Organic substances in xylem sap delivered to above-ground organs by the roots

Squash (Cucurbita maxima) xylem sap, an apoplastic fluid, contains t-zeatin riboside, glutamine, methylglycine, myo-inositol, fructose, oligosaccharides of arabinogalactan, glucan, galacturonan, and pectins (rhamnogalacturonan-I and rhamnogalacturonan-II), as well as various proteins, including arabinogalactan and pathogen-related proteins. These substances are mainly produced in stele (xylem) parenchyma and the pericycle in the root-hair zone where ion transporter genes are expressed. Glycine-rich protein genes (CRGRPs) cloned by antiserum raised against whole xylem sap of cucumber (Cucumis sativus) were abundantly expressed in the parenchyma cells surrounding xylem vessels in the root-hair zone. CRGRP proteins accumulated and immobilized in the lignified walls of metaxylem vessels and perivascular fibers in shoots, suggesting a systemic delivery mechanism of wall materials via xylem sap. A major 30-kDa protein (XSP30) found in cucumber xylem sap was homologous to the B chains of a lectin (ricin) and bound to a nonfucosylated core N-acetylglucosamine dimer of N-linked glycoproteins abundant in leaf parenchyma cells. XSP30 gene expression, abundant in root xylem parenchyma and pericycle, and the level of XSP30 protein fluctuated diurnally under the control of a circadian clock, and the amplitude was up-regulated by gibberellic acid produced in young leaves, suggesting a long-distance control system between organs.     

The tomato xylem sap protein XSP10 is required for full susceptibility to Fusarium wilt disease

XSP10 is an abundant 10 kDa protein found in the xylem sap of tomato. The protein displays structural similarity to plant lipid transfer proteins (LTPs). LTPs are involved in various physiological processes, including disease resistance, and some are able to bind and transfer diverse lipid molecules. XSP10 abundance in xylem sap declines upon infection with Fusarium oxysporum f. sp. lycopersici (Fol), implying involvement of XSP10 in the plant-pathogen interaction. Here, the biochemical characterization of XSP10 with respect to fatty acid-binding properties is reported; a weak but significant binding to saturated fatty acids was found. Furthermore, XSP10-silenced tomato plants were engineered and it was found that these plants exhibited reduced disease symptom development upon infection with a virulent strain of Fol. Interestingly, the reduced symptoms observed did not correlate with an altered expression profile for known reporter genes of plant defence (PR-1 and WIPI). This work demonstrates that XSP10 has lipid-binding properties and is required for full susceptibility of tomato to Fusarium wilt.     

Possible involvement of leaf gibberellins in the clock-controlled expression of XSP30, a gene encoding a xylem sap lectin, in cucumber roots

Root-produced organic compounds in xylem sap, such as hormones and amino acids, are known to be important in plant development. Recently, biochemical approaches have revealed the identities of several xylem sap proteins, but the biological functions and the regulation of the production of these proteins are not fully understood. XYLEM SAP PROTEIN 30 kD (XSP30), which is specifically expressed in the roots of cucumber (Cucumis sativus), encodes a lectin and is hypothesized as affecting the development of above-ground organs. In this report, we demonstrate that XSP30 gene expression and the level of XSP30 protein fluctuate in a diurnal rhythm in cucumber roots. The rhythmic gene expression continues for at least two or three cycles, even under continuous light or dark conditions, demonstrating that the expression of this gene is controlled by a circadian clock. Removal of mature leaves or treatment of shoots with uniconazole-P, an inhibitor of gibberellic acid (GA) biosynthesis, dampens the amplitude of the rhythmic expression; the application of GA negates these effects. These results suggest that light signals perceived by above-ground organs, as well as GA that is produced, possibly, in mature leaves, are important for the rhythmic expression of XSP30 in roots. This is the first demonstration of the regulation of the expression of a clock-controlled gene by GA.     

A tomato xylem sap protein represents a new family of small cysteine-rich proteins with structural similarity to lipid transfer proteins

The coding sequence of a major xylem sap protein of tomato was identified with the aid of mass spectrometry. The protein, XSP10, represents a novel family of extracellular plant proteins with structural similarity to plant lipid transfer proteins. The XSP10 gene is constitutively expressed in roots and lower stems. The decline of XSP10 protein levels in tomato infected with a fungal vascular pathogen may reflect breakdown or modification by the pathogen.     

Presence of a basic secretory protein in xylem sap and shoots of poplar in winter and its physicochemical activities against winter environmental conditions

Journal of Plant Research - XSP25, previously shown to be the most abundant hydrophilic protein in xylem sap of Populus nigra in winter, belongs to a secretory protein family in which the...     

