Monitoring Phloem Unloading and Post-Phloem Transport by Microperfusion of Attached Wheat Grains

Phloem unloading and post-phloem transport in developing wheat (Triticum aestivum L.) grains were investigated by perfusing the endosperm cavities of attached grains. Relative unloading ratio (RUR) and the rate of sucrose release into the endosperm cavity (SRR) were calculated, respectively, from 14C import and from sucrose washout from the cavity. RUR and SRR continued at or near in vivo rates over a wide range of cavity sap osmolality (90 to approximately 500 milliosmolal) and sucrose concentration (14-430 mM) and for long times (29 h). These are much greater ranges than have been observed for the endosperm cavity in vivo (230-300 milliosmolal, and 40-120 mM, respectively), indicating that neither the cavity sap osmolality nor sucrose concentration are controlling factors for the rate of assimilate import into the cavity. The maintenance of in vivo transport rates over a wide range of conditions strongly implicates the role of transport processes within the maternal tissues of the wheat grain, rather than activities of the embryo or endosperm, in determining the rate of assimilate import into the grain. RUR was decreased by high concentrations of sucrose and sorbitol, but not of mannitol. By plasmolyzing some chalazal cells, sorbitol appeared to block symplastic transport across the crease tissues, but neither sucrose nor mannitol caused plasmolysis in maternal tissues of attached grains. The inhibition of RUR by KCN and carbonyl cyanide m-chlorophenyl (CCCP) and the continued import of sucrose into grains against its concentration gradient suggest that solute movement into the endosperm cavity might occur by active membrane transport. However, the evidence is weak, since KCN and CCCP appeared to act primarily on some aspect of symplastic (i.e. nonmembrane) transport. Also, sucrose could move from the endosperm cavity into the maternal tissues (i.e. opposite to the normal direction of sucrose movement), suggesting that transmembrane movement in the nucellus may be a reversible process. Pressure-driven flow into the grain could account for movement against a concentration gradient.     

The Use of Fluorescent Tracers to Characterize the Post-Phloem Transport Pathway in Maternal Tissues of Developing Wheat Grains

Various polar fluorescent tracers were used to characterize the pathways for apoplastic and symplastic transport in the "crease tissues" (i.e. the vascular strand, chalaza, nucellus, and adjacent pericarp) of developing wheat (Triticum aestivum L.) grains. With mostly minor exceptions, the results strongly support existing views of phloem unloading and post-phloem transport pathways in the crease. Apoplastic movement of Lucifer yellow CH (LYCH) from the endosperm cavity into the crease was virtually blocked in the chalazal cell walls before reaching the vascular tissue. However, LYCH could move slowly along the cell wall pathway from the chalaza into the vascular parenchyma. Slow uptake of LYCH into nucellar cell cytoplasm was observed, but no subsequent symplastic movement occurred. Carboxyfluorescein (CF) imported into attached grains moved symplastically from the phloem across the chalaza and into the nucellus, but was not released from the nucellus. In addition, CF moved in the opposite direction (nucellus to vascular parenchyma) in attached grains. Thus, the post-phloem symplastic pathway can accommodate bidirectional transport even when there is an intense net assimilate flux in one direction. When fresh sections of the crease were placed in fluorochrome solutions (e.g. LYCH or pyrene trisulfonate), dye was rapidly absorbed into intact cells, apparently via unsealed plasmodesmata. Uptake was not visibly reduced by cold or by respiratory inhibitors, but was greatly reduced by plasmolysis. Once absorbed, the dye moved intercellularly via the symplast. Based on this finding, a size-graded series of fluorescein-labeled dextrans was used to estimate the size-exclusion limits (SEL) for the post-phloem symplastic pathway. In most, and perhaps all, cells of the crease tissues except for the pericarp, the molecular diameter for the SEL was about 6.2 nm. The SEL in much of the vascular parenchyma may be smaller, but it is still at least 3.6 nm. Channel diameters would likely be about 1 nm larger, or about 4.5 to 7.0 nm in the vascular parenchyma and 7.0 nm elsewhere. These dimensions are substantially larger than those for "conventional" symplastic connections (about 3 nm), and would have a greater than proportionate effect on the per channel diffusive and hydraulic conductivities of the pathway. Thus, relatively small and probably ultrastructurally undetectable adjustments in plasmodesmatal structure may be sufficient to account for assimilate flux through the crease symplast.     

