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The current expansion of Equisetum palustre in wetlands across the Northern Hemisphere has led to an increase in reports of adverse effects in livestock. In light of the limited reduction potential of toxic Equisetum alkaloids through feed conservation measures, it is essential to identify effective strategies to manage E. palustre infested biomass. This study examined the impact and efficacy of biomethanization processes on the biodegradation of Equisetum alkaloids. To monitor the biodegradation in such complex matrices, a sample preparation method and a high-performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS/MS) method were established. The developed method was successfully employed to quantify the total residual content of Equisetum alkaloids in fermentation residues following biomethanization in batch experiments. It was demonstrated that under such conditions a degradation greater than 95 % was achieved. The biomethanization process described here is currently the most effective biological treatment for reducing Equisetum alkaloids. It is notable that the methane yields remained largely unaffected, indicating that field cuttings contaminated with E. palustre can be safely and economically processed via biomethanization which would be a key prerequisite for a circular bioeconomy., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Till Beuerle reports equipment, drugs, or supplies was provided by Deutsche Forschungsgemeinschaft project number 511817356. Dennis Melchert reports equipment, drugs, or supplies was provided by Deutsche Forschungsgemeinschaft project number 511817356. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The aim was to use the agricultural weed and silica (Si) hyperaccumulator Equisetum arvense as Si fertilizer in plant cultivation. We investigated (1) the Si uptake in various Equisetum species, (2) where Si accumulates in the Equisetum plant, (3) processing methods to release as much Si as possible from dried, ground E. arvense plants and (4) which treatment yields gives the highest uptake of Si in young wheat plants cultivated in soil containing ground E. arvense. The results showed that E. arvense containes 22% Si and was among the best Si accumulators. Equisetum arvense accumulates Si as both soluble and firmly bound fractions. Amorphous silica (SiO 2 ) accumulates in the outer cell walls of epidermis of the entire plant. Regarding the processing method, a longer treatment time, greater concentration of Equisetum, boiling, and the addition of sodium bicarbonate increased the Si availability in ground, dried E. arvense. The addition of untreated, ground, dried E. arvense to the soil, corresponding to 160 kg Si ha -1 , increased the available Si in the soil and the Si uptake in wheat plants by five-fold, compared with the control. Boiling the ground E. arvense increased the Si uptake by 10 times, and the of sodium bicarbonate increased the availability and uptake by 40 times, compared with the control. In conclusion, dried, ground E. arvense can be used as a Si fertilizer as is, after boiling for a slightly better effect, or with sodium bicarbonate (up to a similar amount as the ground material) for best effect., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Masson SAS.)

The kinetic properties of Rubisco, a key enzyme for photosynthesis, have been examined in numerous plant species. However, this information on some plant groups, such as ferns, is scarce. This study examined Rubisco carboxylase activity and leaf Rubisco levels in seven ferns, including four Equisetum plants (E. arvense, E. hyemale, E. praealtum, and E. variegatum), considered living fossils. The turnover rates of Rubisco carboxylation (k cat c ) in E. praealtum and E. hyemale were comparable to those in the C 4 plants maize (Zea mays) and sorghum (Sorghum bicolor), whose k cat c values are high. Rubisco CO 2 affinity, estimated from the percentage of Rubisco carboxylase activity under CO 2 unsaturated conditions in k cat c in these Equisetum plants, was low and also comparable to that in maize and sorghum. In contrast, k cat c and CO 2 affinities of Rubisco in other ferns, including E. arvense and E. variegatum were comparable with those in C 3 plants. The N allocation to Rubisco in the ferns examined was comparable to that in the C 3 plants. These results indicate that E. praealtum and E. hyemale have abundant Rubisco with high k cat c and low CO 2 affinity, whereas the carboxylase activity and abundance of Rubisco in other ferns were similar to those in C 3 plants. Herein, the Rubisco properties of E. praealtum and E. hyemale were discussed regarding their evolution and physiological implications., (© 2023. The Author(s) under exclusive licence to The Botanical Society of Japan.)

Plant cell walls are complex structures mainly made up of carbohydrate and phenolic polymers. In addition to their structural roles, cell walls function as external barriers against pathogens and are also reservoirs of glycan structures that can be perceived by plant receptors, activating Pattern-Triggered Immunity (PTI). Since these PTI-active glycans are usually released upon plant cell wall degradation, they are classified as Damage Associated Molecular Patterns (DAMPs). Identification of DAMPs imply their extraction from plant cell walls by using multistep methodologies and hazardous chemicals. Subcritical water extraction (SWE) has been shown to be an environmentally sustainable alternative and a simplified methodology for the generation of glycan-enriched fractions from different cell wall sources, since it only involves the use of water. Starting from Equisetum arvense cell walls, we have explored two different SWE sequential extractions (isothermal at 160 ºC and using a ramp of temperature from 100 to 160 ºC) to obtain glycans-enriched fractions, and we have compared them with those generated with a standard chemical-based wall extraction. We obtained SWE fractions enriched in pectins that triggered PTI hallmarks in Arabidopsis thaliana such as calcium influxes, reactive oxygen species production, phosphorylation of mitogen activated protein kinases and overexpression of immune-related genes. Notably, application of selected SWE fractions to pepper plants enhanced their disease resistance against the fungal pathogen Sclerotinia sclerotiorum. These data support the potential of SWE technology in extracting PTI-active fractions from plant cell wall biomass containing DAMPs and the use of SWE fractions in sustainable crop production., (© 2023. The Author(s).)