Identification of the silicon form in xylem sap of rice (Oryza sativa L.)

Rice (Oryza sativa L.) is a typical silicon (Si)-accumulating plant, but the mechanism responsible for the translocation from the root to the shoot is poorly understood. In this study, the form of Si in xylem sap was identified by (29)Si-nuclear magnetic resonance (NMR) spectroscopy. In rice (cv. Oochikara) cultured in a monosilicic acid solution containing 0.5 mM Si, the Si concentration in the xylem reached 6 mM within 30 min. In the (29)Si-NMR spectra of the xylem sap, only one signal was observed at a chemical shift of -72.6 ppm, which is consistent with that of monosilicic acid. A (1)H-NMR study of xylem sap did not show any significant difference between the wild-type rice and mutant rice defective in Si uptake, and the components of the xylem sap were not affected by the Si supply. The Si concentration in the xylem sap in vitro decreased from an initial 18 mM to 2.6 mM with time. Addition of xylem sap to a solution containing 8 mM Si did not prevent the polymerization of silicic acid. All these results indicate that Si is translocated in the form of monosilicic acid through the xylem and that the concentration of monosilicic acid is high in the xylem only transiently.     

Xylem sap protein composition is conserved among different plant species

Xylem sap from broccoli (Brassica oleracea L. cv. Calabrais), rape (Brassica napus L. cv. Drakkar), pumpkin (Cucurbita maxima Duch. cv. gelber Zentner) and cucumber (Cucumis sativus L. cv. Hoffmanns Giganta) was collected by root pressure exudation from the surface of cut stems of healthy, adult plants. Total protein concentrations were in the range of 100 g ml^–1. One-dimensional gel electrophoresis (SDS–PAGE) resulted in 10–20 visible protein bands in a molecular mass range from 10 to 100 kDa. The main bands were cut out, digested with trypsin, and analysed using tandem mass spectrometry. Fifty bands resulted in amino acid sequence information that was used to perform database similarity searches. Sequences from 30 bands showed high homology to proteins present in databases. Among them, we found mostly peroxidases, but could also identify the lectin-like xylem protein XSP30, a glycine-rich protein, serine proteases, an aspartyl protease family protein, chitinases, and a lipid transfer protein-like polypeptide. Sequence analysis predicted apoplastic secretion signals for all database entries similar to the partial xylem protein sequences. This and the lack of cross-reactivity with phloem protein-specific antibodies suggest that the proteins really originate from the xylem and do not result from phloem contamination. Most of the highly similar proteins probably function in repair and defence reactions. Some of the most abundant proteins (peroxidases, chitinases, serine proteases) were present in xylem exudate of all species analysed, often in more than one band. This indicates an important basic role of these proteins in maintaining xylem function.Abbreviations CHT Chitinase - 1D One-dimensional - GRP Glycine-rich protein - SP Serine protease - SSP Subtilisin-like serine protease - POX Peroxidase     

Chemical forms of aluminum in xylem sap of tea plants (Camellia sinensis L.)

To identify the chemical forms of aluminum (Al) transported from roots to shoots of tea plants (C. sinensis L.), ²⁷Al-nuclear magnetic resonance and ¹⁹F NMR spectroscopy were used to analyze xylem sap.The concentration of Al in collected xylem sap was 0.29 mM, twice as high as that of F. Catechins were not detected in xylem sap. The concentration of malic acid in xylem sap was higher than that of citric acid, whereas the concentration of oxalic acid was negligible.There were two signals in the ²⁷Al NMR spectra of xylem sap, a larger signal at 11 ppm and a smaller one at −1.5 ppm. The former signal was consistent with the peak for an Al-citrate model solution, suggesting that an Al-citrate complex was present in xylem sap. Although the latter signal at −1.5 ppm was thought to indicate the presence of an Al-F complex (at 1.7 ppm) in xylem sap, there was only one signal at −122 ppm in the ¹⁹F NMR spectrum of xylem sap, indicating that the main F complex in xylem sap was F⁻.These results indicate that Al might be translocated as a complex with citrate, while Al-malate, Al-oxalate and Al-F complexes are not major Al complexes in xylem sap of tea plants.     

