The influence of externally supplied sucrose on phloem transport in the maize leaf strip

Sucrose (2,5–1000 mmol l^–1), labeled with [¹⁴C]sucrose, was taken up by the xylem when supplied to one end of a 30-cm-long leaf strip of Zea mays L. cv. Prior. The sugar was loaded into the phloem and transported to the opposite end, which was immersed in diluted Hoagland's nutrient solution. When the Hoagland's solution at the opposite end was replaced by unlabeled sucrose solution of the same molarity as the labeled one, the two solutions met near the middle of the leaf strip, as indicated by radioautographs. In the dark, translocation of ¹⁴C-labeled assimilates was always directed away from the site of sucrose application, its distance depending on sugar concentration and translocation time. When sucrose was applied to both ends of the leaf strip, translocation of ¹⁴C-labeled assimilates was directed toward the lower sugar concentration. In the light, transport of ¹⁴-C-labeled assimilates can be directed (1) toward the morphological base of the leaf strip only (light effect), (2) toward the base and away from the site of sucrose application (light and sucrose effect), or (3) away from the site of sucrose application independent of the (basipetal or acropetal) direction (sucrose effect). The strength of a sink, represented by the darkened half of a leaf strip, can be reduced by applying sucrose (at least 25 mmol l^–1) to the darkened end of the leaf strip. However, equimolar sucrose solutions applied to both ends do not affect the strength of the dark sink. Only above 75 mmol l^–1 sucrose was the sink effect of the darnened part of the leaf strip reduced. Presumably, increasing the sucrose concentration replenishes the leaf tissue more rapidly, and photosynthates from the illuminated part of the leaf strip are imported to a lesser extent by the dark sink.Supported by Deutsche Forschungsgemeinschaft     

Characterization of phloem exudation from castor-bean cotyledons

Exudate was collected fromRicinus communis L. cotyledons after cutting the hypocotyl. It contained high levels of sucrose and potassium, a low level of calcium, and a pH of approx. 7.5. After application of [¹⁴C] sucrose to the cotyledons, radioactivity could be recovered from the exudate, indicating that the exudate was derived from the phloem. Using data from a number of individual seedlings, correlations between loading rates of sucrose, translocation rates, and sucrose and potassium contents were analyzed. A positive correlation was found between the rate of sucrose loading and the rate of sucrose exudation, whereas a negative correlation existed between the contents of sucrose and potassium in the phloem.     

Role of phloem in sucrose transport by Ricinus cotyledons

Sucrose is taken up and accumulated by cotyledons of Ricinus communis L. Autoradiographic studies reveal a predominant accumulation of sucrose in the phloem of the cotyledons. The export of sucrose from the cotyledons to hypocotyl and roots proceeds in the phloem by mass flow. These results, taken together with previous data, are experimental evidence for proton-sucrose symport as the mechanism of phloem loading.     

Ethylene-promoted tomato flower abscission and the possible involvement of an inhibitor

The abscission zone in tomato (Lycopersicon esculentum (L.) Mill. flower pedicels is morphologically distinguishable prior to separation and is delineated by an indentation of the epidermis. Exposure of excised pedicels with the flower attached to ethylene results in abscission within 12 h and this can be accelerated by flower removal. Abscission of excised pedicels with the flower removed takes place in the absence of exogenous ethylene but this is delayed by pretreatment with aminoethoxyvinyl glycine, an inhibitor of ethylene biosynthesis. The data presented support the hypothesis that flower tissue is the source of an abscission inhibitor.Abbreviations AVG aminoethoxyvinyl glycine - IAA indole-3-acetic acid     

Transfer of exogenous auxin from the phloem to the polar auxin transport pathway in pea (Pisum sativum L.)

