bean, stem, phioem unloading, free-space, symplast     

Phloem Loading in Coleus blumei in the Absence of Carrier-Mediated Uptake of Export Sugar from the Apoplast

Phloem loading in Coleus blumei Benth. leaves cannot be explained by carrier-mediated transport of export sugar from the apoplast into the sieve element-companion cell complex, the mechanism by which sucrose is thought to load in other species that have been studied in detail. Uptake profiles of the export sugars sucrose, raffinose, and stachyose into leaf discs were composed of two components, one saturable and the other not. Saturable (carrier-mediated) uptake of all three sugars was almost completely eliminated by the inhibitor p-chloromercuribenzenesulfonic acid (PCMBS). However, when PCMBS was introduced by transpiration into mature leaves it did not prevent accumulation of ¹⁴C-photosynthate in minor veins or translocation of labeled photosynthate from green to nonchlorophyllous regions of the leaf following exposure to ¹⁴CO₂. The efficacy of introducing inhibitor solutions in the transpiration stream was proven by observing saffranin O and calcofluor white movement in the minor veins and leaf apoplast. PCMBS introduced by transpiration completely inhibited phloem loading in tobacco leaves. Phloem loading in C. blumei was also studied in plasmolysis experiments. The carbohydrate content of leaves was lowered by keeping plants in the dark and then increased by exposing them to light. The solute level of intermediary cells increased in the light (phloem loading) in both PCMBS-treated and control tissues. A mechanism of symplastic phloem loading is proposed for species that translocate the raffinose series of oligosaccharides.     

Transport of Photoassimilate in Pea Ovules

Interpretation of tracer washout from an attached empty seedcoat depends on whether photoassimilate within the apoplastof the seed coat is absorbed by the seed coat tissues. Usingsucrose trapping procedures, we were unable to see any evidencefor sucrose uptake from the seed coat apoplast which would beneeded to provide the seed coat with its carbohydrate requirementsif phloem unloading were into the apoplast. Once released intothe apoplast photoassimilate is unavailable to the seed coattissue. Changes between equimolar solutions of sorbitol andsorbitol/sucrose mixes induced small transient responses inseed coat unloading which suggest that sorbitol and sucrosehad different reflection coefficients and gave water relationresponses with rapid, and fatiguable, osmoregulation withinthe seed coat. Immediate inhibition of seed coat unloading with PCMBS is reported,followed by inhibition of import into the entire pod. PCMBSappears to be xylem mobile, thereby quickly being dispersedthroughout the entire experimental pod. A complex CCCP responseis reported, which is consistent with immediate inhibition ofsymplastic transport followed by membrane disruption. AlthoughCCCP inhibited seed coat unloading, there was no effect on ovuleimport. This has been interpreted as evidence that the seedcoat has an active role in control of photoassimilate importinto ovules. Key words: Pisum sativum, phloem unloading, seed coat unloading     

Cellular pathway of photosynthate transport in coats of developing seed of Vicia faba L. and Phaseolus vulgaris L. I. Extent of transport through the coat symplast

The extent of post-phloem solute transport through the coatsymplasts of developing seeds of Vicia faba L. and Phaseolusvulgaris L. was evaluated. For Vicia seed coats, the membrane-impermeantfluorochrome, CF, moved radially from the chalazal vein to reachthe chlorenchyma and thin-walled parenchyma transfer cell layers.Thereafter, the fluorochrome moved laterally in these two celllayers around the entire circumference of the seed coat. Transferof CF from the chalazal vein was inhibited by plasmolysis ofattached ‘empty’ seed coats. In contrast, the spreadof phloem imported CF was restricted to the ground parenchymaof Phaseolus seed coats. Fluorochrome loaded into the outermostground parenchyma cell layer was rendered immobile followingplasmolysis of excised seed-coat halves. Phloem-imported [¹⁴C]sucroseand the slowly membrane permeable sugar, L-[¹⁴C]glucose, werepartitioned identically between the vascular and non-vascularregions of intact Vicia seed coats. For ¹⁴C-photosynthates,these partitioning patterns in attached ‘empty’Vicia seed coats were unaffected by PCMBS, but inhibited byplasmolysis. Tissue autoradiographs of intact Phaseolus seedcoats demonstrated that a pulse of ¹⁴C-photosynthate moved fromthe veins to the grounds tissues. In excised Vicia seed coats,preloaded with ¹⁴C-photosynthates, the cellular distributionof residual ¹⁴C-label was unaffected by PCMBS. In contrast,PCMBS caused the ¹⁴C-photosynthate levels to be elevated inthe veins and ground parenchyma relative to the branch parenchymaof Phaseolusseed coat halves. Based on the above findings, itis concluded that the phloem of Vicia seed coats is interconnectedto two major symplastic domains; one comprises the chlorenchyma,the other the thin-walled parenchyma plus thin-walled parenchymatransfer cells. For Phaseolusseed coats, the phloem forms amajor symplastic domain with the ground parenchyma. Key words: Phaseolus vulgaris L, phloem unloading, photosynthate transport, seed coat, symplast, Vicia faba L     