Accumulation and Conversion of Sugars by Developing Wheat Grains : VI. Gradients Along the Transport Pathway from the Peduncle to the Endosperm Cavity during Grain Filling

Gradients along the transport pathway from the peduncle to the endosperm cavity were examined during grain filling in wheat. Sieve tube exudate was collected from severed aphid stylets established on the peduncle and rachis and on the vascular bundles in the creases of grains. Phloem exudate could also be collected from broken grain pedicels, and by puncturing the vascular bundle in the grain crease with a needle. Stylets on excised grains persisted exuding, indicating that grain sieve tubes are capable of loading solutes. There was little, if any, discernible gradient in osmolality or solute composition (sucrose, total amino acids) of sieve tube contents along the phloem pathway from the peduncle to the rachis or along the rachis itself. Neither was a gradient detected in osmolality along the sieve tube pathway from the rachis through the rachilla and grain stalk to the crease. Demonstrable solute gradients occurred only across those tissues of the grain crease between the crease sieve tubes and the endosperm cavity, a distance of just 1 millimeter. However, while the sucrose concentration in the sieve tubes was almost tenfold that in the endosperm cavity sap, total amino acids were only threefold higher, and the potassium concentrations of the two were equal. Our observations strongly implicate the movement of assimilates from the sieve tubes and across the crease tissues as important control points in grain filling.     

The cellular pathway of photosynthate transfer in the developing wheat grain. II. A structural analysis and histochemical studies of the pathway from the crease phloem to the endosperm cavity

In the developing wheat grain, photosynthate is transferred longitudinally along the crease phloem and then laterally into the endosperm cavity through the crease vascular parenchyma, pigment strand and nucellar projection. In order to clarify this cellular pathway of photosynthate unloading, and hence the controlling mechanism of grain filling, the potential for symplastic and apoplastic transfer was examined through structural and histochemical studies on these tissue types. It was found that cells in the crease region from the phloem to the nucellar projection are interconnected by numerous plasmodesmata and have dense cytoplasm with abundant mitochondria. Histochemical studies confirmed that, at the stage of grain development studied, an apoplastic barrier exists in the cell walls of the pigment strand. This barrier is composed of lignin, phenolics and suberin. The potential capacity for symplastic transfer, determined by measuring plasmodesmatal frequencies and computing potential sucrose fluxes through these plasmodesmata, indicated that there is sufficient plasmodesmatal cross-sectional area to support symplastic unloading of photosynthate at the rate required for normal grain growth. The potential capacity for membrane transport of sucrose to the apoplast was assessed by measuring plasma membrane surface areas of the various cell types and computing potential plasma membrane fluxes of sucrose. These fluxes indicated that the combined plasma membrane surface areas of the sieve element–companion cell (se–cc) complexes, vascular parenchyma and pigment strand are not sufficient to allow sucrose transfer to the apoplast at the observed rates. In contrast, the wall ingrowths of the transfer cells in the nucellar projection amplify the membrane surface area up to 22-fold, supporting the observed rates of sucrose transfer into the endosperm cavity. We conclude that photosynthate moves via the symplast from the se–cc complexes to the nucellar projection transfer cells, from where it is transferred across the plasma membrane into the endosperm cavity. The apoplastic barrier in the pigment strand is considered to restrict solute movement to the symplast and block apoplastic solute exchange between maternal and embryonic tissues. The implications of this cellular pathway in relation to the control of photosynthate transfer in the developing grain are discussed.     

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.     

The cellular pathway of photosynthate transfer in the developing wheat grain. III. A structural analysis and physiological studies of the pathway from the endosperm cavity to the starchy endosperm

