Cell-wall polysaccharides in growing poplar bark tissue

In order to study changes in the cell-wall composition of growing poplar cambium during the seasonal cycle, cell walls were isolated from the cambium and newly formed vascular tissues of poplar branches at three times during the year (winter, beginning of spring, end of summer). Polysaccharide material was isolated by sequential extraction of the cell walls and analysed. The principal polysaccharides identified were pectins and xylose- and glucose-containing polysaccharides, possibly xylans and (xylo)glucans. Our results indicate changes in the relative quantities of these polysaccharides during the seasonal cycle.     

Cell wall polysaccharides in black currants and bilberries—characterisation in berries, juice, and press cake

Cell wall polysaccharides from black currants and bilberries were characterised in three approaches. First, compositions of skin, pulp, and seeds show the distribution of polysaccharides over these tissues. A sequential extraction of cell wall material with different aqueous extractants informs about the extractability of the different polysaccharides, viz. pectins, hemicellulose, and cellulose. Finally, by isolation of cell wall polysaccharides from juice and press cakes obtained by the conventional juice manufacturing. The polysaccharide distribution was followed during juice processing. The main difference between bilberries and black currants is the dominant sugar residue in seeds: mannose for black currants and xylose for bilberries. Most of the hemicellulolytic sugars and cellulose can be found back in the press cake. The sugar composition of the press cake is similar to the composition of the residue after sequential extraction. Black currants contain more pectic sugars than bilberries. Consequently, a commercial enzyme used during processing releases more pectic material into the juice.     

Engineering and characterization of carbohydrate-binding modules for imaging cellulose fibrils biosynthesis in plant protoplasts

Carbohydrate binding modules (CBMs) are noncatalytic domains that assist tethered catalytic domains in substrate targeting. CBMs have therefore been used to visualize distinct polysaccharides present in the cell wall of plant cells and tissues. However, most previous studies provide a qualitative analysis of CBM-polysaccharide interactions, with limited characterization of engineered tandem CBM designs for recognizing polysaccharides like cellulose and limited application of CBM-based probes to visualize cellulose fibrils synthesis in model plant protoplasts with regenerating cell walls. Here, we examine the dynamic interactions of engineered type-A CBMs from families 3a and 64 with crystalline cellulose-I and phosphoric acid swollen cellulose. We generated tandem CBM designs to determine various characteristic properties including binding reversibility toward cellulose-I using equilibrium binding assays. To compute the adsorption (nk_on) and desorption (k_off) rate constants of single versus tandem CBM designs toward nanocrystalline cellulose, we employed dynamic kinetic binding assays using quartz crystal microbalance with dissipation. Our results indicate that tandem CBM3a exhibited the highest adsorption rate to cellulose and displayed reversible binding to both crystalline/amorphous cellulose, unlike other CBM designs, making tandem CBM3a better suited for live plant cell wall biosynthesis imaging applications. We used several engineered CBMs to visualize Arabidopsis thaliana protoplasts with regenerated cell walls using confocal laser scanning microscopy and wide-field fluorescence microscopy. Lastly, we also demonstrated how CBMs as probe reagents can enable in situ visualization of cellulose fibrils during cell wall regeneration in Arabidopsis protoplasts.     

Algal biotechnology industries and research activities in China

In old China there were very few people engaged in the study of the algae,but in new China, freshwater and marine algae are studied by over onehundred old and new phycologists. There is now an algal biotechnologyindustry consisting of an aquaculture industry, producing large amounts ofthe seaweeds Laminaria, Porphyra, Undaria, Gracilaria,eucheumoids, and the microalgae Dunaliella and Spirulina. There is also a phycocolloid industry, producing algin, agar andcarrageenan; an industry producing chemicals and drugs, such as iodine,mannitol, phycocyanin, -carotene, PSS (propylene glycol alginatesulfate) and FPS (fucose-containing sulfated polysaccharides) and anindustry producing food, feed and fertilizer. The Laminariacultivation industry produces about 900,000 t dry Laminaria,probably the largest producer in the world and 13,000 t algin,undoubtedly one of the largest algin producer in the world.     