Sixteen undescribed apocarotenoids (1-16), along with 22 known analogues, were isolated from the aerial parts of Equisetum debile. Their structures, including absolute configurations, were elucidated by NMR, HRESIMS, X-ray diffraction analysis, the modified Mosher's method and the quantum-chemical calculation of electronic circular dichroism (ECD) spectra. Compounds 1-9, 11-12 are the first example of C 16 -apocarotenoids appeared in nature. The plausible biosynthetic pathway of 1-16 was proposed. Moreover, the isolates were evaluated for their lipid-lowering activity, and the results showed that 13, 14, 15, 22, 31, 32 and 33 could remarkably decrease the levels of both TC and TG in FFA induced HepG2 cells at 20 μM. The oil red staining assay further demonstrated the lipid-lowering effects of 13, 14 and 15. The western blot results indicated that compounds 13, 14 and 15 could regulate the lipid metabolism via the activation of the AMPK/ACC/SREBP-1c signaling pathway. A preliminary structure-activity relationship (SAR) study of the isolates indicated that the apocarotenoids with 6/5 ring system displayed more potent lipid-lowering effects., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Mixed β(1,3;1,4)-linkage glucan (MLG) is commonly found in the monocot lineage, at particularly high levels in the Poaceae family, but also in the evolutionally distant genus, Equisetum. MLG has several properties that make it unique from other plant cell wall polysaccharides. It consists of β1,4-linked polymers of glucose interspersed with β1,3-linkages, but the presence of β1,3-linkages provides quite different physical properties compared to its closest form of the cell wall component, cellulose. The mechanisms of MLG biosynthesis have been investigated to understand whether single or multiple enzymes are required to build mixed linkages in the glucan chain. Currently, MLG synthesis by a single enzyme is supported by mutagenesis analyses of cellulose synthase-like F6, the major MLG synthase, but further investigation is needed to gather mechanistic insights. Because of transient accumulation of MLG in elongating cells and vegetative tissues, several hypotheses have been proposed to explain the role of MLG in the plant cell wall. Studies have been carried out to identify gene expression regulators during development and light cycles as well as enzymes involved in MLG organization in the cell wall. A role of MLG as a storage molecule in grains is evident, but the role of MLG in vegetative tissues is still not well understood. Characterization of a cell wall component is difficult due to the complex heterogeneity of the plant cell wall. However, as detailed in this review, recent exciting research has made significant impacts in the understanding of MLG biology in plants., (© The Author(s) 2021. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Certain transglucanases can covalently graft cellulose and mixed-linkage β-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-β-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., (© 2020 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Transglycanases are enzymes that remodel the primary cell wall in plants, potentially loosening and/or strengthening it. Xyloglucan endotransglucosylase (XET; EC 2.4.1.207), ubiquitous in land plants, is a homo-transglucanase activity (donor, xyloglucan; acceptor, xyloglucan) exhibited by XTH (xyloglucan endotransglucosylase/hydrolase) proteins. By contrast, hetero-trans-β-glucanase (HTG) is the only known enzyme that is preferentially a hetero-transglucanase. Its two main hetero-transglucanase activities are MLG : xyloglucan endotransglucosylase (MXE) and cellulose : xyloglucan endotransglucosylase (CXE). HTG is highly acidic and found only in the evolutionarily isolated genus of fern-allies, Equisetum. We now report genes for three new highly acidic HTG-related XTHs in E. fluviatile (EfXTH-A, EfXTH-H and EfXTH-I). We expressed them heterologously in Pichia and tested the encoded proteins' enzymic activities to determine whether their acidity and/or their Equisetum-specific sequences might confer high hetero-transglucanase activity. Untransformed Pichia was found to secrete MLG-degrading enzyme(s), which had to be removed for reliable MXE assays. All three acidic EfXTHs exhibited very predominantly XET activity, although low but measurable hetero-transglucanase activities (MXE and CXE) were also detected in EfXTH-H and EfXTH-I. We conclude that the extremely high hetero-transglucanase activities of Equisetum HTG are not emulated by similarly acidic Equisetum XTHs that share up to 55.5% sequence identity with HTG., Competing Interests: Declaration of Competing Interest A patent application (WO2015044209) has been filed by BASF Agricultural Solutions Belgium NV and The University of Edinburgh for the use of hetero-transglycosylase. F.M., A.H., S.C.F., T.S. and C.H. are inventors., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Current cell-wall models assume no covalent bonding between cellulose and hemicelluloses such as xyloglucan or mixed-linkage β-d-glucan (MLG). However, Equisetum hetero-trans-β-glucanase (HTG) grafts cellulose onto xyloglucan oligosaccharides (XGOs) - and, we now show, xyloglucan polysaccharide - in vitro, thus exhibiting CXE (cellulose:xyloglucan endotransglucosylase) activity. In addition, HTG also catalyzes MLG-to-XGO bonding (MXE activity). In this study, we explored the CXE action of HTG in native plant cell walls and tested whether expansin exposes cellulose to HTG by disrupting hydrogen bonds. To quantify and visualize CXE and MXE action, we assayed the sequential release of HTG products from cell walls pre-labeled with substrate mimics. We demonstrated covalent cellulose-xyloglucan bonding in plant cell walls and showed that CXE and MXE action was up to 15% and 60% of total transglucanase action, respectively, and peaked in aging, strengthening tissues: CXE in xylem and cells bordering intercellular canals and MXE in sclerenchyma. Recombinant bacterial expansin (EXLX1) strongly augmented CXE activity in vitro. CXE and MXE action in living Equisetum structural tissues potentially strengthens stems, while expansin might augment the HTG-catalyzed CXE reaction, thereby allowing efficient CXE action in muro. Our methods will enable surveys for comparable reactions throughout the plant kingdom. Furthermore, engineering similar hetero-polymer formation into angiosperm crop plants may improve certain agronomic traits such as lodging tolerance., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Horsetail (Equisetum arvense) plants grew healthily for 10 weeks under both Si-deficient and Si-replete conditions. After 10 weeks, plants grown under Si-deficient conditions succumbed to fungal infection. We have used NanoSIMS and fluorescence microscopy to investigate silica deposition in the tissues of these plants. Horsetail grown under Si-deficient conditions did not deposit identifiable amounts of silica in their tissues. Plants grown under Si-replete conditions accumulated silica throughout their tissues and especially in the epidermis of the outer side of the leaf and the furrow region of the stem where it was continuous and often, as a double layer suggestive of a barrier function. We have previously shown, both in vivo (in horsetail and thale cress) and in vitro (using an undersaturated solution of Si(OH) 4 ), that callose is a "catalyst" of plant silica deposition. Here we support this finding by comparing the deposition of silica to that of callose and by showing that they are co-localized. We propose the existence of a synergistic mechanical protection by callose and silica against pathogens in horsetail, whereby the induction of callose synthesis and deposition is the first, biochemical line of defence and callose-induced precipitation of silica is the second, adventitious mechanical barrier., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Silicon (Si) is a beneficial element to plants, and its absorption via transporters leads to protective effects against biotic and abiotic stresses. In higher plants, two groups of root transporters for Si have been identified: influx transporters (Lsi1) and efflux transporters (Lsi2). Lsi1 transporters belong to the NIPIII aquaporins, and functional Lsi1s have been found in many plants species. Much less is known about Lsi2s that have been characterized in only a few species. Horsetail (Equisetum arvense), known among the highest Si accumulators in the plant kingdom, is a valuable model to study Si absorption and deposition. In this study, we first analyzed discrete Si deposition patterns in horsetail shoots, where ubiquitous silicification differs markedly from that of higher plants. Then, using the sequenced horsetail root transcriptome, two putative Si efflux transporter genes, EaLsi2-1 and EaLsi2-2, were identified. These genes share low sequence similarity with their homologues in higher plants. Further characterisation of EaLsi2-1 in transient expression assay using Nicotiana benthamiana epidermal cells confirmed transmembrane localization. In order to determine their functionality, the EaLsi2-1 was expressed in Xenopus oocytes, confirming that the translated protein was efficient for Si efflux. Both genes were equally expressed in roots and shoots, but interestingly, showed a much higher expression in the shoots than in the roots in contrast to Lsi2s found in other plants, a result consistent with the specific anatomy of horsetail and its rank as one of the highest Si accumulators among plant species., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