Photosynthate Unloading from Seed Coats of Phaseolus vulgaris L.--Nature and Cellular Location of Turgor-Sensitive Unloading

Patrick, J. W., Jacobs, E., Offler, C. E. and Cram, W. J. 1986.Photosynthate unloading from seed coats of Phaseolus vulgarisL.—Nature and cellular location of turgor-sensitive unloading—J.exp. Bot. 37: 1006–1019. Unloading rates of ¹⁴C-Photosynthates from excised seed-coathalves of Phaseolus vulgaris L. plants were sharply increasedat cell turgor potentials in excess of 5 ? 10⁵ Pa. Turgor-sensitiveunloading occurred in the absence of any change in the passivepermeability of, and active sucrose influx across, the plasmalemmaand tonoplast membranes. The proton ionophore CCCP, and lowtemperature significantly slowed turgor-sensitive unloadingwhile PCMBS, a non-permeating sulphydryl-modifying compound,was without effect. Turgor-sensitive unloading significantlydepressed the ¹⁴C-Photosynthate content of the ground and branchparenchyma, but had no effect on the ¹⁴C-Photosynthate levelsin the vascular tissues. Cycling of cell turgor potentials aboveand below 5 ? 10⁵ Pa elicited reproducible responses in theunloading rate of ¹⁴C-Photosynthates. Increasing turgor above5 ? 10⁵ Pa resulted in a burst of ¹⁴C-Photosynthate unloading.Reversal to turgors less than 5 ? 10⁵ Pa caused a rapid depressionin unloading rate. It is proposed that turgor-sensitive unloadingis facilitated by a specific turgor-sensitive porter locatedon the plasmalemma of the ground and/or branch parenchyma cellsof bean seed coats. Key words: Bean, seed coat, turgor-sensitive unloading, phloem     

Pathways of Uptake and Accumulation of Sugars in Tomato Fruit

The route of sucrose unloading from the conducting tissue, theregulation of sucrose hydrolysis and the uptake and subsequentmetabolism of sugars were investigated in the rapidly growingtomato fruit. During the first two weeks of fruit enlargement, the vacuoleaccounted for more than 85% of the protoplast volume and theintercellular space accounted for 20% of the fruit placentaltissue. The plasmodesmatal frequency was highest between phloemparenchyma cells and lowest between phloem sieve cells and phloemparenchyma. The total invertase activity was about 8 µmolglucose g^–1 d. wt min^–1 during the rapid growingperiod and increased six-fold at ripening. The wall-bound invertaseaccounted for less than 11% of the total activity. Invertaseactivity increased with increasing sucrose concentrations (upto 50 mM) in the incubation medium, but decreased at higherconcentrations. Sucrose synthase activity could only be detectedwhen fruit was older than 19 d. The uptake and metabolism of sugars by fruit cells were investigatedby incubation of fruit slices with ¹⁴C-sugars for 3 h. The uptakeof sucrose increased with the sucrose concentration up to 200mM. The rate of glucose uptake and its conversion to the ethanol-insolublefraction were higher than those of sucrose. The uptake of sucrosedid not compete with that of glucose or vice versa, providedthe osmotic potential of the incubation solution was maintainedconstant. The uptake of sucrose was not inhibited by metabolicinhibitors such as PCMBS, CCCP, sodium azide or vanadate. TheATPase activity in the fruit tissue was low. These findings did not identify conclusively the mode of sucroseunloading. However, the uptake of sugars by fruit cells is non-specificand does not appear to require a membrane carrier or plasmalemmaATPase to provide energy for sucrose uptake. Fruit, invertase, Lycopersicon esculentum, phloem unloading, plasmodesmata, sucrose     

Phloem Unloading in Developing Leaves of Sugar Beet : I. Evidence for Pathway through the Symplast