Chemical characterization of cell-wall polysaccharides from tank-cultivated and wild plants ofDelesseria sanguinea (Hudson) Lamouroux (Ceramiales,delesseriaceae): culture patterns and potent anticoagulant activity

Tank cultivation ofDelesseria sanguinea was investigated in order to manipulate conditions for vegetative growth and to provide biomass for the analysis of cell wall polysaccharides. Seasonality is subject to short-day photoperiodic control. Night-break or long-day conditions prevented fertility in tetrasporophytes and gametophytes and triggered outgrowth of new blades. Long-day illuminations allowed a 1% daily growth rate. Seawater temperature below 13 °C was necessary for inducing formation of new blades. Both wild and cultivated ofD. sanguinea plants contained a non gelling sulfated heteropolysaccharide composed of a galactosyl backbone branched with xylosyl residues. The hot water extract at neutral pH displayed the highest anticoagulant activity (5 μg ml^-1 polysaccharide concentration in APTT clotting assay). No obvious differences were found in polysaccharide chemical composition and properties between gametophytes and sporophytes or between cultivated and wild plants.     

The specific identity and the life history of JapaneseSyringoderma (Syringodermatales,phaeophyceae)

Observations on JapaneseSyringoderma from Rishiri Island, which had been previously identified asS. australe, were made based on newly collected fertile material. These results and comparisons with the type specimen ofSyringoderma abyssicola (=Chlanidophora abyssicola) revealed that the Rishiri Island plants should be identified asSyringoderma abyssicola. Syringoderma abyssicola from Rishiri Island formed unilocular sporangia among the paraphyses on the fan-shaped blades in winter. The first products (uni-spores) in the unilocular sporangia form flagella, and soon after form cell walls before release. Then these reduced gametophytes divide into tetrads and form swarmers, each of which contains a chloroplast with a stigma. These swarmers germinate into branched filaments, from which thicker erect filaments of apical growth tissue. At 5–10 C these erect filaments formed fan-shaped blades under long-day conditions, and unilocular sporangia under short-day conditions corresponding respectively to spring and winter at Rishiri Island. Accordingly, the seasonal growth pattern of the species is considered to be controlled by responses to photoregime and temperature. Dedicated to the memory of the late Professor Muneao Kurogi.     

Cell wall composition of leaves of an inherently fast- and an inherently slow-growing grass species

Differences in the relative growth rules of the inherently slow-growing Deschampsia flexuosa L. and the inherently fast-growing Holcus lanatus L. were reflected in cell wall synthesis in the elongation zone of the leaves. Leaf elongation rates depended on the size of the plant and ranged from 6 to 14 mm d^?1 in Deschampsia and from 12 to 42 mm d^?1 in Holcus. Anatomical data showed that the epidermis and vascular tissue are the important tissues controlling leaf extension. The cell wall polysaccharides of fully expanded leaves of the two species were identical in sugar composition. Enzymatic hydrolysis of polymeric sugars in the cell walls of the sheath and the lamina gave glucose (85%), arabinose (3.5%), fucose (0.5%), xylose (5.0%), mannose (0.5%), galaclose (0.8%) and galacturonic acid (3–4%). This composition applied throughout the blade and the sheath and did not change with ageing. Polysaccharides in the meristems of the two species showed identical sugar compositions with 51–55% glucose, 13–15% galactoronic acid and 13–14% arabinose as the main components. The extension zone was marked by a gradual increase of driselase-digestable polymers (per mm tissue) and a concurrent shift in sugar composition. The massive increase of glucose in the cell wall polymers of the elongation zone is probably caused by cellulose synthesis. The rate of synthesis of cell wall polysaccharides in Holcus was twice as high as that in Deschampsia. The slower-growing Deschampsia has more ferulic acid esterified with cell walls, which might contribute to the slowing of leaf growth. Lignin is not significantly deposited until growth has essentially ceased and is not responsible for the difference in growth rate.     