A phloem-sap feeder mixes phloem and xylem sap to regulate osmotic potential

Phloem-sap feeders (Hemiptera) occasionally consume the dilute sap of xylem, a behaviour that has previously been associated with replenishing water balance following dehydration. However, a recent study reported that non-dehydrated aphids ingested xylem sap. Here, we tested the hypothesis that the consumption of xylem sap, which has a low osmolality, is a general response to osmotic stresses other than dehydration. Alate aphids were subjected to different treatments and subsequently transferred onto a plant, where electrical penetration graph (EPG) was used to estimate durations of passive phloem sap consumption and active sucking of xylem sap. The proportion of time aphids fed on xylem sap (i.e., time spent feeding on xylem sap/total time spent feeding on phloem plus xylem sap) was used as a proxy of the solute concentration of the uptake. The proportion of time alate aphids fed on xylem sap increased: (1) with the time spent imbibing an artificial diet containing a solution of sucrose, which is highly concentrated in phloem sap and is mainly responsible for the high osmotic potential of phloem sap; (2) with the osmotic potential of the artificial diet, when osmotic potential excess was not related to sucrose concentration; and (3) when aphids were deprived of primary symbionts, a condition previously shown to lead to a higher haemolymph osmotic potential. All our results converge to support the hypothesis that xylem sap consumption contributes to the regulation of the osmotic potential in phloem-sap feeders.     

Water relations in silver birch during springtime: How is sap pressurised?

• Positive sap pressures are produced in the xylem of birch trees in boreal conditions during the time between the thawing of the soil and bud break. During this period, xylem embolisms accumulated during wintertime are refilled with water. The mechanism for xylem sap pressurization and its environmental drivers are not well known.
• We measured xylem sap flow, xylem sap pressure, xylem sap osmotic concentration, xylem and whole stem diameter changes, and stem and root non‐structural carbohydrate concentrations, along with meteorological conditions at two sites in Finland during and after the sap pressurisation period.
• The diurnal dynamics of xylem sap pressure and sap flow during the sap pressurisation period varied, but were more often opposite to the diurnal pattern after bud burst, i.e. sap pressure increased and sap flow rate mostly decreased when temperature increased. Net conversion of soluble sugars to starch in the stem and roots occurred during the sap pressurisation period. Xylem sap osmotic pressure was small in comparison to total sap pressure, and it did not follow changes in environmental conditions or tree water relations.
• Based on these findings, we suggest that xylem sap pressurisation and embolism refilling occur gradually over a few weeks through water transfer from parenchyma cells to xylem vessels during daytime, and then the parenchyma are refilled mostly during nighttime by water uptake from soil. Possible drivers for water transfer from parenchyma cells to vessels are discussed. Also the functioning of thermal dissipation probes in conditions of changing stem water content is discussed.

     

Control of Sodium Transport in Sunflower Roots

Electrochemical potential differences (driving forces) for sodiumdistributed between the outside solution and the exuding sapof water-culture-grown sunflower plants (Helianthus annuius)have been determined. The results indicated that sodium wasmoving from the outside solution to the xylem against the electrochemicalpotential gradient at external concentrations below approximately0.30 mM Na. At higher external concentrations sodium appearedto be actively excluded from the xylem. An electrical potential difference between the exuding sap andthe external solution of approximately 30 mV was observed. Itwas unaffected by the external sodium concentration. Use ofa short-circuiting technique indicated that the trans-root potentialresides at the plasmalemma of the cortical cells. Driving forces on sodium distributed between the external solutionand the root and between the xylem sap and the root were calculated.They indicated that the root is able to accumulate sodium activelyboth from the external solution and the xylem sap. It is concludedthat sodium transport to the xylem in this species is controlledby the balance of these two opposing forces.     

Xylem sap composition: A tool for investigating mineral uptake and cycling in adult spruce

Xylem sap composition of spruce is influenced by several factors, such as the sampled organ, the sampling period, the availability of soil nutrients, and the soil water potential. Based on literature data and ongoing investigations carried out with adult trees, we present an overview on the main factors influencing xylem sap concentrations of Norway spruce. Direct measurements of nutrient fluxes in the xylem sap are then used to suggest a general scheme of mineral element cycling within adult trees. In Norway spruce (Picea abies Karst.), nutrient concentration in the xylem sap was higher in twigs and fine roots compared to the bottom of the trunk, the highest concentrations beeing observed in spring during the shoot elongation. Xylem sap concentrations were higher in spruce growing at nutrient rich sites than at poor sites. The combination of twig and trunk xylem sap analysis, together with xylem flow measurements in the trunk during the course of a vegetation period allowed the quantification of mineral fluxes via xylem sap flow in the trunk and twigs. These results were compared to gross mineral uptake measurements at the same site. Ca flux in the trunk xylem sap was lower than the gross uptake of Ca. Mg flux in trunk sap was approximately equivalent to Mg gross uptake whereas P and K fluxes in trunk sap were much higher than the gross uptake. Fluxes of Ca, Mg, K and P in the twig sap were much higher than that in trunk sap. Data suggest that internal cycling is responsible for a large part of the nutrient fluxes in the xylem sap of the crown. Xylem sap composition thus appears to be a tool which can complement other sources of information on mineral uptake and cycling in adult spruce     

Squash xylem sap has activities that inhibit proliferation and promote the elongation of tobacco BY-2 cell protoplasts.