When [1-¹⁴C]indol-3yl-acetic acid ([1-¹⁴C]IAA) was applied to the upper surface of a mature foliage leaf of garden pea (Pisum sativum L. cv. Alderman), ¹⁴C effluxed basipetally but not acropetally from 30-mm-long internode segments excised 4 h after the application of [1-¹⁴C]IAA. This basipetal efflux was strongly inhibited by the inclusion of 3.10^–6 mol· dm³ N-1-naphthylphthalamic acid (NPA) in the efflux buffer. In contrast, when [¹⁴C] sucrose was applied to the leaf, the efflux of label from stem segments excised subsequently was neither polar nor sensitive to NPA. The [1-¹⁴C]IAA was initially exported from mature leaves in the phloem — transport was rapid and apolar; label was recovered from aphids feeding on the stem; and label was recovered in exudates collected from severed petioles in 20 mM ethylenediaminetetraacetic acid. No ¹⁴C was detected in aphids feeding on the stems of plants to which [1-¹⁴C]IAA had been applied apically, even though the internode on which they were feeding transported considerable quantities of label. Localised applications of NPA to the stem strongly inhibited the basipetal transport of apically applied [1-¹⁴C]IAA, but did not affect transport of [1-¹⁴C]IAA in the phloem. These results demonstrate for the first time that IAA exported from leaves in the phloem can be transferred into the extravascular polar auxin transport pathway but that reciprocal transfer probably does not occur. In intact plants, transfer of foliar-applied [1-¹⁴C]IAA from the phloem to the polar auxin transport pathway was confined to immature tissues at the shoot apex. In plants in which all tissues above the fed leaf were removed before labelling, a limited transfer of IAA occurred in more mature regions of the stem.Abbreviations IAA indol-3yl-acetic acid - EDTA ethylenediaminetetraacetic acid - NPA N-1-naphthylphthalamic acid We are grateful to the Nuffield Foundation for supporting this research under the NUF-URB95 scheme and for the provision of a bursary to A.J.C. We thank Professor Dennis A. Baker for constructive comments on a draft of this paper and Mrs. Rosemary Bell for her able technical assistance.     

Reactivation of phloem export in mature maize leaves after a dark period

Mature leaves of corn plants (Zea mays L. cv. Prior) which were darkened for 48 h contain neither bundle-sheath starch nor glucose, and their sucrose content is below 5 M. In such leaves phloem export has ceased. When re-illuminated, photosynthetic sucrose production starts without delay, but the sucrose: glucose ratio is 1.25:1. Obviously, most of the new-formed sugar is utilized locally. Labeling with ¹⁴CO₂ has shown that phloen export starts 30 to 40 min after the onset of photosynthesis, when the sucrose: glucose ratio has increased to 13:1. The first newly formed starch can be detected when phloem export is reactivated. Glucose content remains constantly low af about 2 M for at least 2 h, and it never exceeds 10 M. Radioactivity in the exporting veins is about five times higher after 2 to 7 h of re-illumination than in the 14-h-day plant. Therefore, phloem export is either intensified during the period of reactivation or exported assimilates are partly unloaded along their way. Comparison of photosynthetic activity of equal-sized leaf strips has shown that both accumulation of photosynthates and radioactivity of exporting veins are about three times higher in the detached strip than in the strip which remained attached to the mother plant.     

Sucrose uptake in isolated phloem of celery is a single saturable transport system

The uptake of different sugars was studied in segments of isolated phloem from petioles of celery (Apium graveolens L.) in order to determine the kinetics and specificity of phloem loading in this highly uniform conductive tissue. The uptake kinetics of sucrose and the sugar alcohol, mannitol, which are both phloem-translocated, indicated presence of a single saturable system, while uptake of non-phloem sugars (glucose and 3-O-methylglucose) exhibited biphasic kinetics with lower uptake rates than those for sucrose and mannitol. The presence of unlabeled mannitol, 3-O-methylglucose and maltose in the incubation solution did not cause inhibition of labeled-sucrose uptake, indicating high carrier specificity and lack of sucrose hydrolysis in vivo. The pH optimum for sucrose uptake was 5–6. Furthermore, a rapid and transient alkalinization of the external media by sucrose indicated a sugar/H⁺-cotransport mechanism. Dual-labeling experiments showed that sucrose influx continued at a constant rate (V _max=15 mol·h^-1·(g FW)^-1), whereas sucrose efflux was low and insensitive to external concentration. Therefore, the saturable uptake kinetics for sucrose did not appear to be the result of an equilibrium between rates of sucrose influx and efflux.Abbreviations 3-OMG 3-O-methylglucose - PCMBS p-chloromercuribenzene sulfonate - SE-CC sieve element-companion cell - VB vascular bundle     

Source-to-sink gradient of potassium in the phloem

The potassium contents of bark strips of cassava (Manihot esculenta Crantz) and of phloem exudate of castor bean (Ricinus communis L.) were analyzed at different regions of the stem. In cassava, a peak in potassium content was observed near the first mature leaf, leveling off both above and below this point. In castor bean, only a downward decreasing gradient was observed. In both plants, the direction of the potassium gradient coincided with the presumed direction of assimilate flow.     

Microautoradiographic studies of phloem loading and transport in the leaf of Zea mays L.