Composition of the walls of pollen grains of the seagrass Amphibolis antarctica

The composition of walls isolated from pollen grains of the seagrass Amphibolis antarctica was determined. Glucose, galactose, and rhamnose were the major neutral monosaccharides in the wall polysaccharides, and fucose, arabinose, xylose, and mannose were present in minor proportions. No apiose, a monosaccharide present in the wall polysaccharides of the vegetative parts of the seagrass Heterozostera tasmanica, was found. Large amounts of uronic acid (mainly as galacturonic acid) were found in the walls. The monosaccharides were probably present in cellulose and pectic polysaccharides, the latter comprising neutral pectic galactans, and rhamnogalacturonans containing high proportions of rhamnose. The walls contained a small amount of protein; glycine and lysine were the amino acids present in the highest proportions. Histochemical examination of isolated walls confirmed the presence of polyanionic components (pectic polysaccharides), -glucans (cellulose), and protein. The composition of the walls is discussed in relation to analyses of the walls of pollen grains and vegetative organs of other plants.     

Hetero‐trans‐β‐glucanase,an enzyme unique to Equisetum plants,functionalizes cellulose

Cell walls are metabolically active components of plant cells. They contain diverse enzymes, including transglycanases (endotransglycosylases), enzymes that ‘cut and paste’ certain structural polysaccharide molecules and thus potentially remodel the wall during growth and development. Known transglycanase activities modify several cell‐wall polysaccharides (xyloglucan, mannans, mixed‐linkage β‐glucan and xylans); however, no transglycanases were known to act on cellulose, the principal polysaccharide of biomass. We now report the discovery and characterization of hetero‐trans‐β‐glucanase (HTG), a transglycanase that targets cellulose, in horsetails (Equisetum spp., an early‐diverging genus of monilophytes). HTG is also remarkable in predominantly catalysing hetero‐transglycosylation: its preferred donor substrates (cellulose or mixed‐linkage β‐glucan) differ qualitatively from its acceptor substrate (xyloglucan). HTG thus generates stable cellulose–xyloglucan and mixed‐linkage β‐glucan–xyloglucan covalent bonds, and may therefore strengthen ageing Equisetum tissues by inter‐linking different structural polysaccharides of the cell wall. 3D modelling suggests that only three key amino acid substitutions (Trp → Pro, Gly → Ser and Arg → Leu) are responsible for the evolution of HTG's unique specificity from the better‐known xyloglucan‐acting homo‐transglycanases (xyloglucan endotransglucosylase/hydrolases; XTH). Among land plants, HTG appears to be confined to Equisetum, but its target polysaccharides are widespread, potentially offering opportunities for enhancing crop mechanical properties, such as wind resistance. In addition, by linking cellulose to xyloglucan fragments previously tagged with compounds such as dyes or indicators, HTG may be useful biotechnologically for manufacturing stably functionalized celluloses, thereby potentially offering a commercially valuable ‘green’ technology for industrially manipulating biomass.     

Morphogenesis of complex plant cell shapes: the mechanical role of crystalline cellulose in growing pollen tubes

Cellulose is the principal component of the load-bearing system in primary plant cell walls. The great resistance to tensile forces of this polysaccharide and its embedding in matrix components make the cell wall a material similar to a fiber composite. In the rapidly growing pollen tube, the amount of cellulose in the cell wall is untypically low. Therefore, we want to investigate whether the load-bearing function of cellulose is nevertheless important for the architecture of this cell. Enzymatic digestion with cellulase and inhibition of cellulose crystal formation with CGA (1-cyclohexyl-5-(2,3,4,5,6-pentafluorophenoxy)-1λ4,2,4,6-thiatriazin-3-amine) resulted in the formation of tubes with increased diameter in Solanum chacoense and Lilium orientalis when present during germination. In pre-germinated tubes, application of both agents resulted in the transient arrest of growth accompanied by the formation of an apical swelling indicating a role in the mechanical stabilization of this cellular region. Once growth resumed in the presence of cellulase, however, the cell wall in the newly formed tube showed increased amounts of pectins, possibly to compensate for the reduced amount of cellulose. Scanning electron microscopy of pollen tubes subjected to digestion of matrix polysaccharides revealed the mechanical anisotropy of the cell wall. In both Lilium and Solanum, the angle of highest stability revealed by crack formation was significantly below 45°, an indication that in the mature part of the cell cellulose may not the main stress-bearing component against turgor pressure induced tensile stress in circumferential direction.     

