Open questions in plant cell wall synthesis.

Plant cells are surrounded by strong yet flexible polysaccharide-based cell walls that support cells while also allowing growth by cell expansion. Plant cell wall research has advanced tremendously in recent years. Sequenced genomes of model and crop plants have facilitated cataloguing and characterization of many enzymes involved in cell wall synthesis. Structural information has been generated for several important cell wall-synthesizing enzymes. Important tools have been developed including antibodies raised against a variety of cell wall polysaccharides and glycoproteins, collections of enzyme clones and synthetic glycan arrays for characterizing enzymes, herbicides that specifically affect cell wall synthesis, live-cell imaging probes to track cell wall synthesis, and an inducible secondary cell wall synthesis system. Despite these advances, and often because of the new information they provide, many open questions about plant cell wall polysaccharide synthesis persist. This article highlights some of the key questions that remain open, reviews the data supporting different hypotheses that address these questions, and discusses technological developments that may answer these questions in the future.

Signals and Their Perception for Remodelling, Adjustment and Repair of the Plant Cell Wall.

The integrity of the cell wall is important for plant cells. Mechanical or chemical distortions, tension, pH changes in the apoplast, disturbance of the ion homeostasis, leakage of cell compounds into the apoplastic space or breakdown of cell wall polysaccharides activate cellular responses which often occur via plasma membrane-localized receptors. Breakdown products of the cell wall polysaccharides function as damage-associated molecular patterns and derive from cellulose (cello-oligomers), hemicelluloses (mainly xyloglucans and mixed-linkage glucans as well as glucuronoarabinoglucans in Poaceae) and pectins (oligogalacturonides). In addition, several types of channels participate in mechanosensing and convert physical into chemical signals. To establish a proper response, the cell has to integrate information about apoplastic alterations and disturbance of its wall with cell-internal programs which require modifications in the wall architecture due to growth, differentiation or cell division. We summarize recent progress in pattern recognition receptors for plant-derived oligosaccharides, with a focus on malectin domain-containing receptor kinases and their crosstalk with other perception systems and intracellular signaling events.

Defining natural factors that stimulate and inhibit cellulose:xyloglucan hetero-transglucosylation.

Certain transglucanases can covalently graft cellulose and mixed-linkage ß-glucan (MLG) as donor substrates onto xyloglucan as acceptor substrate and thus exhibit cellulose:xyloglucan endotransglucosylase (CXE) and MLG:xyloglucan endotransglucosylase (MXE) activities in vivo and in vitro. However, missing information on factors that stimulate or inhibit these hetero-transglucosylation reactions limits our insight into their biological functions. To explore factors that influence hetero-transglucosylation, we studied Equisetum fluviatile hetero-trans-ß-glucanase (EfHTG), which exhibits both CXE and MXE activity, exceeding its xyloglucan:xyloglucan homo-transglucosylation (XET) activity. Enzyme assays employed radiolabelled and fluorescently labelled oligomeric acceptor substrates, and were conducted in vitro and in cell walls (in situ). With whole denatured Equisetum cell walls as donor substrate, exogenous EfHTG (extracted from Equisetum or produced in Pichia) exhibited all three activities (CXE, MXE, XET) in competition with each other. Acting on pure cellulose as donor substrate, the CXE action of Pichia-produced EfHTG was up to approximately 300% increased by addition of methanol-boiled Equisetum extracts; there was no similar effect when the same enzyme acted on soluble donors (MLG or xyloglucan). The methanol-stable factor is proposed to be expansin-like, a suggestion supported by observations of pH dependence. Screening numerous low-molecular-weight compounds for hetero-transglucanase inhibition showed that cellobiose was highly effective, inhibiting the abundant endogenous CXE and MXE (but not XET) action in Equisetum internodes. Furthermore, cellobiose retarded Equisetum stem elongation, potentially owing to its effect on hetero-transglucosylation reactions. This work provides insight and tools to further study the role of cellulose hetero-transglucosylation in planta by identifying factors that govern this reaction.

Hemicellulose-remodelling transglycanase activities from charophytes: towards the evolution of the land-plant cell wall.