Microautoradiographs showed that [¹⁴C]sucrose taken up in the xylem of small and intermediate (longitudinal) vascular bundles of Zea mays leaf strips was quickly accumulated by vascular parenchyma cells abutting the vessels. The first sieve tubes to exhibit ¹⁴C-labeling during the [¹⁴C]sucrose experiments were thick-walled sieve tubes contiguous to the more heavily labeled vascular parenchyma cells. (These two cell types typically have numerous plasmodesmatal connections.) With increasing [¹⁴C]sucrose feeding periods, greater proportions of thick- and thin-walled sieve tubes became labeled, but few of the labeled thin-walled sieve tubes were associated with labeled companion cells. (Only the thin-walled sieve tubes are associated with companion cells.) When portions of leaf strips were exposed to ¹⁴CO₂ for 5 min, the vascular parenchyma cells-regardless of their location in relation to the vessels or sieve tubes-were the most consistently labeled cells of small and intermediate bundles, and label (¹⁴C-photosynthate) appeared in a greater proportion of thin-walled sieve tubes than thick-walled sieve tubes. After a 5-min chase with ¹²CO₂, the thin-walled sieve tubes were more heavily labeled than any other cell type of the leaf. After a 10-min chase with ¹²CO₂, the thin-walled sieve tubes were even more heavily labeled. The companion cells generally were less heavily labeled than their associated thin-walled sieve tubes. Although all of the thick-walled sieve tubes were labeled in portions of leaf strips fed ¹⁴CO₂ for 5 min and given a 10-min ¹²CO₂ chase, only five of 72 vascular bundles below the ¹⁴CO₂-exposed portions contained labeled thick-walled sieve tubes. Moreover, the few labeled thick-walledsieve tubes of the transport region always abutted ¹⁴C-labeled vascular parenchyma cells. The results of this study indicate that (1) the vascular parenchyma cells are able to retrieve at least sucrose from the vessels and transfer it to the thick-walled sieve tubes, (2) the thick-walled sieve tubes are not involved in long-distance transport, and (3) the thin-walled sieve tubes are capable themselves of accumulating sucrose and photosynthates from the apoplast, without the companion cells serving as intermediary cells.     

Turnover and transport of quinolizidine alkaloids. Diurnal fluctuations of lupanine in the phloem sap,leaves and fruits of Lupinus albus L.

Quinolizidine alkaloids formed in the leaves of Lupinus albus L. are translocated via the phloem to the other plant organs, especially the maturing fruits. Compared with amino-acid transport in the phloem, the alkaloids contribute about 8% to the overall nitrogen being exported from the leaf. Since it is likely that the alkaloids are subsequently degraded in the target tissues a minor role of quinolizidine alkaloids might be nitrogen transport. A marked diurnal fluctuation of alkaloids was observed in the leaves, the phloem sap, the roots and the fruits with an increase during the day and an amplitude of several hundred percent thus providing evidence for a rapid turnover of endogenous alkaloids.Abbreviations QA quinolizidine alkaloids - GLC gas-liquid chromatography     

Effect of ethylene on flower abscission: a survey

The effect of ethylene on flower abscission was investigated in monocotyledons and eudicotyledons, in about 300 species from 50 families. In all species studied except Cymbidium, flower abscission was highly sensitive to ethylene. Flower fall was not consistent among the species in any family studied. It also showed no relationship with petal senescence or abscission, nor with petal colour changes or flower closure. Results suggest that flower abscission is generally mediated by endogenous ethylene, but that some exceptional ethylene-insensitive abscission occurs in the Orchidaceae.     

Phloem transport in Picea abies (L.) Karst. in mid-winter

Summary Translocation of ¹⁴C assimilates was studied on four different transport systems of Picea abies branches after induced activation in January. ¹⁴CO₂ assimilation of terminal shoots for 48 h at 25° C resulted in phloem loading and basipetal transport of ¹⁴C photosynthate into the following, older shoot generations. ¹⁴C import was enhanced, when these older shoot generations were kept in the dark. Microautoradiographs of the labelled terminal shoots showed that ¹⁴C assimilates were exported from needles via sieve elements of the leaf traces and loaded into the latest increment of the axial secondary phloem. No ¹⁴C label appeared in the obliterated sieve cells or in the tracheids. In addition, ¹⁴C photosynthate accumulated densely in the chlorophyllous cells of the cortex and in cells of the resin ducts, indicating certain sink activity. In the darkened 2-year-old shoot, imported ¹⁴C photosynthate was concentrated in the functional secondary phloem, while some ¹⁴C label was unloaded into the latest xylem increment. When 6-year-old shoots were exposed to ¹⁴CO₂ for 48 h in the light, ¹⁴C assimilates accumulated in the phloem of the leaf trace and in the latest increment of the axial secondary phloem. However, a substantial amount of radioactivity was unloaded into ray cells and phloem parenchyma cells. Thus, the presence of functioning phloem in needles and twigs of P. abies during winter allows long-distance translocation and radial distribution of assimilates according to existing source-sink relations.     