The cellular pathway of sucrose transfer from the endosperm cavity to the starchy endosperm of developing grains of wheat (Triticum turgidum) has been elucidated. The modified aleurone and sub-aleurone cells exhibit a dense cytoplasm enriched in mitochondria and endoplasmic relicilium. Significantly, the sub-aleurone cells are characterized by secondary wall ingrowths. Numerous plasmodesmata interconnect all cells between the modified aleurone and starchy endosperm. The pro-tonophore carbonylcyanide-m-chlorophenyl hydrazone (CCCP) slowed [¹⁴C]sucrose uptake by grain tissue slices enriched in modified aleurone and sub-aleurone cells but had no effect on uptake by the starchy endosperm. The fluorescent weak acid sulphorhodamine G (SRG) was preferentially accumulated by the modified aleurone and sub-aleurone cells, and this uptake was sensitive to CCCP. The combined plasma membrane surface areas of the modified aleurone and sub-aleurone cells appeared to be sufficient to support the in vivo rates of sucrose transfer to the starchy endosperm. Plasmolysis of intact excised grain inhibited [¹⁴C]sucrose transfer from the endosperm cavity to the starchy endosperm. The sulphydryl group modifier p-chloromercuribenzenesulphonie acid (PCMBS) decreased [¹⁴C]sucrose uptake by the modified aleurone and sub-aleurone cells but had little effect on uptake by the starchy endosperm. In contrast, when PCMBS and [¹⁴C]sucrose were supplied to the endosperm cavity of intact excised grain, PCMBS slowed accumulation by all tissues equally. Estimates of potential sucrose fluxes through the interconnecting plasmodesmata were found to be within the published range. It is concluded that the bulk of sucrose is accumulated from the endosperm cavity by the modified aleurone and sub-aleurone cells and subsequently transferred through the symplast to the starchy endosperm.     

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     

Sucrose Release into the Endosperm Cavity of Wheat Grains Apparently Occurs by Facilitated Diffusion across the Nucellar Cell Membranes

Nutrients required for the growth of the embryo and endosperm of developing wheat (Triticum aestivum L.) grains are released into the endosperm cavity from the maternal tissues across the nucellar cell plasma membranes. We followed the uptake and efflux of sugars into and out of the nucellus by slicing grains longitudinally through the endosperm cavity to expose the nucellar surface to experimental solutions. Sucrose uptake and efflux are passive processes. Neither was sensitive to metabolic inhibitors, pH, or potassium concentration. p-Chloromercuribenzene sulfonate, however, strongly inhibited both uptake and efflux, although not equally. Except for p-chloromercuribenzene sensitivity, these characteristics of efflux and the insensitivity of Suc movement to turgor pressure are similar to those of sucrose release from maize pedicels, but they contrast with legume seed coats. Although the evidence is incomplete, movement appears to be carrier mediated rather than channel mediated. In vitro rates of sucrose efflux were similar to or somewhat less than in vivo rates, suggesting that transport across the nucellar cell membranes could be a factor in the control of assimilate import into the grain.     

Photoassimilate translocation in the petiole of Cyclamen and Primula is independent of lateral retrieval

Phloem translocation of photoassimilates between source andsink is considered to be linked with active retrieval of sugarsleaked to the vascular apoplast. This hypothesis was evaluatedby studying photo-assimilate movement in petioles of intactplants of Cyclamen persicum and Primula obconica in the presenceof inhibitors affecting sucrose retrieval (PCMBS, CCCP). Inhibitorsolutions were applied by rinsing locally isolated petiole bundlesor by injection into the petioe parenchyma. PCMBS and CCCP reduced[¹⁴C]sucrose retrieval from the petiole apoplast by the vascularcells and altered the distribution pattern of ¹⁴C-photoassimilateswithin the petiole tissues. However, these treatments did notaffect translocation through the petiole phloem. Evidence isprovided that the reagents were present in the vascular apoplastsurrounding the translocating phloem. It was concluded thatassimilate movement in the petiole of Cyclamen and Primula wasindependent of apoplastic retrieval. Key words: Cyclamen, Primula, phloem, transport, path, sucrose, retrieval     

Gradients in water potential and turgor pressure along the translocation pathway during grain filling in normally watered and water-stressed wheat plants

The water relations parameters involved in assimilate flow into developing wheat (Triticum aestivum L.) grains were measured at several points from the flag leaf to the endosperm cavity in normally watered (Psi approximately -0.3 MPa) and water-stressed plants (Psi approximately -2 MPa). These included direct measurement of sieve tube turgor and several independent approaches to the measurement or calculation of water potentials in the peduncle, grain pericarp, and endosperm cavity. Sieve tube turgor measurements, osmotic concentrations, and Psi measurements using dextran microdrops showed good internal consistency (i.e. Psi = Psi(s) + Psi(p)) from 0 to -4 MPa. In normally watered plants, crease pericarp Psi and sieve tube turgor were almost 1 MPa lower than in the peduncle. This suggests a high hydraulic resistance in the sieve tubes connecting the two. However, observations concerning exudation rates indicated a low resistance. In water-stressed plants, peduncle Psi and crease pericarp Psi were similar. In both treatments, there was a variable, approximately 1-MPa drop in turgor pressure between the grain sieve tubes and vascular parenchyma cells. There was little between-treatment difference in endosperm cavity sucrose or osmotic concentrations or in the crease pericarp sucrose pool size. Our results re-emphasize the importance of the sieve tube unloading step in the control of assimilate import.     