Transglycanases remodel cell-wall polymers, having a critical impact on many physiological processes. Unlike xyloglucan endotransglucosylase (XET) activity, widely studied in land plants, very little is known about charophyte wall-modifying enzymes - information that would promote our understanding of the 'primordial' wall, revealing how the wall matrix is remodelled in the closest living algal relatives of land plants, and what changed during terrestrialisation. We conducted various in-vitro assays for wall-remodelling transglycosylases, monitoring either (a) polysaccharide-to-[3 H]oligosaccharide transglycosylation or (b) non-radioactive oligosaccharide-to-oligosaccharide transglycosylation. We screened a wide collection of enzyme extracts from charophytes (and early-diverging land plants for comparison) and discovered several homo- and hetero-transglycanase activities. In contrast to most land plants, charophytes possess high trans-ß-1,4-mannanase activity, suggesting that land plants' algal ancestors prioritised mannan remodelling. Trans-ß-1,4-xylanase activity was also found, most abundantly in Chara, Nitella and Klebsormidium. Exo-acting transglycosidase activities (trans-ß-1,4-xylosidase and trans-ß-1,4-mannosidase) were also detected. In addition, charophytes exhibited homo- and hetero-trans-ß-glucanase activities (XET, mixed-linkage glucan [MLG]:xyloglucan endotransglucosylase and cellulose:xyloglucan endotransglucosylase) despite the paucity or lack of land-plant-like xyloglucan and MLG as potential donor substrates in their cell walls. However, trans-α-xylosidase activity (which remodels xyloglucan in angiosperms) was absent in charophytes and early-diverging land plants. Transglycanase action was also found in situ, acting on endogenous algal polysaccharides as donor substrates and fluorescent xyloglucan oligosaccharides as acceptor substrates. We conclude that trans-ß-mannanase and trans-ß-xylanase activities are present and thus may play key roles in charophyte walls (most of which possess little or no xyloglucan and MLG, but often contain abundant ß-mannans and ß-xylans), comparable to the roles of XET in xyloglucan-rich land plants.

Porous Carbon Spheres Derived from Hemicelluloses for Supercapacitor Application.

With the increasing demand for dissolving pulp, large quantities of hemicelluloses were generated and abandoned. These hemicelluloses are very promising biomass resources for preparing carbon spheres. However, the pore structures of the carbon spheres obtained from biomass are usually poor, which extensively limits their utilization. Herein, the carbon microspheres derived from hemicelluloses were prepared using hydrothermal carbonization and further activated with different activators (KOH, K2CO3, Na2CO3, and ZnCl2) to improve their electrochemical performance as supercapacitors. After activation, the specific surface areas of these carbon spheres were improved significantly, which were in the order of ZnCl2 > K2CO3 > KOH > Na2CO3. The carbon spheres with high surface area of 2025 m2/g and remarkable pore volume of 1.07 cm3/g were achieved, as the carbon spheres were activated by ZnCl2. The supercapacitor electrode fabricated from the ZnCl2-activated carbon spheres demonstrated high specific capacitance of 218 F/g at 0.2 A/g in 6 M KOH in a three-electrode system. A symmetric supercapacitor was assembled in 2 M Li2SO4 electrolyte, and the carbon spheres activated by ZnCl2 showed excellent electrochemical performance with high specific capacitance (137 F/g at 0.5 A/g), energy densities (15.4 Wh/kg), and good cyclic stability (95% capacitance retention over 2000 cycles).

Redesigning plant cell walls for the biomass-based bioeconomy.

Lignocellulosic biomass-the lignin, cellulose, and hemicellulose that comprise major components of the plant cell well-is a sustainable resource that could be utilized in the United States to displace oil consumption from heavy vehicles, planes, and marine-going vessels and commodity chemicals. Biomass-derived sugars can also be supplied for microbial fermentative processing to fuels and chemicals or chemically deoxygenated to hydrocarbons. However, the economic value of biomass might be amplified by diversifying the range of target products that are synthesized in living plants. Genetic engineering of lignocellulosic biomass has previously focused on changing lignin content or composition to overcome recalcitrance, the intrinsic resistance of cell walls to deconstruction. New capabilities to remove lignin catalytically without denaturing the carbohydrate moiety have enabled the concept of the "lignin-first" biorefinery that includes high-value aromatic products. The structural complexity of plant cell-wall components also provides substrates for polymeric and functionalized target products, such as thermosets, thermoplastics, composites, cellulose nanocrystals, and nanofibers. With recent advances in the design of synthetic pathways, lignocellulosic biomass can be regarded as a substrate at various length scales for liquid hydrocarbon fuels, chemicals, and materials. In this review, we describe the architectures of plant cell walls and recent progress in overcoming recalcitrance and illustrate the potential for natural or engineered biomass to be used in the emerging bioeconomy.