To elucidate the physiological functions of the substances in xylem sap, we analyzed the biological activities of xylem sap from squash (Cucurbita maxima Duch.) root using tobacco BY-2 (Nicotiana tabacum L. cv. Bright Yellow 2) cell protoplasts. When BY-2 cell protoplasts were cultivated with the total substance of squash xylem sap, the protoplasts elongated remarkably, and cell division was inhibited. Although trans-zeatin riboside (ZR), the most abundant cytokinin in squash xylem sap, had a concentration-dependent effect similar to that of total squash xylem sap, ZR concentrations several orders of magnitude greater than those found endogenously in squash xylem sap (i.e. 2 x 10(-8) M) were required to affect the growth of BY-2 cell protoplasts. The ability to stimulate cell elongation and inhibit cell division in BY-2 cell protoplasts was observed for the ethyl acetate phase fraction (pH 2) of squash xylem sap and an acetonitrile-eluate fraction from reverse-phase chromatography. The xylem sap also showed inhibitory activity for auxin-induced elongation of excised cucumber hypocotyls. These results suggest that an organic substance other than ZR is produced in the root and transported to above-ground organs through the xylem via the transpiration stream, where it is involved in regulating cell proliferation and elongation in the shoot, possibly as an auxin antagonist.     

Sap Concentrations in Halophytes and Some Other Plants

Freezing point depression in xylem sap of mangroves was found to range from 0.05 to 0.5 degrees , in desert plants from 0.01 to 0.16 degrees . In crush juices from leaves of Batis and Salicornia, 90% or more of the freezing point depression was made up of sodium and chlorine ions; in mangroves they constituted 50 to 70%, the rest probably being organic solutes. Plants growing in seawater have -30 to -60 atmospheres pressure in the xylem sap. As shown earlier, at zero turgor pressure the intracellular freezing point of the parenchyma cells matches closely the negative pressure in the xylem sap. This agrees with the present data, that the fluid which exudes from the xylem by applying gas pressure on the leaves is practically pure water; freezing point is rarely above 0.01 to 0.02 degrees . To perform this ultrafiltration, the plasma membrane is subjected to a hydrostatic pressure gradient which in some cases may exceed 100 atmospheres.     

Stomatal conductance and xylem sap properties of aspen (Populus tremuloides) in response to low soil temperature

The mechanisms regulating stomatal response following exposure to low (5°C) soil temperature were investigated in aspen ( Populus tremuloides Michx.) seedlings. Low soil temperature reduced stomatal conductance within 4 h, but did not alter shoot xylem pressure potential within 24 h. The xylem sap composition was altered and its pH increased from 6.5 to 7.1 within the initial 4 h of the low temperature treatment. However, the increase in abscisic acid (ABA) concentration in xylem sap was observed later, after 8 h of treatment. These changes were accompanied by a reduction in the electrical conductivity and an increase in the osmotic potential of the xylem sap. The timing of physiological responses to low soil temperature suggests that the rapid pH change of the xylem sap and accompanying changes in ion concentration were the initial factors which triggered stomatal closure in low temperature-treated seedlings, and that the role of the more slowly accumulating ABA was likely to reinforce the stomatal closure. When leaf discs were exposed to xylem sap extracted from low soil temperature-treated plants, stomatal aperture was negatively correlated with ABA and positively correlated with K⁺ concentrations of the xylem sap. The stomatal opening in the leaf discs linearly increased in response to exogenous KCl concentrations when K⁺ concentrations were in the similar range to those detected in the xylem sap. The lowest concentration of exogenous ABA to induce stomatal closure was several-fold higher compared with the concentration present in the xylem sap.     

Terpenoids are transported in the xylem sap of Norway spruce

Norway spruce is a conifer storing large amounts of terpenoids in resin ducts of various tissues. Parts of the terpenoids stored in needles can be emitted together with de novo synthesized terpenoids. Since previous studies provided hints on xylem transported terpenoids as a third emission source, we tested if terpenoids are transported in xylem sap of Norway spruce. We further aimed at understanding if they might contribute to terpenoid emission from needles. We determined terpenoid content and composition in xylem sap, needles, bark, wood and roots of field grown trees, as well as terpenoid emissions from needles. We found considerable amounts of terpenoids—mainly oxygenated compounds—in xylem sap. The terpenoid concentration in xylem sap was relatively low compared with the content in other tissues, where terpenoids are stored in resin ducts. Importantly, the terpenoid composition in the xylem sap greatly differed from the composition in wood, bark or roots, suggesting that an internal transport of terpenoids takes place at the sites of xylem loading. Four terpenoids were identified in xylem sap and emissions, but not within needle tissue, suggesting that these compounds are likely derived from xylem sap. Our work gives hints that plant internal transport of terpenoids exists within conifers; studies on their functions should be a focus of future research.     