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., (© 2015 The Authors The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Unicellular model plant systems (vegetative microspores of horsetail Equisetum arvense and pollen of six plant species Corylus avellana, Dolichothele albescens Populus balsamifera, Salix caprea, Saintpaulia ionantha, Tulipa hybridum, on which autofluorescence and fluorescence after histochemical treatment studied, have been represented as bioindicators of ozone. It has found that low doses of ozone 0.005 or 0.008 μl/l did not affect or stimulate the autofluorescence of the samples with the ability to germinate in an artificial medium. In higher ozone concentrations (0.032 μl/l) either the decrease in the intensity of the emission or changing in the position of the maxima in the fluorescence spectrum (new 515-520 nm maximum characteristic for the green-and yellow area has appeared) were observed. In dose of 0.2 μl/l, higher than above the threshold of danger to human health, autofluorescence in all samples fell down to up to zero, and there was no the ability to germinate. In this case the formation of lipofuscin-like compounds fluoresced in blue with maxima from 440 to 485 nm was observed. Stress metabolites, known as neurotransmitters biogenic amines, were found in treated cells as determined on the characteristic fluorescence at 460-480 nm in the samples after a specific histochemical reactions for catecholamines (with glyoxylic acid) or for histamine (with o-phthalic aldehyde). Increased intensity of the emission under the treatment with ozone (total doses from 0.012 to 0.032 μl/l) was associated with an increase in the concentrations of catecholamines and histamine. The fluorescent analysis on undamaged cells-possible bioindicators of ozone can be useful in ecomonitoring for earlier warning about health hazardous concentrations of this compound in the air.