The combined effects of black oxidising table olive process and ripening on the cell wall polysaccharides of olive pulp

Olive fruits harvested at cherry and black stages of ripening were processed according the table olive black oxidising processing and sampled after the three main steps: storage in brine, lye treatment and thermal treatment (final product). The results show that the storage in brine contributed positively to the stabilisation of cell wall polysaccharides of olive pulp as the amounts of main polysaccharides practically were maintained in both stages of ripening. The lye treatment introduced degradation of cell walls due to the generalised loss of pectic and hemicellulosic polysaccharides and cellulose, caused by the breakage of ester and hydrogen bonds. On the other hand, the lye treatment introduced shifts in the solubilisation of polysaccharides rendering them more difficult to extract by alkali solutions and enabling their retention in the cellulosic residue, which should contribute positively to cell wall firmness. The thermal treatment introduced degradation of cellulose and increased the solubilisation of polysaccharides with a higher extent in the black olives. This work showed that the differences on the cell wall polysaccharides between stages of ripening are magnified after processing and allowed to conclude that the stage of ripening of olive fruits is determinant for obtaining a final product with adequate texture properties for table olive consumption.     

Cell wall reconstruction and DNA damage repair play a key role in the improved salt tolerance effects of He-Ne laser irradiation in tall fescue seedlings

The improved salt tolerance effects of He–Ne laser were further studied through the estimation of ROS levels, cell viability, DNA damage phenomena, physicochemical properties, and monosaccharide compositions of cell wall polysaccharides in tall fescue seedlings. Salt stress produced deleterious effects on seedlings growth and development. ROS levels and genomic DNA damage were markedly increased compared with controls. Physicochemical activities and monosaccharide proportions of cell wall polysaccharide were also pronouncedly altered. He–Ne laser irradiation improved plant growth retardation via increasing cell viability and reverting physicochemical parameters. According to the results of Fourier transform infrared (FTIR) scanning spectra and DNA apopladder analysis, He–Ne laser was showed to efficiently ameliorate cell wall polysaccharide damage and DNA fragmentation phenomena. The treatment with DNA synthesis inhibitor further demonstrated that DNA damage repair was correlated with the improvement effects of the laser. Therefore, our data illustrated that He–Ne laser irradiation resulted in cell wall reconstruction and genomic DNA injury repair in vivo in salt-stressed seedlings, then enhanced salt tolerance probably via interactions between plant cell wall and related resistance gene expression pattern.     

Biophysical consequences of remodeling the neutral side chains of rhamnogalacturonan I in tubers of transgenic potatoes

Two lines of transgenic potato (Solanum tuberosum L.) plants modified in their cell wall structure were characterized and compared to wild type with regard to biomechanical properties in order to assign functional roles to the particular cell wall polysaccharides that were targeted by the genetic changes. The targeted polymer was rhamnogalacturonan I (RG-I), a complex pectic polysaccharide comprised of mainly neutral oligosaccharide side chains attached to a backbone of alternating rhamnosyl and galacturonosyl units. Tuber rhamnogalacturonan I molecules from the two transformed lines are reduced in linear galactans and branched arabinans, respectively. The transformed tuber tissues were found to be more brittle when subjected to uniaxial compression and the side-chain truncation was found to be correlated with the physical properties of the tissue. Interpretation of the force–deflection curves was aided by a mathematical model that describes the contribution of the cellulose microfibrils, and the results lead to the proposition that the pectic matrix plays a role in transmitting stresses to the load-bearing cellulose microfibrils and that even small changes to the rheological properties of the matrix have consequences for the biophysical properties of the wall.     

Turnover of cell wall polysaccharides of a Vinca rosea suspension culture

Turnover of cell wall components was examined in two growth phases of a batch suspension culture of Vinca rosea L. Three-day-cultured cells (cell division phase) and 5-day-cultured cells (cell expansion phase) were incubated with d -[U-¹⁴C]glucose. After various periods of incubation, extra-cellular polysaccharides (ECP) and cell walls were isolated, and then the cell walls were fractionated to pectic substance, hemicellulose, and cellulose fractions. The results of the measurement of radioactivities and amounts of total carbohydrate in the ECP and cell wall fractions indicated that synthesis of pectic substance was more active in the cell division phase than in the cell expansion phase. From the results of the pulse-chase experiments, in which cells prelabelled by incubation with d -[U-¹⁴C]glucose for 3 h were incubated in a medium containing unlabelled glucose for various periods, the gross degradation, net synthesis, and gross synthesis of cell wall components were estimated. Active degradation and synthesis were observed in the hemicellulose fraction, indicating that active turnover occurred in the hemicellulose fraction, while little degradation was found in the pectic substance and cellulose fractions.     