Facial Synthesis of Adsorbent from Hemicelluloses for Cr(VI) Adsorption.

It is challenging work to develop a low-cost, efficient, and environmentally friendly Cr(VI) adsorbent for waste water treatment. In this paper, we used hemicelluloses from chemical fiber factory waste as the raw material, and prepared two kinds of carbon materials by the green hydrothermal method as adsorbent for Cr(VI). The results showed that hemicelluloses hydrothermally treated with citric acid (HTC) presented spherical shapes, and hemicelluloses hydrothermally treated with ammonia solution (HTC-NH2) provided spongy structures. The adsorption capacity of the samples can be obtained by the Langmuir model, and the adsorption kinetics could be described by the pseudo-second-order model at pH 1.0. The maximum adsorption capacity of HTC-NH2 in the Langmuir model is 74.60 mg/g, much higher than that of HTC (61.25 mg/g). The green hydrothermal treatment of biomass with ammonia solution will provide a simple and feasible way to prepare adsorbent for Cr(VI) in waste water treatment.

Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi.

Fungal pathogens have evolved combinations of plant cell-wall-degrading enzymes (PCWDEs) to deconstruct host plant cell walls (PCWs). An understanding of this process is hoped to create a basis for improving plant biomass conversion efficiency into sustainable biofuels and bioproducts. Here, an approach integrating enzyme activity assay, biomass pretreatment, field emission scanning electron microscopy (FESEM), and genomic analysis of PCWDEs were applied to examine digestibility or degradability of selected woody and herbaceous biomass by pathogenic fungi. Preferred hydrolysis of apple tree branch, rapeseed straw, or wheat straw were observed by the apple-tree-specific pathogen Valsa mali, the rapeseed pathogen Sclerotinia sclerotiorum, and the wheat pathogen Rhizoctonia cerealis, respectively. Delignification by peracetic acid (PAA) pretreatment increased PCW digestibility, and the increase was generally more profound with non-host than host PCW substrates. Hemicellulase pretreatment slightly reduced or had no effect on hemicellulose content in the PCW substrates tested; however, the pretreatment significantly changed hydrolytic preferences of the selected pathogens, indicating a role of hemicellulose branching in PCW digestibility. Cellulose organization appears to also impact digestibility of host PCWs, as reflected by differences in cellulose microfibril organization in woody and herbaceous PCWs and variation in cellulose-binding domain organization in cellulases of pathogenic fungi, which is known to influence enzyme access to cellulose. Taken together, this study highlighted the importance of chemical structure of both hemicelluloses and cellulose in host PCW digestibility by fungal pathogens.

Expression profiling of endo-xylanases during ripening of strawberry cultivars with contrasting softening rates. Influence of postharvest and hormonal treatments.

BACKGROUND: Softening is one of the main features that determine fruit quality during strawberry (Fragaria x ananassa, Duch.) ripening and storage. Being closely related to textural changes, the molecular and biochemical bases underlying strawberry cell-wall metabolism is a matter of interest. Here we investigated the abundance of transcripts encoding putative strawberry endo-xylanases in plant tissues, during fruit ripening and under postharvest and hormonal treatments. Total xylanase activity and expression of related genes in strawberry varieties with contrasting firmness were analyzed. RESULTS: FaXynA and FaXynC mRNA abundance was significantly higher than FaXynB in each plant tissue studied. Higher total xylanase activity was detected at the end of the ripening of the softer cultivar ('Toyonoka') in comparison with the firmer one ('Camarosa'), correlating with the abundance of FaXynA and FaXynC transcripts. Postharvest 1-methylcyclopropene treatment up-regulated FaXynA and FaXynC expressions. FaXynC mRNA abundance decreased with heat treatment but the opposite was observed for FaXynA. Calcium chloride treatment down-regulated FaXynA and FaXynC expression. Both genes responded differently to plant growth regulators' exposure. FaXynC expression was down-regulated by auxins and gibberellins treatment and up-regulated by abscisic acid. FaXynA was up-regulated by auxins, while no changes in mRNA levels were evident by abscisic acid and gibberellins treatment. Ethephon exposure did not change FaXynA and FaXynC expressions. CONCLUSION: New knowledge about the presence of xylanases in ripening strawberry fruit and their response to postharvest and hormonal treatments is provided. Our findings suggest a role for endo-xylanases in hemicelluloses depolymerization and possibly in strawberry fruit softening. © 2020 Society of Chemical Industry.