Phloem transport and differential unloading in pea seedlings after source and sink manipulations

Phloem transport was investigated in pea seedlings after application of [¹⁴C]sucrose to the cotyledons. The accumulation of the label in segments of young seedlings shows a differential unloading along the plant axis. Shoot and root exhibit tip-to-base gradients of sink strength. In the primary root, the sink-strength profiles reflect not only the importance of the apical meristem, but show also the starting points of cambial activity and production of secondary vascular elements. Experiments including partial removal of the source and manipulations of the sink strength indicate that translocation of pea seedlings is sink-regulated and responds rapidly to changed apoplastic conditions in the apical root region. Here, a lowered water potential leads to an increase of phloem unloading that is suggested to supply the assimilate demand for the short-term osmoregulation of affected cells via the symplasmic pathway.Abbreviation PCMBS parachloromercuribenzenesulfonic acid Discussions with Prof. R. Kollmann, Botanisches Institut, Universität Kiel, and financial support from the Deutsche Forschungsgemeinschaft are gratefully acknowledged.     

Sieve element unloading: cellular pathway, mechanism and control

The transport and distribution of phloem – mobile solutes is predominantly determined by transport processes located at the sink end of the source – transport – sink system. Transport across the sieve element boundary, sieve element unloading, is the first of a series of sink transport processes. Unloading of solutes from the sieve elements may follow an apo- or symplastic route. It is speculated that the unloading pathway is integrated with sink function and that apoplastic unloading is restricted to situations in which movement through the symplast is not compatible with sink function. These situations include axial transport and storage of osmotically active solutes against concentration and turgor gradients between the sieve elements and sink cells. Coupled with alteration in sink function, the cellular pathway of unloading can switch in stems and possibly other sinks. Experimental systems and approaches used to elucidate the mechanism of sieve element unloading are reviewed. Unloading fluxes to the apoplast can largely be accounted for by membrane diffusion in axial sinks. However, the higher fluxes in storage sinks suggests dependence on some form of facilitated transport. Proton sucrose symport is assessed to be a possible mechanism for facilitated efflux of solutes across the sieve element plasma membrane to the sink apoplast. Unloading through the symplast may occur by diffusion or mass flow. The latter mechanism serves to dissipate phloem water and hence prevent the potential elevation of sieve element turgor that would otherwise slow phloem import into the sink. The possibility of energised plasmodesmatal transport is raised. Sieve element unloading must be integrated with subsequent compartmentation and metabolism of the unloaded solute. Solute levels are an obvious basis for control of sieve element unloading, but are found to offer limited scope for a mass action mechanism. Apoplastic, cellular pathway, sieve element, solute transport, symplastic. Translated into a turgor signal, solute levels could regulate the rate of unloading, metabolism and compartmentation forming part of a turgor homeostat irrespective of the pathway of unloading.     

Characterization of the uptake of sucrose and glucose by isolated seed coat halves of developing pea seeds. Evidence that a sugar facilitator with diffusional kinetics is involved in seed coat unloading

Uptake of ¹⁴C-labelled sucrose and glucose by isolated seed coat halves of pea (Pisum sativum L. cv. Marzia) seeds was measured in the concentration range <0.1 μM to 100 mM. The initial influx of sucrose was strictly proportional to the external concentration, with a coefficient of proportionality (k) of 6.2 μmol·(g FW)^?1·min^?1·M^?1. Sucrose influx was not affected by 10 μM carbonylcyanide m-chlorophenylhydrazone (CCCP), but it was inhibited by 40% in the presence of 2.5 mM p-chloromercuribenzenesulfonic acid (PCMBS). Influx with diffusional kinetics was also observed for glucose (k = 4.8 μmol·(g FW)^?1·min ^?1·M ^?1) and mannitol (k = 5.1 μmol·(g FW)^?1·min^?1·M^?1). For glucose an additional saturable system was found (K_m = 0.26 mM, V _max = 4.2 nmol·(g FW)^?1·min^?1), which appeared to be completely inhibited by CCCP and partly by PCMBS. In contrast to the diffusional pathway, uptake by this saturable system was slightly pH-dependent, with an optimum at pH 5.5. The influx of sucrose appears to be by the same pathway as the efflux of endogenous sucrose, which was inhibited by 36% in the presence of 2.5 mM PCMBS (De Jong A, Wolswinkel P, 1995, Physiol Plant 94: 78–86). It is argued that passive transport may be the only mechanism for sucrose transport through the plasma membrane of seed coat parenchyma cells. The estimated permeability coefficient of the plasma membrane for sucrose (P = 3.5·10^?7 cm·s^?1) is more than 1 × 10⁶-fold higher than that reported for artificial lipid membranes. This relatively high permeability is hypothesized to result from pore-forming proteins that allow the diffusion of sucrose. Furthermore, it is shown that a sucrose gradient across the plasma membrane of the seed coat parenchyma of only 22 mM will suffice to result in the net efflux of sucrose which is required to feed the embryo.     