Physiological and transport data are presented in support of a symplastic pathway of phloem unloading in importing leaves of Beta vulgaris L. (`Klein E multigerm'). The sulfhydryl reagent p-chloromercuribenzene sulfonic acid (PCMBS) at concentration of 10 millimolar inhibited uptake of exogenous [¹⁴C]sucrose by sink leaf tissue over sucrose concentrations of 0.1 to 5.0 millimolar. Inhibited uptake was 24% of controls. The same PCMBS treatment did not affect import of ¹⁴C-label into sink leaves during steady state labeling of a source leaf with ¹⁴CO₂. Lack of inhibition of import implies that sucrose did not pass through the free space during unloading. A passively transported xenobiotic sugar, l-[¹⁴C]glucose, imported by a sink leaf through the phloem, was evenly distributed throughout the leaf as seen by whole-leaf autoradiography. In contrast, l-[¹⁴C]glucose supplied to the apoplast through the cut petiole or into a vein of a sink leaf collected mainly in the vicinity of the major veins with little entering the mesophyll. These patterns are best explained by transport through the symplast from phloem to mesophyll.     

Sectorial Patterns in Leaves on Fruit Tree Shoots Produced by Radioactive Assimilates and Solutions

In vigorously growing shoots of apple and plum ¹⁴C-assimilateswere translocated from a ‘fed’ leaf to particularsectors of other leaves in a distribution pattern associatedwith the phyllotaxis; the same sectorial and distribution patternswere produced by ³²P phosphate solution taken into the shootthrough a cut petiole. The frequency with which a given sectorialpattern occurred at a particular position on the phyllotacticspiral was ascertained. Such patterns were not observed abovethe third rolled leaf in the apple shoot apex. Killing the phloem in the petiole prevented egress of labelledassimilate but not of ³²P solution. Barkringing above the sourceleaf reduced, but did not completely prevent, assimilate movementup the stem, suggesting some translocation in the xylem. Distribution of label from ⁴⁵CaCl₂, ⁸⁶RbCl and [³H]asparagine,incorporated through cut petioles, did not follow the same patternas label from ³²P solutions. Malus pumila Mill., apple, Prunus domestica L., Prunus insititia. L., leaf plum, patterns, transport of radioisotopes, vascular phyllotaxis     

Translocation in Plants Possessing Supernumerary Phloem: II. THE INCLUDED PHLOEM OF BOUGAINVILLEA GLABRA CHOISY AND THE BICOLATERAL BUNDLES OF CUCUMBER (CUCUMIS SATIVUS L.)

The translocation path in the included phloem of Bougainvilleaand in the bicolateral bundles of cucumber was studied by exposingyoung branches to ¹⁴CO₂ and detecting the radioactive compoundsby autoradiography. In Bougainvillea, the structure and functionof the phloem system is comparatively uncomplicated and uniform.All phloem bundles, i. e. those which are located in parenchymatoustissue of the central zone and those embedded in the secondaryxylem, become labelled. Exogenous IAA was translocated in thebundles, but the exact mode of translocation was not ascertained.Apical dominance was not affected by girdling. The implicationof this fact is discussed with respect to the translocationof the auxin that determines the correlative inhibition involvedin apical dominance. In cucumber the inner phloem became labelled throughout theplant to a lesser extent than the outer phloem. However, inthe petiole of the assimilating leaf the intensity of the labelwas the same in both inner and outer phloem, although the innerphloem has fewer elements. Below the treated leaf the innerphloem translocated less than the outer phloem. Above this leafthe inner phloem was entirely unlabelled     

Assimilation of gaseous ammonia and the transport of its products in barley and spinach leaves

The interactions between the assimilation and transport of nitrogenand carbon were investigated in barley and spinach leaves. Bothplants were fumigated with NH₃ (1 mg m^–3 and the contentof amino acids, sucrose and carbon intermediates of amino acidmetabolism were analysed in the leaves, apoplast and phloemsap. The following changes took place in the C- and N-metabolismof barley leaves during 5 h of fumigation with NH₃ (a) The contentsof amino acids, especially glutamine, largely increased andthe contents of sucrose, 2-oxoglutarate, phosphoenolpyruvate,and glycerate-3-phosphate declined. (b) A decrease in the phophoenolpyruvatecontent was accompanied by an increased activity of phosphoenolpyruvatecarboxylase. (c) The altered cytosolic concentrations of aminoacids and sucrose during NH₃ fumigation correlated with similarchanges in the apoplast and phloem sap. The altered percentageof each amino acid relative to the total amino acid concentrationin the cytosol, caused by NH₃ fumigation, is reflected in theapoplast and the phloem sap. The results indicate that the concentrations of amino acids in the cytosol determine their concentrationsin the phloem. Key words: Amino acids, ammonia fumigation, barley leaves, C: N partitioning, phosphoenolpyruvate carboxylase, phloem sap, spinach leaves     

The Pathway of Radial Transfer of Photosynthate in Decapitated Stems of Phaseolus vulgaris L.