Fractionation of Waste MDF by Steam Refining.

In view of the expected increase in available waste medium-density fiberboard (MDF) and the current insufficient and unsatisfactory disposal capacities, efficient ways of recycling the waste material need to be developed. In this study, the potential of steam refining as a method to hydrolyze the resins, isolate fibers, and obtain a hemicellulose-rich extract available for further utilization in the context of a biorefinery was assessed. Two different MDF waste samples, as well as poplar (Populus spp.) and spruce (Picea spp.) wood chips for benchmarking, were treated over a severity range from 2.47 to 3.95. The separated fiber and extract fractions were analyzed with regard to yield, content of carbohydrates, acids, degradation products, and nitrogen. A fiber fraction of more than 70% yield and an extract containing up to 30% of carbohydrates for further processing can be gained by steam-refining waste MDF. At low severities, most of the nitrogen-based compounds are solubilized. Increasing the severity leads to a decrease in nitrogen in the extract as the nitrogen compounds are converted into volatiles. A non-hydrolysable resin residue remains on the fibers, independent of the treatment severity. In comparison to the benchmark samples, the extract fraction of waste MDF shows a high pH of 8 and high amounts of acetic and formic acid. The generation of furfural and 5-hydroxymethylfurfural (5-HMF) on the other hand is suppressed. Distinct differences in carbohydrate hydrolysis behavior between waste MDF and conventional wood can be observed. Especially, the mannose-containing constituents seem to be resistant to hydrolysis reactions in the milieu created in MDF fractionation.

Probing the Molecular Structure and Orientation of the Leaf Surface of Brassica oleracea L. by Polarization Modulation-Infrared Reflection-Absorption Spectroscopy.

The surface of most aerial plant organs is covered with the cuticle, a membrane consisting of a variety of organic compounds, including waxes, cutin (a polyester) and polysaccharides. The cuticle serves as the multifunctional interface between the plant and the environment, and plays a major role in protecting plants against various environmental stress factors. Characterization of the molecular arrangements in the intact cuticle is critical for the fundamental understanding of its physicochemical properties; however, this analysis remains technically challenging. Here, we describe the nondestructive characterization of the intact cuticle of Brassica oleracea L. leaves using polarization modulation-infrared (IR) reflection-absorption spectroscopy (PM-IRRAS). PM-IRRAS has a probing depth of less than several hundreds of nanometers, and reveals the crystalline structure of the wax covering the cuticle surface (epicuticular wax) and the nonhydrogen-bonding character of cutin. Combined analysis using attenuated total reflection-IR spectra suggested that hemicelluloses xylan and xyloglucan are present in the outer cuticle region close to the epicuticular wax, whereas pectins are dominant in the inner cuticle region (depth of ≤2 µm). PM-IRRAS can also determine the average orientation of the cuticular molecules, as indicated by the positive and negative spectral peaks. This unique advantage reveals the orientational order in the intact cuticle; the hydrocarbon chains of the epicuticular wax and cutin and the backbones of hemicelluloses are oriented perpendicular to the leaf surface. PM-IRRAS is a versatile, informative and easy-to-use technique for studying plant cuticles because it is nondestructive and does not require sample pretreatment and background measurements.

Characteristics of the Water- and Alkali-Soluble Hemicelluloses Fractionated by Sequential Acidification and Graded-Ethanol from Sweet Maize Stems.