The rhizome is responsible for the invasiveness and competitiveness of many plants with great economic and agricultural impact worldwide. Besides its value as an invasive organ, the rhizome plays a role in the establishment and massive growth of forage, providing biomass for biofuel production. Despite these features, little is known about the molecular mechanisms that contribute to rhizome growth, development, and function in plants. In this work, we characterized the proteome of rhizome apical tips and elongation zones from different species using a GeLC-MS/MS (one-dimensional electrophoresis in combination with liquid chromatography coupled online with tandem mass spectrometry) spectral-counting proteomics strategy. Five rhizomatous grasses and an ancient species were compared to study the protein regulation in rhizomes. An average of 2200 rhizome proteins per species were confidently identified and quantified. Rhizome-characteristic proteins showed similar functional distributions across all species analyzed. The over-representation of proteins associated with central roles in cellular, metabolic, and developmental processes indicated accelerated metabolism in growing rhizomes. Moreover, 61 rhizome-characteristic proteins appeared to be regulated similarly among analyzed plants. In addition, 36 showed conserved regulation between rhizome apical tips and elongation zones across species. These proteins were preferentially expressed in rhizome tissues regardless of the species analyzed, making them interesting candidates for more detailed investigative studies about their roles in rhizome development.

Concentrations of (137)Cs and (40)K in different tissues of edible wild herbaceous plants, that is, leaf blade and petiole for giant butterbur (Petasites japonicas (Siebold et Zucc.) Maxim.), and leaf, stem and strobilus for fertile shoot of field horsetail (Equisetum arvense L.) were measured in 2012-2014 to clarify the effect in Japan from the Fukushima Daiichi Nuclear Power Plant accident. The concentrations of (137)Cs decreased with time with effective half-lives of ca. 450 d and 360 d for giant butterbur and field horsetail, respectively. The ANOVA test revealed that (40)K and (137)Cs distributions in leaf blade and petiole for giant butterbur and leaf and stem for field horsetail were different. Therefore, other plants, leaf and stem for Japanese knotweed (Fallopia japonica (Houtt.) Ronse Decr.) and Canada goldenrod (Solidago canadensis L.), and leaf blade and petiole for gingko (Ginkgo biloba L.) and Someiyoshino cherry (Cerasus × yedoensis (Matsum.) A.V.Vassil. 'Somei-yoshino') were collected from the same sampling field and their (137)Cs and (40)K concentrations were compared to those in the giant butterbur and field horsetail parts. For (137)Cs, concentrations in leaf blade and leaf parts were 1.1-6.0 times higher than those in petiole and stem parts for all six plants. On the other hand, (40)K concentrations in leaf blade and leaf parts were 0.40-0.97 of those observed in petiole and stem parts. Discrimination ratios of (40)K/(137)Cs of leaf blade to petiole or leaf to stem were then calculated and they ranged from 0.09 to 0.57. These results suggested that Cs and K did not behave similarly in these plants. Thus, to understand the radiocesium fate in plants, K measurement results should not be used as an analog for Cs behavior although Cs is known to have a similar chemical reactivity to that of K., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

(1,3; 1,4)-β-D-glucan, also known as mixed linkage glucan (MLG), is a polysaccharide that in flowering plants is unique to the cell walls of grasses and other related members of Poales. MLG is highly abundant in endosperm cell walls, where it is considered a storage carbohydrate. In vegetative tissues, MLG transiently accumulates in the primary cell walls of young, elongating organs. In evolutionary distant species such as Equisetum, MLG accumulates predominantly in old tissues in the stems. Similarly, we have recently shown that rice accumulates a large amount of MLG in mature stems, which prompted us to re-evaluate the hypothesis that MLG is solely related to growth in grass vegetative tissues. Here, we summarize data that confirms the presence of MLG in secondary cell walls and mature tissues in rice and other grasses. Along with these results, we discuss additional evidence indicating a broader role for MLG than previously considered.

Plants benefit greatly from silicon (Si) absorption provided that they contain Si transporters. The latter have recently been identified in the roots of some higher plants known to accumulate high concentrations of Si, and all share a high level of sequence identity. In this study, we searched for transporters in the primitive vascular plant Equisetum arvense (horsetail), which is a valuable but neglected model plant for the study of Si absorption, as it has one of the highest Si concentrations in the plant kingdom. Our initial attempts to identify Si transporters based on sequence homology with transporters from higher plants proved unsuccessful, suggesting a divergent structure or property in horsetail transporters. Subsequently, through sequencing of the horsetail root transcriptome and a search using amino acid sequences conserved in plant aquaporins, we were able to identify a multigene family of aquaporin Si transporters. Comparison of known functional domains and phylogenetic analysis of sequences revealed that the horsetail proteins belong to a different group than higher-plant Si transporters. In particular, the newly identified proteins contain a STAR pore as opposed to the GSGR pore common to all previously identified Si transporters. In order to determine its functionality, the proteins were heterologously expressed in both Xenopus oocytes and Arabidopsis, and the results showed that the horsetail proteins are extremely efficient a transporting Si. These findings offer new insights into the elusive properties of Si and its absorption by plants., (© 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.)