[¹⁴C]Sucrose was found to be the predominant component of the¹⁴C-photosynthates that accumulated in the free space of decapitatedstems of P. vulgaris plants. The ¹⁴C-photosynthates appearedto occupy the entire free-space volume of the stems at totalsugar concentrations in the range of 3–12 mM. The free-spacesugar levels were found to rapidly decline once photosynthatetransfer to the stems was halted. Moreover, it was found thatestimates of the rate of in vitro sucrose uptake by the stemscould account fully for the decline in free-space sugar levels.Overall, the evidence indicated that at least part of the radialpathway of photosynthate transfer in bean stems involved thestem apoplast. It is tentatively proposed that, based on celland tissue distribution of ¹⁴C-photosynthates, the apoplasticpathway extends from the membrane boundary of the sieve element/companion-cellcomplex to all other cells of the stem. Apoplast, Phaseolus vulgaris L., bean, phloem unloading, photosynthates, symplast     

Germination Behaviour of Solanum nigrum Seeds

Sucrose has been found in the apoplast of bean stems at a concentrationof 25–60 mM with an axial concentration gradient in theappropriate direction for Munch translocation. Removal of theepidermis from a 50 mm length of stem enabled the washout oflabelled photosynthate from the apoplast. The rate of labelwashout was strongly dependent on temperature, and the rateincreased on blockage of phloem pathways to the main sink forthat assimilate. Washout did not reduce when the bathed tissuewas plasmolyzed. We propose that sucrose is unloaded from thephloem into the apoplast, and a sucrose concentration is maintainedthere by a balance of sucrose uptake into sink tissue or reloadinginto the phloem. It is proposed that the apoplastic pool ofphotosynthate can act to buffer sudden changes in phloem contentswhen there are rapid changes in source-sink configuration. Key words: Sucrose, Phaseolus vulgaris, Apoplast, Phloem unloading     

An Investigation of the Relay Hypothesis of Phloem Transport in Ricinus communis L.

To test for the existence of an apoplastic unloading/reloadingstep in phloem translocation, as envisaged in the relay hypothesisof phloem transport, isotopic trapping experiments were performedon Ricinus comunis L. var. Gibsonii. A CO₂ buffer system wasused to supply ¹⁴CO₂ at constant partial pressure and constantspecific activity to a photosynthesizing leaf. The subtendinginternode was perfused with solutions of sugars or of mannitoland transolation of ¹⁴C past the perfused zone was monitoredby the collection of phloem exudate. Trapping of activity inthe perfusate was enhanced by the presence of sugars as wouldbe expected with an unloading/reloading process. However, therewas no evidence that introduction of the unlabelled sugars tothe apoplast also reduced activity in the phloem exudate. Moreover,the rate of loss of activity to the perfusing solutions representedonly 1% of the rate of appearance in the exudation. It is suggestedthat the trapping results may reflect an unloading of tracerfrom the phloem associated with a subsequent reloading by adjacenttissues rather than by the sieve tubes. To investigate the length of sieve tube continuity in Ricinus,a horizontal incision was made to the bark and the rate of exudationof phloem sap was monitored. Successive circumferential cutswere made above the exuding incision and progressively closerto it. In general, a girdling incision produced a transientdecrease in the rate of change of exudation rate (i.e. the firstderivative became more negative/less positive). The magnitudeof this response rose with exudation rate and fell with thedistance at which a girdling cut was made. Fitting an appropriatemodel yielded an estimate for contributory length of 69 ±6 cm. This was comparable with the distance of the initial tangentialincision from the stem apex, suggesting a continuous sieve tubesystem in Ricinus. A similar investigation on the petiole yieldedan estimate of around 7.0 cm. This lower estimate for contributorylength is believed to reflect a rapid sealing process that limitsthe distance of propagation of turgor-release rather than alimited length of sieve tube continuity. The results of this investigation do not support the relay hypothesisof phloem transport. Rather they suggest a continuous sievetube system which has a distributed capacity to load and unloadsolutes, and which may exhibit a sealing response when injured. Key words: Ricinus communis L, phloem transport, phleom unloading     