Composition of the cell walls of different asparagus (Asparagus officinalis) tissues

Portions of stems from the base of asparagus spears (Asparagus officinalis L. cv. Connovor Collossus) were dissected to give the following tissues: (1) pith, which was free of vascular bundles, (2) two surrounding layers, parenchyma and fibre I and II (PFI and PFII), containing parenchyma and vascular bundles, (3) sclerenchyma sheath, (4) epidermis and sub-epidermal layers and (5) asparagus vascular fibre (AVF). The alcohol-insoluble residues (AIRs) from these tissues were shown to be free of starch. They were analysed for moisture and protein, and the component sugars were released by two hydrolytic procedures, which helped to distinguish the sugars from non-cellulosic polysaccharides and cellulose. The AIRs from pith and epidermal tissues were relatively low in xylose, but were rich in cellulosic glucose, and sugars associated with pectic polysaccharides such as galacturonic acid, galactose and arabinose. Their major component polysaccharides (in decreasing amounts) were inferred to be pectic polysaccharides, cellulose, and hemicelluloses. AIR from sclerenchyma was rich in glucose and xylose, suggesting the presence of much cellulose and (acidic) xylans. The AIRs of PFI, PFII and AVF contained significant amounts of xylose in addition tn other sugars, and the major polysaccharides inferred to be present were pectic polysaccharides, cellulose and hemicelluloses, a significant proportion of which may be acidic xylans. Methylation analysis of the AIRs confirmed the above inferences. The bulk of the glucosyl residues were (1–4)-linked, and there were small but significant amounts of (1–4, 6)-linked glucosyl residues (the linkage characteristic of xyloglucans) in all the preparations. The presence of (1–4)-linked galactosyl, (1–5)-linked arabinosyl, terminal galactosyl, terminal arabinosyl, (1–2)- and (1–2, 4)-linked rhamnosyl residues in all the AIRs except that from sclerenchyma, confirmed the presence of significant levels of pectic polysaccharides in all the parenchyma tissues. All the preparations containing vascular tissues contained significant amounts of (1–4)-linked xylosyl residues, probably derived from acidic xylans. Even in the AIR of pith, a significant amount of (1–4)-linked xylosyl residues were detected. This may be due to the ability of these cells and the parenchyma cells associated with the vascular bundles, to undergo lignification in mature asparagus plants.     

Composition of cell wall phenolics and polysaccharides of the potential bioenergy crop –Miscanthus

The composition and concentrations of cell wall polysaccharides and phenolic compounds were analyzed in mature stems of several Miscanthus genotypes, in comparison with switchgrass and reed (Arundo donax), and biomass characteristics were correlated with cell wall saccharification efficiency. The highest cellulose content was found in cell walls of M. sinensis‘Grosse Fontaine’ (55%) and in A. donax (47%) and lowest (about 32%) in M. sinensis‘Adagio’. There was little variation in lignin contents across M. sinensis samples (all about 22–24% of cell wall), however, Miscanthus×giganteus (M × g) cell walls contained about 28% lignin, reed – 23% and switchgrass – 26%. The highest ratios of cellulose/lignin and cellulose/xylan were in M. sinensis‘Grosse Fontaine’ across all samples tested. About the same total content of ester‐bound phenolics was found in different Miscanthus genotypes (23–27 μg/mg cell wall), while reed cell walls contained 17 μg/mg cell wall and switchgrass contained a lower amount of ester‐bound phenolics, about 15 μg/mg cell wall. Coumaric acid was a major phenolic compound ester‐bound to cell walls in plants analyzed and the ratio of coumaric acid/ferulic acid varied from 2.1 to 4.3, with the highest ratio being in M × g samples. Concentration of ether‐bound hydroxycinnamic acids varied greatly (about two‐three‐fold) within Miscanthus genotypes and was also the highest in M × g cell walls, but at a concentration lower than ester‐bound hydroxycinnamic acids. We identified four different forms of diferulic acid esters bound to Miscanthus cell walls and their concentration and proportion varied in genotypes analyzed with the 5‐5‐coupled dimer being the predominant type of diferulate in most samples tested. The contents of lignin and ether‐bound phenolics in the cell wall were the major determinants of the biomass degradation caused by enzymatic hydrolysis.     