Background and Aims: Horsetails (Equisetopsida) diverged from other extant eusporangiate monilophytes in the Upper Palaeozoic. They are the only monilophytes known to contain the hemicellulose mixed-linkage (1 → 3, 1 → 4)-β-d-glucan (MLG), whereas all land plants possess xyloglucan. It has been reported that changes in cell-wall chemistry often accompanied major evolutionary steps. We explored changes in hemicelluloses occurring during Equisetum evolution., Methods: Hemicellulose from numerous monilophytes was treated with lichenase and xyloglucan endoglucanase. Lichenase digests MLG to di-, tri- and tetrasaccharide repeat-units, resolvable by thin-layer chromatography., Key Results: Among monilophytes, MLG was confined to horsetails. Our analyses support a basal trichotomy of extant horsetails: MLG was more abundant in subgenus Equisetum than in subgenus Hippochaete, and uniquely the sister group E. bogotense yielded almost solely the tetrasaccharide repeat-unit (G4G4G3G). Other species also gave the disaccharide, whereas the trisaccharide was consistently very scarce. Tetrasaccharide : disaccharide ratios varied interspecifically, but with no consistent difference between subgenera. Xyloglucan was scarce in Psilotum and subgenus Equisetum, but abundant in subgenus Hippochaete and in the eusporangiate ferns Marattia and Angiopteris; leptosporangiate ferns varied widely. All monilophytes shared a core pattern of xyloglucan repeat-units, major XEG products co-chromatographing on thin-layer chromatography with non-fucosylated hepta-, octa- and nonasaccharides and fucose-containing nona- and decasaccharides., Conclusions: G4G4G3G is the ancestral repeat-unit of horsetail MLG. Horsetail evolution was accompanied by quantitative and qualitative modification of MLG; variation within subgenus Hippochaete suggests that the structure and biosynthesis of MLG is evolutionarily plastic. Xyloglucan quantity correlates negatively with abundance of other hemicelluloses; but qualitatively, all monilophyte xyloglucans conform to a core pattern of repeat-unit sizes.

Primary cell walls from plants are composites of cellulose tethered by cross-linking glycans and embedded in a matrix of pectins. Cell wall composition varies between plant species, reflecting in some instances the evolutionary distance between them. In this work the monosaccharide compositions of isolated primary cell walls of nine fern species and one lycophyte were characterized and compared with those from Equisetum and an angiosperm dicot. The relatively high abundance of mannose in these plants suggests that mannans may constitute the major cross-linking glycan in the primary walls of pteridophytes and lycophytes. Pectin-related polysaccharides contained mostly rhamnose and uronic acids, indicating the presence of rhamnogalacturonan I highly substituted with galactose and arabinose. Structural and fine-structural analyses of the hemicellulose fraction of leaves of Adiantum raddianum confirmed this hypothesis. Linkage analysis showed that the mannan contains mostly 4-Man with very little 4,6-Man, indicating a low percentage of branching with galactose. Treatment of the mannan-rich fractions with endo-β-mannanase produced characteristic mannan oligosaccharides. Minor amounts of xyloglucan and xylans were also detected. These data and those of others suggest that all vascular plants contain xyloglucans, arabinoxylans, and (gluco)mannans, but in different proportions that define cell wall types. Whereas xyloglucan and pectin-rich walls define Type I walls of dicots and many monocots, arabinoxylans and lower proportion of pectin define the Type II walls of commelinoid monocots. The mannan-rich primary walls with low pectins of many ferns and a lycopod indicate a fundamentally different wall type among land plants, the Type III wall., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background and Aims: The anatomy of Equisetum stems is characterized by the occurrence of vallecular and carinal canals. Previous studies on the carinal canals in several Equisetum species suggest that they convey water from one node to another., Methods: Cell wall composition and ultrastructure have been studied using immunocytochemistry and electron microscopy, respectively. Serial sectioning and X-ray computed tomography were employed to examine the internode-node-internode transition of Equisetum ramosissimum., Key Results: The distribution of the LM1 and JIM20 extensin epitopes is restricted to the lining of carinal canals. The monoclonal antibodies JIM5 and LM19 directed against homogalacturonan with a low degree of methyl esterification and the CBM3a probe recognizing crystalline cellulose also bound to this lining. The xyloglucan epitopes recognized by LM15 and CCRC-M1 were only detected in this lining after pectate lyase treatment. The carinal canals, connecting consecutive rings of nodal xylem, are formed by the disruption and dissolution of protoxylem elements during elongation of the internodes. Their inner surface appears smooth compared with that of vallecular canals., Conclusions: The carinal canals in E. ramosissimum have a distinctive lining containing pectic homogalacturonan, cellulose, xyloglucan and extensin. These canals might function as water-conducting channels which would be especially important during the elongation of the internodes when protoxylem is disrupted and the metaxylem is not yet differentiated. How the molecularly distinct lining relates to the proposed water-conducting function of the carinal canals requires further study. Efforts to elucidate the spatial and temporal distribution of cell wall polymers in a taxonomically broad range of plants will probably provide more insight into the structural-functional relationships of individual cell wall components or of specific configurations of cell wall polymers.