Sweet maize stems were treated with hot water and potassium hydroxide to fractionate hemicellulosic polymers. The results showed that the water-soluble hemicelluloses were mainly composed of glucose (27.83%), xylose (27.32%), and galactose (16.81%). In comparison, alkali-soluble hemicelluloses fractionated by acidification and a graded ethanol solution (10%, 20%, 35%, 50%, 65%, and 80%) were mainly composed of xylose (69.73 to 88.62%) and arabinose (5.41 to 16.20%). More highly branched hemicelluloses tended to be precipitated in a higher concentration of ethanol solution, as revealed by the decreasing xylose to arabinose ratio from 16.43 to 4.21. Structural characterizations indicated that alkali-soluble hemicelluloses fractionated from sweet maize stems were mainly arabinoxylans. The results provided fundamental information on hemicelluloses composition and structure and their potential utilization in the fields of biofuels, biochemicals, and biomaterials.

The Ceratopteris (fern) developing motile gamete walls contain diverse polysaccharides, but not pectin.

MAIN CONCLUSION: Unlike most plant cell walls, the five consecutive walls laid down during spermatogenesis in the model fern Ceratopteris contain sparse cellulose, lack pectin and are enriched with callose and hemicelluloses. Seed-free plants like bryophytes and pteridophytes produce swimming male gametes for sexual reproduction. During spermatogenesis, unique walls are formed that are essential to the appropriate development and maturation of the motile gametes. Other than the detection of callose and general wall polysaccharides in scattered groups, little is known about the sequence of wall formation and the composition of these walls during sperm cell differentiation in plants that produce swimming sperm. Using histochemistry and immunogold localizations, we examined the distribution of callose, cellulose, mannan and xylan-containing hemicelluloses, and homogalacturonan (HG) pectins in the special walls deposited during spermatogenesis in Ceratopteris. Five walls are produced in sequence and each has a unique fate. The first wall (W1) contains callose and sparse xylan-containing hemicelluloses. Wall two (W2) is thin and composed of cellulose crosslinked by xylan-containing hemicelluloses. The third wall (W3) is thick and composed entirely of callose, and the fourth wall (W4) is built of cellulose heavily crosslinked by galactoxyloglucan hemicelluloses. Wall five (W5) is an arabinogalactan protein (AGP)-rich matrix in which the gamete changes shape and multiple flagella elongate. We detected no esterified or unesterified HG pectins in any of the walls laid down during spermatogenesis. To consider evolutionary modifications in cell walls associated with motile gametes, comparisons are presented with male gametophyte and spermatogenous cell walls across plant groups.

Reconstruction of lignin and hemicelluloses by aqueous ethanol anti-solvents to improve the ionic liquid-acid pretreatment performance of Arundo donax Linn.

Ionic liquid (IL)-acid pretreatment is known to not only enhance the enzymatic hydrolysis efficiency of lignocellulose but also to generate deposits on the surface of fiber by conventional water regeneration, which retard the increment. In this study, ethanol aqueous solution regeneration was developed as a new method to change the substrates characteristics for IL-acid pretreatment and their effects on the enzymatic hydrolysis were evaluated. Following the IL-acid reaction, the biomass slurry was subjected to ethanol aqueous solution at various concentration. Results indicated that anti-solvent choice significantly influenced the reconstruction of both hemicelluloses and lignin as a result of the competition between water and ethanol. The partial removal of hemicelluloses and suitable lignin re-localization contributed to a more porous structure. Consequently, the cellulose digestibility of aqueous ethanol regenerated samples was dramatically enhanced to ∼100% and approximately 11- and 2-fold higher than that of untreated and conventional water regenerated pretreated samples, respectively. A giant leap in the initial rate of enzymatic hydrolysis was also detected in 50% ethanol aqueous solution regenerated samples and only about 10 hr was needed to convert 80% of cellulose to glucose due to the appearance of cellulose II hydrate-like and more porous structure.

Polysaccharide compositions of collenchyma cell walls from celery (Apium graveolens L.) petioles.