Changes in cell-wall polysaccharide composition of developingNitella internodes

Changes in the uronide, neutral-polysacharide, and cellulose composition of the cell wall ofNitella axillaris Braun were followed throughout development of the internodes and correlated with changes in growth rate. As the cells increased in length from 4 to 80 mm during development, the relative growth rate decreased. Cell wall thickness, as measured by wall density, increased in direct proportion to diameter, indicating that cell-wall stress did not change during elogation. Cell-wall analyses were adapted to allow determination of the composition of the wall of single cells. The total amounts of uronides, neutral sugars and cellulose all increased during development. However, as the growth rate decreased, the relative proportions of uronides and neutral sugars, expressed as percent of the dry weight of the wall, decreased, while the proportion of cellulose increased. The neutral sugars liberated upon hydrolysis ofNitella walls are qualitatively similar to those found in hydrolysates of higher plant cell walls: glucose, xylose, mannose, galactose, arabinose fucose and rhamnose. Only the percentage of galactose was found to increase in walls of mature cells, while the percentage of all other sugars decreased. The rate of apposition (g of wall material deposited per unit wall surface area per hour) of neutral polysaccharides decreased rapidly with decreasing growth rate during the early stages of development. The rate of apposition of uronides decreased more steadily throughout development, while that of cellulose, after an early decline, remained constant until dropping off at the end of the elongation period. These correlations between decreasing growth rate and decreasing rate of apposition of neutral sugars and uronides indicate that synthesis of these cell-wall components could be involved in the regulation of the rate of cell elongation inNitella.     

Macroalgal farming in the sea: water motion and nitrate uptake

A better understanding of water motion effects on nutrient uptake by marine crop plants should make it possible to farm the sea more effectively. Farms in China, Japan and the Philippines now grow plants on slack lines or nets that move with passing waves and currents. Nutrient uptake rates are increased onLaminaria farms in China by adding nitrogen-containing fertilizer. In contrast, forests of the giant kelp,Macrocystis grow in California at low nutrient levels without fertilization. The giant kelp, compared as a structure with the slack Chinese farms, has float-supported, spring-like stipes that stretch and recoil as waves pass. This motion seems likely to enhance flow over the thallus surface. In thus study we modified flow around kelp blades in a water tunnel in the laboratory by changing orifice plates, and flow around Chinese-style long-line farms in the sea by tightening them under various sea conditions. Our measurements suggest that if marine farms were designed and operated to increase water movement over the plants being grown, their rates of nutrient uptake, and growth would increase.This paper was presented at the Symposium on Applied Phycology at the Fourth International Phycological Congress, Duke University.     

Indigestibility of plant cell wall by the Australian plague locust, Chortoicetes terminifera

The plant cell wall may play an important role in defence against herbivores since it can be both a barrier to, and nutrient diluter of, the easily digested cell contents. The aim of this study was to investigate the digestibility of the cell wall of three grasses, Triticum aestivum L., Dactyloctenium radulans (R. Br.) Beauv., and Astrebla lappacea (Lindl.) Domin, by the Australian plague locust, Chortoicetes terminifera Walker (Orthoptera: Acrididae, Acridinae) as determined by the Van Soest method [ Van Soest PJ, Robertson JB & Lewis BA (1991) Methods for dietary fiber, neutral detergent fiber, and nonstarch polysaccharides in relation to animal nutrition. Journal of Dairy Science 74: 3583–3597]. Determination of plant cell wall digestion by locusts required a precise methodological procedure to determine both the exact intake and the concentration of cell wall in the diet and the faeces. Plant cell wall determination is affected by the particle size distribution of the dried plant material. All three grasses differed in the percentage of cell wall per gram dry matter and the proportions of hemicellulose, cellulose, and acid‐detergent sulphuric lignin within the cell wall. The locust was unable to digest the cell wall of any of the grasses. Thus, plant cell walls are a mechanical barrier hindering locusts assimilating nutrients. That is, access, rather than nutrient concentration per se, may be limiting nutrient factor.