Background: The horsetails (Equisetum sp) are known biosilicifiers though the mechanism underlying silica deposition in these plants remains largely unknown. Tissue extracts from horsetails grown hydroponically and also collected from the wild were acid-digested in a microwave oven and their silica 'skeletons' visualised using the fluor, PDMPO, and fluorescence microscopy., Results: Silica deposits were observed in all plant regions from the rhizome through to the stem, leaf and spores. Numerous structures were silicified including cell walls, cell plates, plasmodesmata, and guard cells and stomata at varying stages of differentiation. All of the major sites of silica deposition in horsetail mimicked sites and structures where the hemicellulose, callose is known to be found and these serendipitous observations of the coincidence of silica and callose raised the possibility that callose might be templating silica deposition in horsetail. Hydroponic culture of horsetail in the absence of silicic acid resulted in normal healthy plants which, following acid digestion, showed no deposition of silica anywhere in their tissues. To test the hypothesis that callose might be templating silica deposition in horsetail commercially available callose was mixed with undersaturated and saturated solutions of silicic acid and the formation of silica was demonstrated by fluorimetry and fluorescence microscopy., Conclusions: The initiation of silica formation by callose is the first example whereby any biomolecule has been shown to induce, as compared to catalyse, the formation of silica in an undersaturated solution of silicic acid. This novel discovery allowed us to speculate that callose and its associated biochemical machinery could be a missing link in our understanding of biosilicification.

Equisetum arvense L. (Equisetaceae-horsetail) accumulates various flavones and flavonols in infertile shoot. Enzyme assays conducted with crude extracts of the green tissue revealed chalcone synthase activity and also three further activities assigned to flavonoid biosynthesis and identified as flavone synthase I, flavanone 3β-hydroxylase and flavonol synthase. The latter three activities were characterized as soluble, 2-oxoglutarate-dependent dioxygenases by their typical cofactor requirements and peculiar inhibition. Notably, this is the first report of flavone synthase I which had been considered to be restricted solely to species of the Apiaceae from a distant plant taxon., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

• The very high rates of convective ventilation reported recently in Equisetum telmateia (up to 120 cm(3) min(-1); internal wind speed, 10 cm s(-1)) prompted this study of a further eight species for the presence or absence of convection and the possible reasons for this. • Convection rates were examined in relation to anatomical pathways, internal resistance to applied pressurized gas flow and stomata. • Only species with interconnecting cortical aerenchyma in branches (when present), shoots and rhizomes induced convection. Rapid humidity-induced convection (HIC) occurred in E. palustre (up to 13 cm(3) min(-1)), with slower rates in E. × schaffneri and E. ramosissimum (≤ 6 and 3 cm(3) min(-1), respectively). Excised shoots of E. hyemale and E. fluviatile showed the potential for HIC (≤ 0.5 and 0.15 cm(3) min(-1), respectively), but not into the rhizomes. High rates were linked to low internal gas flow resistance. No convection was detected in E. scirpoides, E. sylvaticum or E. arvense due to the extremely high resistance to pressure flow, for example, from intercalary meristems and, in the last two, to nonaerenchymatous branches. • Of the nine Equisetum species studied so far, four showed through-flow convection; the other species must rely solely on diffusion for underground aeration in wet soils., (© 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.)

Significant pressurized (convective) ventilation has been demonstrated in some flowering wetland plants, for example water-lilies and reeds, but not previously in nonflowering plants. Here we investigated convective flows in the great horsetail, Equisetum telmateia, and the possibility that convections aerated the massive rhizomes of the Calamites, extinct giant horsetails of the Carboniferous. Convection in E. telmateia was examined in relation to induction sites, anatomical pathways, relative humidity (RH), external wind-speed, diurnal effects, rhizome resistance and pressure-gradients. A mathematical model, incorporating Calamite aeration anatomy, was applied in assessing potentials for convective aeration. Individual shoots of E. telmateia generated extremely high rates of humidity-induced convection: < or = 120 cm(3) min(-1) (internal wind-velocity: 10 cm s(-1)) with rates proportional to branch numbers and 1/RH. Flows passed through branches, stem and rhizome via low-resistance lacunae (vallecular canals) and vented via stubble. Stomata supported internal pressures up to 800 Pa. Anatomically, E. telmateia resembles the Calamites and modelling predicted possible flows of 70 l min(-1) per Calamite tree. This is the first demonstration of significant convective flow in a nonflowering species, indicating that plant ventilation by a type of 'molecular gas-pump' may date back 350 million yr or more. Stomatal form and low-resistance pathways may facilitate high flow rates.

Mixed-linkage (1-->3),(1-->4)-beta-D-glucan (MLG) is widely considered to be a defining feature of the cell walls of plants in the Poales order. However, we conducted an extensive survey of cell-wall composition in diverse land plants and discovered that MLG is also abundant in the walls of the horsetail Equisetum arvense. MALDI-TOF MS and monosaccharide linkage analysis revealed that MLG in E. arvense is an unbranched homopolymer that consists of short blocks of contiguous 1,4-beta-linked glucose residues joined by 1,3-beta linkages. However, in contrast to Poaceae species, MLG in E. arvense consists mostly of cellotetraose rather than cellotetriose, and lacks long 1,4-beta-linked glucan blocks. Monosaccharide linkage analyses and immunochemical profiling indicated that, in E. arvense, MLG is a component of cell walls that have a novel architecture that differs significantly from that of the generally recognized type I and II cell walls. Unlike in type II walls, MLG in E. arvense does not appear to be co-extensive with glucuroarabinoxylans but occurs in walls that are rich in pectin. Immunofluorescence and immunogold localization showed that MLG occurs in both young and old regions of E. arvense stems, and is present in most cell types apart from cells in the vascular tissues. These findings have important implications for our understanding of cell-wall evolution, and also demonstrate that plant cell walls can be constructed in a way not previously envisaged.