BACKGROUND: Collenchyma serves as a mechanical support tissue for many herbaceous plants. Previous work based on solid-state NMR and immunomicroscopy suggested collenchyma cell walls (CWs) may have similar polysaccharide compositions to those commonly found in eudicotyledon parenchyma walls, but no detailed chemical analysis was available. In this study, compositions and structures of cell wall polysaccharides of peripheral collenchyma from celery petioles were investigated. RESULTS: This is the first detailed investigation of the cell wall composition of collenchyma from any plant. Celery petioles were found to elongate throughout their length during early growth, but as they matured elongation was increasingly confined to the upper region, until elongation ceased. Mature, fully elongated, petioles were divided into three equal segments, upper, middle and lower, and peripheral collenchyma strands isolated from each. Cell walls (CWs) were prepared from the strands, which also yielded a HEPES buffer soluble fraction. The CWs were sequentially extracted with CDTA, Na2CO3, 1 M KOH and 4 M KOH. Monosaccharide compositions of the CWs showed that pectin was the most abundant polysaccharide [with homogalacturonan (HG) more abundant than rhamnogalacturonan I (RG-I) and rhamnogalacturonan II (RG-II)], followed by cellulose, and other polysaccharides, mainly xyloglucans, with smaller amounts of heteroxylans and heteromannans. CWs from different segments had similar compositions, but those from the upper segments had slightly more pectin than those from the lower two segments. Further, the pectin in the CWs of the upper segment had a higher degree of methyl esterification than the other segments. In addition to the anticipated water-soluble pectins, the HEPES-soluble fractions surprisingly contained large amounts of heteroxylans. The CDTA and Na2CO3 fractions were rich in HG and RG-I, the 1 M KOH fraction had abundant heteroxylans, the 4 M KOH fraction was rich in xyloglucan and heteromannans, and cellulose was predominant in the final residue. The structures of the xyloglucans, heteroxylans and heteromannans were deduced from the linkage analysis and were similar to those present in most eudicotyledon parenchyma CWs. Cross polarization with magic angle spinning (CP/MAS) NMR spectroscopy showed no apparent difference in the rigid and semi-rigid polysaccharides in the CWs of the three segments. Single-pulse excitation with magic-angle spinning (SPE/MAS) NMR spectroscopy, which detects highly mobile polysaccharides, showed the presence of arabinan, the detailed structure of which varied among the cell walls from the three segments. CONCLUSIONS: Celery collenchyma CWs have similar polysaccharide compositions to most eudicotyledon parenchyma CWs. However, celery collenchyma CWs have much higher XG content than celery parenchyma CWs. The degree of methyl esterification of pectin and the structures of the arabinan side chains of RG-I show some variation in the collenchyma CWs from the different segments. Unexpectedly, the HEPES-soluble fraction contained a large amount of heteroxylans.

Hetero-trans-ß-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 ß-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-ß-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 ß-glucan) differ qualitatively from its acceptor substrate (xyloglucan). HTG thus generates stable cellulose-xyloglucan and mixed-linkage ß-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.

Cell wall dynamics during apple development and storage involves hemicellulose modifications and related expressed genes.

BACKGROUND: Fruit quality depends on a series of biochemical events that modify appearance, flavour and texture throughout fruit development and ripening. Cell wall polysaccharide remodelling largely contributes to the elaboration of fleshy fruit texture. Although several genes and enzymes involved in cell wall polysaccharide biosynthesis and modifications are known, their coordinated activity in these processes is yet to be discovered. RESULTS: Combined transcriptomic and biochemical analyses allowed the identification of putative enzymes and related annotated members of gene families involved in cell wall polysaccharide composition and structural changes during apple fruit growth and ripening. The early development genes were mainly related to cell wall biosynthesis and degradation with a particular target on hemicelluloses. Fine structural evolutions of galactoglucomannan were strongly correlated with mannan synthase, glucanase (GH9) and ß-galactosidase gene expression. In contrast, fewer genes related to pectin metabolism and cell expansion (expansin genes) were observed in ripening fruit combined with expected changes in cell wall polysaccharide composition. CONCLUSIONS: Hemicelluloses undergo major structural changes particularly during early fruit development. The high number of early expressed ß-galactosidase genes questions their function on galactosylated structures during fruit development and storage. Their activity and cell wall substrate remains to be identified. Moreover, new insights into the potential role of peroxidases and transporters, along with cell wall metabolism open the way to further studies on concomitant mechanisms involved in cell wall assembly/disassembly during fruit development and storage.