Characterization of the maize xylem sap proteome

The xylem in plants has mainly been described as a conduit for water and minerals, but emerging evidence also indicates that the xylem contains protein. To study the proteins in xylem sap, we characterized the identity and composition of the maize xylem sap proteome. The composition of the xylem sap proteome in maize revealed proteins related to different phases of xylem differentiation including cell wall metabolism, secondary cell wall synthesis, and programmed cell death. Many proteins were found to be present as multiple isoforms and some of these isoforms are glycosylated. Proteins involved in defense mechanisms were also present in xylem sap and the sap proteins were shown to have antifungal activity in bioassays.     

Effects of iron deficiency on the composition of the leaf apoplastic fluid and xylem sap in sugar beet. Implications for iron and carbon transport

The effects of iron deficiency on the composition of the xylem sap and leaf apoplastic fluid have been characterized in sugar beet (Beta vulgaris Monohil hybrid). pH was estimated from direct measurements in apoplastic fluid and xylem sap obtained by centrifugation and by fluorescence of leaves incubated with 5-carboxyfluorescein and fluorescein isothiocyanate-dextran. Iron deficiency caused a slight decrease in the pH of the leaf apoplast (from 6.3 down to 5.9) and xylem sap (from 6.0 down to 5.7) of sugar beet. Major organic acids found in leaf apoplastic fluid and xylem sap were malate and citrate. Total organic acid concentration in control plants was 4.3 mM in apoplastic fluid and 9.4 mM in xylem sap and increased to 12.2 and 50.4 mM, respectively, in iron-deficient plants. Inorganic cation and anion concentrations also changed with iron deficiency both in apoplastic fluid and xylem sap. Iron decreased with iron deficiency from 5.5 to 2.5 microM in apoplastic fluid and xylem sap. Major predicted iron species in both compartments were [FeCitOH](-1) in the controls and [FeCit(2)](-3) in the iron-deficient plants. Data suggest the existence of an influx of organic acids from the roots to the leaves via xylem, probably associated to an anaplerotic carbon dioxide fixation by roots.     

Role of xylem consumption on osmoregulation in Macrosiphum euphorbiae (Thomas)

Aphids are phloem feeders that occasionally ingest xylem sap. The duration of xylem consumption by Macrosiphum euphorbiae (Hemiptera: Aphididae) was positively correlated with the level of dehydration of alate aphids of different ages after a period of starvation, supporting the hypothesis that aphids ingest xylem sap to replenish their water balance. However, the duration of xylem sap ingestion but not phloem sap consumption varied in unstarved alate adults of different ages. Furthermore, both alate and apterous aphids ingested xylem sap at the end of their life, when aphids were not dehydrated but when fecundity started to decrease. Fecundity was negatively correlated with the proportion of time spent ingesting xylem sap, and that over the entire reproductive life of alate and apterous aphids. The lower proportion of xylem ingested by apterous than by alate aphids during the first few days of adult life may be related to a higher symbiont density in apterous morphs. As previous studies have demonstrated a relationship between sucrose assimilation, which is directly influenced by fecundity and symbiont density, and osmoregulation, we suggest that xylem consumption may play a role in the osmoregulation of haemolymph of aphids.     

转Bt 基因抗虫棉(Gossyposium)伤流中Bt 毒蛋白的运输

本文以常规棉品种‘石远321’和抗虫棉品种‘SGK321’与‘99B’为材料, 通过考马斯亮蓝和ELISA方法分析了伤流中可溶性蛋白和Bt(苏云金芽孢杆菌)毒蛋白含量。结果发现, 转基因抗虫棉伤流中可溶性蛋白含量显著高于常规棉, 并在抗虫棉伤流中检测到了Bt毒蛋白, 而常规棉伤流中没有Bt毒蛋白的存在。通过嫁接实验研究发现, 将常规棉嫁接到转Bt基因抗虫棉砧木上, 叶片中也有Bt毒蛋白的积累。这些结果说明, Bt毒蛋白可以通过木质部伤流液向地上部运输, 并在叶片中积累, Bt毒蛋白由根系向地上部的运输对棉花的抗虫性是有益的。