Equisetaceae has been of research interest for decades, as it is one of the oldest living plant families, and also due to its high accumulation of silica up to 25% dry wt. Aspects of silica deposition, its association with other biomolecules, as well as the chemical composition of the outer strengthening tissue still remain unclear. These questions were addressed by using high resolution (<1 microm) Confocal Raman microscopy. Two-dimensional spectral maps were acquired on cross sections of Equisetum hyemale and Raman images calculated by integrating over the intensity of characteristic spectral regions. This enabled direct visualization of differences in chemical composition and extraction of average spectra from defined regions for detailed analyses, including principal component analysis (PCA) and basis analysis (partial least square fit based on model spectra). Accumulation of silica was imaged in the knobs and in a thin layer below the cuticula. In the spectrum extracted from the knob region as main contributions, a broad band below 500 cm(-1) attributed to amorphous silica, and a band at 976 cm(-1) assigned to silanol groups, were found. From this, we concluded that these protrusions were almost pure amorphous, hydrated silica. No silanol group vibration was detected in the silicified epidermal layer below and association with pectin and hemicelluloses indicated. Pectin and hemicelluloses (glucomannan) were found in high levels in the epidermal layer and in a clearly distinguished outer part of the hypodermal sterome fibers. The inner part of the two-layered cells revealed as almost pure cellulose, oriented parallel along the fiber.

Mixed-linkage (1-->3,1-->4)-beta-d-glucan (MLG) is a hemicellulose reputedly confined to certain Poales. Here, the taxonomic distribution of MLG, and xyloglucan, especially in early-diverging pteridophytes, has been re-investigated. Polysaccharides were digested with lichenase and xyloglucan endoglucanase (XEG), which specifically hydrolyse MLG and xyloglucan, respectively. The oligosaccharides produced were analysed by thin-layer chromatography (TLC), high-pressure liquid chromatography (HPLC) and alkaline peeling. Lichenase yielded oligo-beta-glucans from all Equisetum species tested (Equisetum arvense, Equisetum fluviatile, Equisetum scirpoides, Equisetum sylvaticum and Equisetum xtrachyodon). The major product was the tetrasaccharide beta-glucosyl-(1-->4)-beta-glucosyl-(1-->4)-beta-glucosyl-(1-->3)-glucose (G4G4G3G), which was converted to cellotriose by alkali, confirming its structure. Minor products included G3G, G4G3G and a nonasaccharide. By contrast, poalean MLGs yielded G4G3G > G4G4G3G > nonasaccharide > dodecasaccharide. No other pteridophytes tested contained MLG, including Psilotum and eusporangiate ferns. No MLG was found in lycopodiophytes, bryophytes, Chara or Nitella. XEG digestion showed that Equisetum xyloglucan has unusual repeat units. Equisetum, an exceedingly isolated genus whose closest living relatives diverged > 380 million years ago, has evolved MLG independently of the Poales. Equisetum and poalean MLGs share basic structural motifs but also exhibit clear-cut differences. Equisetum MLG is firmly wall-bound, and may tether neighbouring microfibrils. It is also suggested that MLG acts as a template for silica deposition, characteristic of grasses and horsetails.

Horsetail (Equisetum spp.) is known as one of the strongest accumulators of silicon among higher terrestrial plants. We use the combination of position-resolved analytical techniques, namely microtomography, energy-dispersive X-Ray elemental mapping, Raman microscopy, as well as small-angle and wide-angle scattering of X-rays, to study the type, distribution and nanostructure of silica in the internodes of Equisetum hyemale. The predominant silicification pattern is a thin continuous layer on the entire outer epidermis with the highest density in particular knob regions of the long epidermal cells. The knob tips contain up to 33 wt% silicon in the form of pure hydrated amorphous silica, while the silica content is lower in the inner part of the knobs and on the continuous layer. In contrast to the knob tips, the silica in these regions lacks silanol groups and is proposed to be in close association with polysaccharides. No mentionable amount of crystalline silica is detected by wide-angle X-ray scattering. The small-angle X-ray scattering data are consistent with the presence of colloidal, sheet-like silica agglomerates with a thickness of about 2 nm. From these results we conclude that there are at least two distinct forms of silica in E. hyemale which may have different functions. The close association of silica with cell wall polymers suggests that they may act as a polymeric template that controls the shape and size of the colloidal silica particles similar to many other biominerals and mineralised tissues. We propose that owing to its specific distribution in E. hyemale, a protective role and possibly also an important biomechanical role are among the most likely functions of silica in these plants.