Sugar Efflux from Maize (Zea mays L.) Pedicel Tissue

Sugar release from the pedicel tissue of maize (Zea mays L.) kernels was studied by removing the distal portion of the kernel and the lower endosperm, followed by replacement of the endosperm with an agar solute trap. Sugars were unloaded into the apoplast of the pedicel and accumulated in the agar trap while the ear remained attached to the maize plant. The kinetics of ¹⁴C-assimilate movement into treated versus intact kernels were comparable. The rate of unloading declined with time, but sugar efflux from the pedicel continued for at least 6 hours and in most experiments the unloading rates approximated those necessary to support normal kernel growth rates. The unloading process was challenged with a variety of buffers, inhibitors, and solutes in order to characterize sugar unloading from this tissue.

Unloading was not affected by apoplastic pH or a variety of metabolic inhibitors. Although p-chloromercuribenzene sulfonic acid (PCMBS), a nonpenetrating sulfhydryl group reagent, did not affect sugar unloading, it effectively inhibited extracellular acid invertase. When the pedicel cups were pretreated with PCMBS, at least 60% of sugars unloaded from the pedicel could be identified as sucrose. Unloading was inhibited up to 70% by 10 millimolar CaCl₂. Unloading was stimulated by 15 millimolar ethyleneglycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid which partially reversed the inhibitory effects of Ca²⁺. Based on these results, we suggest that passive efflux of sucrose occurs from the maize pedicel symplast followed by extracellular hydrolysis to hexoses.

     

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.     

灵武长枣果实维管束韧皮部及周围细胞的超微结构特征

为了探讨灵武长枣果实光合同化物韧皮部卸载和运输的途径,该研究采用透射电镜技术,对不同发育时期灵武长枣果实维管束韧皮部及其周围薄壁细胞的超微结构特征进行了分析.结果表明:筛管/伴胞复合体及其周围韧皮薄壁细胞间在果实膨大前期富含胞间连丝,而韧皮薄壁细胞与周围库细胞以及相邻库细胞间几乎不存在胞间连丝,形成共质体隔离;筛管/伴...     

Transporter SlSWEET15 unloads sucrose from phloem and seed coat for fruit and seed development in tomato

Tomato (Solanum lycopersium), an important fruit crop worldwide, requires efficient sugar allocation for fruit development. However, molecular mechanisms for sugar import to fruits remain poorly understood. Expression of sugars will eventually be exported transporters (SWEETs) proteins is closely linked to high fructose/glucose ratios in tomato fruits and may be involved in sugar allocation. Here, we discovered that SlSWEET15 is highly expressed in developing fruits compared to vegetative organs. In situ hybridization and β-glucuronidase fusion analyses revealed SlSWEET15 proteins accumulate in vascular tissues and seed coats, major sites of sucrose unloading in fruits. Localizing SlSWEET15-green fluorescent protein to the plasma membrane supported its putative role in apoplasmic sucrose unloading. The sucrose transport activity of SlSWEET15 was confirmed by complementary growth assays in a yeast (Saccharomyces cerevisiae) mutant. Elimination of SlSWEET15 function by clustered regularly interspaced short palindromic repeats (CRISPRs)/CRISPR-associated protein gene editing significantly decreased average sizes and weights of fruits, with severe defects in seed filling and embryo development. Altogether, our studies suggest a role of SlSWEET15 in mediating sucrose efflux from the releasing phloem cells to the fruit apoplasm and subsequent import into storage parenchyma cells during fruit development. Furthermore, SlSWEET15-mediated sucrose efflux is likely required for sucrose unloading from the seed coat to the developing embryo.