Mineral partitioning in the phloem during autumn senescence of beech leaves

Summary During the period of leaf senescence in fall, the minerals Mg, Ca, K, P, Cl, S, and Si were compared for occurrence and density in tissue compartments of leaf blade, petiole, and subtending stem of beech (Fagus sylvatica L.). Measurements were made by energy-dispersive X-ray microanalysis. The plant material was collected on 2,9, 16 and 23 October, and showed green, greenyellow, yellow, and red-brown autumn leaf coloration. Mg, K, and P were retrieved from the leaf blade prior to shedding, and deposited mainly in cortex and pith tissues of the stem. S and Ca remained in the leaf, and Si and Cl appeared to accumulate in the leaf prior to shedding. During the four stages of leaf senescence, the phloem compartments of the petiole showed considerable changes in mineral content. In addition, leaf senescence in several cases was accompanied by ion shifting from symplastic to apoplastic compartments and vice versa.     

Phloem transport of sulfur in Ricinus

Mature leaves of Ricinus communis fed with ³⁵SO ₄ ^2- in the light export labeled sulfate and reduced sulfur compounds by phloem transport. Only 1–2% of the absorbed radiosulfur is exported to the stem within 2–3 h, roughly 12% of ³⁵S recovered was in reduced form. The composition of phloem translocate moving down the stem toward the root was determined from phloem exudate: 20–40% of the ³⁵S moved in the form of organic sulfur compounds, however, the bulk of sulfur was transported as inorganic sulfate. The most important organic sulfur compound translocated was glutathione, carrying about 70% of the label present in the organic fraction. In addition, methionine and cysteine were involved in phloem sulfur transport and accounted for roughly 10%. Primarily, the reduced forms of both, glutathione and cysteine are prsent in the siever tubes.Abbreviations CySH cysteine - GSH glutathione - GSSG glutathione disulfide - NEM N-ethylmaleimide - CyS-SCy cystine     

Study of sucrose and mannitol transport in plasma-membrane vesicles from phloem and non-phloem tissues of celery (Apium graveolens L.) petioles

The mature petiole of celery is an organ with versatile sink/source capacities where sucrose and mannitol are unloaded from and reloaded into the phloem cells. Plasma-membrane vesicles were purified by twophase partitioning either from phloem strands isolated from mature petioles of celery (Apium graveolens L.) or from mature petioles devoid of vascular bundles. Both types of vesicle were comparable in purity (more than 86% of plasma-membrane origin), size (135 nm diameter) and orientation (72% right-side-out). Plasma-membrane vesicles from phloem tissues had a higher vanadate-sensitive ATPase activity than plasma-membrane vesicles from petioles. Plasma-membrane vesicles from phloem tissues accumulated mannitol and sucrose in response to an artificial proton-motive force, in agreement with the existence of proton/substrate carriers. Plasma-membrane vesicles from petioles devoid of vascular bundles accumulated only mannitol following application of an artificial proton-motive force. The data suggest the volvement of apoplasmic transport events. The pathway for sucrose uptake in storage parenchyma cells is discussed in the light of the available physiological data.     

Wound phloem in transition to bundle phloem in primary roots ofPisum sativum L.

Summary 48 hours after interrupting the root stele ofPisum, wound phloem initiated (proximally or distally to the wound) to reconnect the vascular stumps was found to contain some nucleate wound-sieve elements. At the elongating end of an incomplete wound-sieve tube these elements exhibit a sequence of ultrastructural changes as known from protophloem-sieve tubes. Elongation occurs by the addition of newly divided (wound-) sieve-element/companion-cell complexes. In order to dedifferentiate and assume a new specialization formerly quiescent stelar or cortical cells require at least one (mostly more) preliminary division. Companion cells are consequently obligatory sister cells to wound-sieve elements.By reconstruction using serial sections it could be shown that wound-sieve tubes elongate bidirectionally, starting in an early activated procambial cell of the stele. The elongation is directed by the existence of plasmodesmata, preferably when lying in primary pit fields, and by the plane of preceding divisions. Thus, the developing wound-sieve tube can deviate from the damaged bundle and radiate into the cortex as soon as the plane of the preceding divisions is favourable. In the opposite direction, elongating wound-sieve tubes run parallel to pre-existing phloem traces, thus broading their base at the bundle for the deviating part of the wound-sieve tube. Frequently an individual wound-sieve tube is supplemented at the bundle by a further wound-sieve tube which is partly running parallel to it. Both sieve tubes are interlinked with sieve plates by three-poled sieve elements.Ultrastructurally, the developmental changes of nucleate wound-sieve elements follow the known pattern. In spite of its contrasting origin and odd shape a mature wound-sieve element eventually has the same contents as regular sieve elements: sieve-element plastids, mitochondria, stacked ER and small amounts of P-protein within an electronlucent cytoplasm.     