Apoplastic Phloem Unloading in the Stem of Bean

Sucrose has been found in the apoplast of bean stems at a concentrationof 25–60 mM with an axial concentration gradient in theappropriate direction for Munch translocation. Removal of theepidermis from a 50 mm length of stem enabled the washout oflabelled photosynthate from the apoplast. The rate of labelwashout was strongly dependent on temperature, and the rateincreased on blockage of phloem pathways to the main sink forthat assimilate. Washout did not reduce when the bathed tissuewas plasmolyzed. We propose that sucrose is unloaded from thephloem into the apoplast, and a sucrose concentration is maintainedthere by a balance of sucrose uptake into sink tissue or reloadinginto the phloem. It is proposed that the apoplastic pool ofphotosynthate can act to buffer sudden changes in phloem contentswhen there are rapid changes in source-sink configuration. Key words: Sucrose, Phaseolus vulgaris, Apoplast, Phloem unloading     

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; )     

Phloem loading in Vicia faba leaves: Effect of N-ethylmaleimide and parachloromercuribenzenesulfonic acid on H+ extrusion,K+ and sucrose uptake

The effects of a penetrating (NEM) and a non-penetrating (PCMBS) sulfhydryl-specific reagent on proton extrusion, ⁸⁶Rb and [U-¹⁴C]sucrose uptake by Vicia faba leaves have been studied. Proton extrusion was strongly or completely inhibited by 0.1 mM NEM. ⁸⁶Rb and [U-¹⁴C]sucrose uptake were markedly reduced by NEM concentrations equal to or higher than 0.5 mM. Under our experimental conditions, PCMBS (1 mM) exerted a strong inhibition on [¹⁴C]sucrose uptake but did not inhibit proton extrusion and ⁸⁶Rb uptake. The sensitivity of phloem loading to PCMBS is thought to be a consequence of sugar-carrier blockage and not of inhibition of the proton pump.Abbreviations CCCP carbonylcyanide-m-chlorophenylhydrazone - DES diethylstilbestrol - DCCD dicyclohexylcarbodiimide - FC Fusicoccin - NEM N-ethylmaleimide - PCMBS p-chloromercuribenzenesulfonic acid     

A three-step screening procedure to identify the mode of phloem loading in intact leaves

A three-step screening method was developed to identify the mode of phloem loading in intact leaves. Phloem loading of ¹⁴CO₂-derived photosynthate was challenged by p-chloromercuribenzenesulfonic acid (PCMBS) in leaves of dicotyledons with either a symplasmic (type 1, with intermediary cells as companion cells) or apoplasmic (type 2b, with transfer cells as companion cells) minor-vein configuration. Firstly, photosynthate export as the result of phloem loading was measured by collection of phloem exudate from the petiole. The PCMBS had virtually no effect on photosynthate export in representatives of type-1 families (Lamiaceae, Lythraceae, Onagraceae, Saxifragaceae). In contrast, photosynthate export was strongly reduced by PCMBS in representatives of type-2b families (Asteraceae, Balsaminaceae, Dipsacaceae, Linaceae, Tropaeolaceae, Valerianaceae) and type-2b members of polytypical families (Fabaceae, Scrophulariaceae). Secondly, densitometric measurements of leaf autoradiographs demonstrated that the contrast between the mesophyll and the lower-order veins was hardly affected by PCMBS treatment in type-1 species, whereas PCMBS strongly reduced the contrast in type-2b species. Thirdly, separate ¹⁴C-radioassays of vein and mesophyll tissues confirmed this observation. The three-step procedure thus revealed a strong and consistent reduction of phloem loading by PCMBS in type-2b species which was absent in type-1 species. In conclusion, phloem loading in type-2b species occurs via the apoplast and type-1 species execute an alternative — most likely symplasmic — mode of phloem loading.Abbreviations PCMBS p-chloromercuribenzenesulfonic acid - SE/CC-complex sieve element/companion cell complex We gratefully acknowledge the expert help of Dr. Maarten Terlou, Department of Image Processing and Design, University of Utrecht, in carrying out the densitometric measurements.