SlSWEET15, a specific sucrose uniporter in tomato, mediates apoplasmic sucrose unloading from phloem cells and seed coat to support fruit expansion and seed filling.     

菜豆茎韧皮部卸出的细胞途径以及激素的促进作用(英文)

通过缩小叶面积和去茎尖改变源库比率,以调节韧皮部卸出的途径,证明了韧皮部卸出的共质体与质外体途径的季节变化,和由对氯高汞苯磺酸所诱发的从质外体向共质体途径的转变,是与光合产物的输入有关。缩小叶面积而降低源库比率,能增加夏季生长植株茎韧皮部的质外体卸出,但对冬季生长植株无影响。去尖而增加源库比率,则促进共质体卸出。赤霉酸和激动素能促进共质体的横向转运,但对质外体转运无作用。当质外体为主要运输途径时,赤霉酸和激动素开启共质体途径。赤霉酸和激动素刺激光合产物,通过共质体从筛管一伴胞复合体向韧皮部薄壁纽胞输送,并可能在韧皮部薄壁细胞被动扩散到自由空间。由此可进一步说明蔗糖在激素处理部位自由空间的增加。     

Phloem unloading follows an extensive apoplasmic pathway in cucumber (Cucumis sativus L.) fruit from anthesis to marketable maturing stage

The phloem unloading pathway remains unclear in fruits of Cucurbitaceae, a classical stachyose-transporting species with bicollateral phloem. Using a combination of electron microscopy, transport of phloem-mobile symplasmic tracer carboxyfluorescein, assays of acid invertase and sucrose transporter, and [(14)C]sugar uptake, the phloem unloading pathway was studied in cucumber (Cucumis sativus) fruit from anthesis to the marketable maturing stage. Structural investigations showed that the sieve element-companion cell (SE-CC) complex of the vascular bundles feeding fruit flesh is apparently symplasmically restricted. Imaging of carboxyfluorescein unloading showed that the dye remained confined to the phloem strands of the vascular bundles in the whole fruit throughout the stages examined. A 37 kDa acid invertase was located predominantly in the cell walls of SE-CC complexes and parenchyma cells. Studies of [(14)C]sugar uptake suggested that energy-driven transporters may be functional in sugar trans-membrane transport within symplasmically restricted SE-CC complex, which was further confirmed by the existence of a functional plasma membrane sucrose transporter (CsSUT4) in cucumber fruit. These data provide a clear evidence for an apoplasmic phloem unloading pathway in cucumber fruit. A presumption that putative raffinose or stachyose transporters may be involved in soluble sugars unloading was discussed.     

Photoassimilate-transport characteristics of nonchlorophyllous and green tissue in variegated leaves of Coleus blumei Benth

The nonchlorophyllous (albino) tissue of mature C. blumei leaves is a sink for photoassimilate. Transport from the green to the albino region of the same leaf was inhibited by cold and anoxia. When the green tissue of mature leaves was removed, the remaining albino portion imported labeled translocate from other mature leaves in the phloem. Photoassimilate unloading in the albino region of mature leaves was studied by quantitative autoradiography. The unloading was inhibited by cold but not by anoxia. No labeled photoassimilate could be detected in the free space of mature albino tissue by compartmental efflux analysis as phloem unloading proceeded in a N₂ atmosphere, indicating that unloading, may occur by a symplastic pathway as it apparently does in sink leaves of other species. The minor veins of mature albino leaf tissue did not accumulate exogenous [¹⁴C]sucrose. Minor veins of green tissue in the same leaves accumulated [¹⁴C]sucrose but, in contrast to other species studied to date, this accumulation was insensitive to the inhibitor p-chloromercuribenzensulfonic acid (PCMBS).In its capacity to import and unload photoassimilate, and in the inability, of the minor veins to accumulate exogenous sucrose, the albino region of the mature C. blumei lamina differs from mature albino tobacco leaves and darkened mature leaves of other species. This, together with evidence indicating that phloem loading in C. blumei and other species may occur by different routes and with different sensitivity to PCMBS, indicates that the mechanism of transfer of photoassimilates between veins and surrounding tissues, and the mechanism of the sink-source transition, may not be the same in the leaves of all species. It is speculated that the unusual properties of the C. blumei leaf may be a consequence of the presence, in the minor veins, of intermediary cells, large companion cells connected to the bundle sheath by abundant plasmodesmata.Abbreviation PCMBS p-chloromercuribenzenesulfonic acid