Long-distance transport of sulfur in Nicotiana tabacum

Sulfur reduction in tobacco plants is a light-enhanced process that predominantly takes place in the leaves rather than the roots. The amount of sulfate reduced in mature leaves can exceed their own requirement and enables an export of reduced sulfur, both basipetal toward the roots as well as acropetal toward the growing parts of the stem. Evidence is presented that translocation of reduced sulfur toward the roots occurs in the phloem. TLC and paper chromatography reveal that glutathione is the main transport form of reduced sulfur in tobacco plants; 67–70% of reduced ³⁵S was confined to glutathione, 27–30% to methionine, and 2–8% to cysteine.Abbreviation TLC thin-layer chromatography     

Transport of hexoses by the phloem of Ricinus communis L. seedlings

The sieve-tube sap of Ricinus communis L. seedlings has been analysed to determine whether or not hexoses can be taken up by the phloem. Under natural conditions, i.e. with the endosperm attached to the cotyledons, glucose and fructose occurred only in trace amounts in the sieve-tube sap. Incubation of the cotyledons with hexoses in the concentration range 25–200 mM caused a rapid and substantial uptake of hexoses into the phleom, where they appeared eventually in the sieve-tube sap at the same concentration as in the incubation medium. Phloem loading of glucose, 3-O-methyl-glucose and sorbitol occurred easily, whereas fructose was less well loaded. glucose and to a larger extent fructose were also transformed to sucrose, which was loaded into the phloem. The loading of hexoses into the sieve tubes as observed in the experimental exudation system also occurred in the intact seedling, but transloction in the latter soon came to a standstill, probably because of lack of consumption by the sink tissues. These results indicate that the virtual absence of hexoses in the sievetube sap under in-vivo conditions is not because of the inability of the phloem-loading system to transport the monosaccharides but because of the absence of sufficiently high concentrations in the apoplast.     

Retention of xenobiotics along the phloem path

Detached leaves of Cyclamen persicum Mill. can be used as a simple source-sink system. Phloem transport in the excised material was monitored by the noninvasive ¹¹C-technique. Assimilate movement stopped immediately when the petiole was cut off. However, within 20 min a recovery of transport was observed. The translocation rate in the detached leaf was only 13% of that in the intact plant. ¹⁴C-Xenobiotics and [³H]sucrose were injected into the upper petiole parenchyma (source). They moved downstream by a symplastic route. The stump of the petiole was inserted into a buffer solution containing ethylenediaminetetraacetic acid (sink). After 3 h, the distribution of sucrose and xenobiotics was determined in five subsequent segments of the petiole (path). The retention coefficient (r) was calculated from the ratio of radioactivity in the vascular bundle to that in the petiole parenchyma. The distribution along the vascular path was given by a geometric progression, whereas its constant was the transport coefficient (q). Values of r and q corresponded with the degree of phloem mobility and ambimobility. Four groups of compounds were classified: (i) acidic substances with log K_ow = — 2 to — 2.4 (K_ow is the partition coefficient octanol/water) at pH 8 (pH of sieve tube sap), retained by ion trapping and exhibiting small lateral efflux (q0.7; maleic hydrazide, dalapon); (ii) acidic substances with log K_ow = — 0.7 to — 0.8 at pH 8, retained by ion trapping and subjected to a moderate lateral efflux (0.7>q> 0.5; 2,4-dichlorophenoxyacetic acid, 2-methyl-4-chlorophenoxyacetic acid, bromoxynil); (iii) nonionised substances retained by optimum permeability, exhibiting a considerable lateral leakage (q<0.5; glyphosate, amitrole); (iv) substances without basipetal transport in the phloem (atrazine, diuron). Retention of sucrose corresponded quantitatively with that shown in group (i). This classification was also supported by results of uptake and efflux experiments using the isolated conducting tissue. Theoretical translocation profiles were calculated from the determined transport coefficients (q).Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - K_ow partition coefficient octanol/water - MCPA 2-methyl-4-chloro-phenoxyacetic acid - q transport coefficient in the vascular bundle - r retention coefficient in the vascular bundle The authors gratefully acknowledge the assistance of H. Fiedler and M. Neugebauer. We are particularly grateful to K. Dutschka, G. Hudepokl, and Dr. J. Knust for producing ¹¹CO₂.