Mannan biotechnology: from biofuels to health.

Mannans of different structure and composition are renewable bioresources that can be widely found as components of lignocellulosic biomass in softwood and agricultural wastes, as non-starch reserve polysaccharides in endosperms and vacuoles of a wide variety of plants, as well as a major component of yeast cell walls. Enzymatic hydrolysis of mannans using mannanases is essential in the pre-treatment step during the production of second-generation biofuels and for the production of potentially health-promoting manno-oligosaccharides (MOS). In addition, mannan-degrading enzymes can be employed in various biotechnological applications, such as cleansing and food industries. In this review, fundamental knowledge of mannan structures, sources and functions will be summarized. An update on various aspects of mannan-degrading enzymes as well as the current status of their production, and a critical analysis of the potential application of MOS in food and feed industries will be given. Finally, emerging areas of research on mannan biotechnology will be highlighted.

Rheological Flow Behavior of Structural Polysaccharides from Edible Tender Cladodes of Wild, Semidomesticated and Cultivated 'Nopal' (Opuntia) of Mexican Highlands.

The aim of this study was to quantify the content of polysaccharides of edible tender cladodes (nopalitos) of three species of Opuntia and to evaluate the rheological flow behavior of isolated polysaccharides. A completely randomized experimental design was used to characterize a wild (O. streptacantha), a semidomesticated (O. megacantha) and a domesticated (O. ficus-indica) species. Mucilage content was higher (4.93 to 12.43 g 100 g-1 dry matter), tightly bound hemicelluloses were lower (3.32 to 1.81 g 100 g-1 dry matter) and pectins and loosely bound hemicelluloses were not different in wild than in domesticated species. Aqueous solution/suspensions of mucilage, pectins, hemicellulose and cellulose of all species showed non-Newtonian behavior under simple shear flow. The flow behavior of the structural polysaccharides was well described by the Ostwald de-Waele model. Pectins and mucilages exhibited the highest consistency indexes (K values ranged from 0.075 to 0.177 Pasn) with a moderated shear-thinning behavior (n values ranged from 0.53 to 0.67). Cellulose dispersions exhibited the most shear-thinning behavior (n values ranged from 0.17 to 0.41) and hemicelluloses showed a tendency to Newtonian flow (n values ranged from 0.82 to 0.97). The rheological flow properties of these polysaccharides may be useful to improve the textural and sensory qualities of some foods and pharmaceutical materials. Moreover, they can emerge as functional ingredients mainly due to the nutraceutical properties that have been attributed to nopalitos.

Selectively fractionate hemicelluloses with high molecular weight from poplar thermomechanical pulp by tetramethylammonium hydroxide.

Selective fractionation of hemicelluloses is of great significance for realizing high-value application of hemicelluloses and comprehensive utilization of lignocellulosic biomass. Tetramethylammonium hydroxide (TMAH) solvent has been confirmed as a promising solvent to selectively fractionate hemicelluloses from holocellulose. Herein, TMAH solvent was adopted to pretreat poplar thermomechanical pulp (PTMP) for the selective fractionation of hemicelluloses and enhancement of enzymatic hydrolysis performance of residues. The maximal hemicelluloses yield (65.0 %) and excellent cellulose retention rate (93.3 %) were achieved after pretreatment by the 25 wt% TMAH solvent, while the delignification was only 33.9 %. The hemicelluloses fractions could be selectively fractionated with high molecular weights (109,800-118,500 g/mol), the contents of Klason lignin in them were low (3.2-5.9 %), and the dominating structure of them was 4-O-methylglucurono-ß-D-xylan. Compared to the H2SO4 and NaOH methods, the hemicelluloses fractionated by TMAH method exhibited higher yields, more complete structures and higher molecular weights. Furthermore, the crystalline structure of cellulose practically remained stable, and the highest yield of enzymatic hydrolysis glucose was 57.5 %, which was 3.3 times of that of PTMP. The fractionation effectiveness of TMAH solvent was not significantly reduced after repeatedly recycling. This work demonstrated TMAH solvent could selectively fractionate hemicelluloses from PTMP and efficiently promote sustainable poplar-based biorefinery.