In many organisms, meiotic chromosomes are bundled at their telomeres to form a "bouquet" arrangement. The bouquet formation plays an important role in homologous chromosome pairing and therefore progression of meiosis. As meiotic telomere clustering occurs in response to mating pheromone signaling in fission yeast, we looked for factors essential for bouquet formation among genes induced under mating pheromone signaling. This genome-wide search identified two proteins, Bqt1 and Bqt2, that connect telomeres to the spindle-pole body (SPB; the centrosome equivalent in fungi). Neither Bqt1 nor Bqt2 alone functions as a connector, but together the two proteins form a bridge between Rap1 (a telomere protein) and Sad1 (an SPB protein). Significantly, when both Bqt1 and Bqt2 are ectopically expressed in mitotic cells, they also form a bridge between Rap1 and Sad1. Thus, a complex including Bqt1 and Bqt2 is essential for connecting telomeres to the SPB.

The study with pot culture experiment showed that Equisetum ramosissirmunm did not appear obvious poisoning symptoms when treated with low concentration Cu (500 mg x kg(-1)), while serious injuries were found when treated with high concentration Cu (1000 to approximately 3000 mg x kg(-1)), which reflected in the severe damage of cell membrane and cytoarchitecture as well as the structure and function of main organelles, and the significant decrease of the contents of leaf chlorophyll a and b and stem soluble monosaccharose. The cell membrane osmolarity and the average MDA content of the plant exposed to heavy copper pollution was 1 to approximately 2 and 1 to approximately 3 times greater than the control, respectively. It could be concluded that high concentration Cu disturbed the physiological metabolism, and critically threatened the normal growth of E. ramosissimum. The activities of protective enzyme, especially of SOD and POD, were enhanced with increasing Cu concentration, and had a positive correlation with Cu concentration (rPOD = 0.978, rSOD = 0.926, P < 0.05).

The intricate siliceous architectures of diatom species have inspired our exploration of biosilicification. In vitro studies of natural systems within the area of silica biosynthesis are complicated. Previous studies, which included biomimetic approaches, often failed to recognize the chemistry of silicic acid and its analogues. To better understand the role of various proteins in the biosilicification process, recent studies have been conducted to test the ability of enzymes to catalyze the formation and cleavage of siloxane bonds. Notably, biocatalysis at silicon was observed. Further understanding of the biotransformation strategy in the design and synthesis of structurally complex materials would be beneficial.

We have determined the crystal structure, at 1.2-A resolution, of Equisetum arvense ferredoxin isoform II (FdII), which lacks residues equivalent to Arg(39) and Glu(28) highly conserved among other ferredoxins (Fds). In other Fds these residues form an intramolecular salt bridge crucial for stabilization of the [2Fe-2S] cluster, which is disrupted upon complex formation with Fd-NADP(+) oxidoreductase (FNR) to form two intermolecular salt bridges. The overall structure of FdII resembles the known backbone structures of E. arvense isoform I (FdI) and other plant-type Fds. Dramatically, in the FdII structure a unique, alternative salt bridge is formed between Arg(22) and Glu(58). This results in a different relative orientation of the alpha-helix formed by Leu(23)-Glu(29) and eliminates the possibility of forming three of the five intermolecular salt bridges identified on formation of a complex between maize FdI and maize FNR. Mutation of FdII, informed by structural differences with FdI, showed that the alternative salt bridge and the absence of an otherwise conserved Tyr residue are important for the alternative stabilization of the FdII [2Fe-2S] cluster. We also investigated FdI and FdII electron transfer to FNR on chloroplast thylakoid membranes. The K(m) and V(max) values of FdII are similar to those of FdI, contrary to previous measurements of the reverse reaction, from FNR to Fd. The affinity between reduced FdI and oxidized FNR is much greater than that between oxidized FdI and reduced FNR, whereas this is not the case with FdII. The pH dependence of electron transfer by FdI, FdII, and an FdII mutant with FdI features was measured and further indicated that the binding mode to FNR differs between FdI and FdII. Based on this evidence, we hypothesize that binding modes with other Fd-dependent reductases may also vary between FdI and FdII. The structural differences between FdI and FdII therefore result in functional differences that may influence partitioning of electrons into different redox metabolic pathways.

Phenomenon of autofluorescence from vegetative microspores of spore-breding plant Equisetum arvense has been studied by methods of laser-scanning confocal microscopy (LSCM) and microspectrofluorimetry during the development of the cells. The microspores have demonstrated a difference between structures: blue-fluorescing cover and red-fluorescing chloroplasts. The fluorescence spectra of the studied cells was also measured by original microspectrofluorimeter. The character of the spectra and the color of fluorescence was changed during the microspores germination. The red fluorescence of the microspores was, mainly, due to the presence of chlorophyll and azulenes. The unicellular microspores may be recommended as natural probes of cellular viability and development.

Effects of shading on the growth of Equisetum arvense during the growing season were studied in terms of the dynamics of non-structural carbohydrates (starch, sucrose, glucose). Tubers of 0.04 g dry mass were planted in pots. Plants were cultivated under different radiation conditions (100%, 3%, 100%-->3%, and 100%-->3%-->100%). The carbohydrate concentration in belowground parts responded sharply to the irradiance conditions. Under 3% relative photon flux density (PFD), they could not grow beyond the initial mass and decayed. Dry mass per length of rhizomes was highly correlated with the starch concentration, which was correlated with the mass of current tubers. The rhizomes of low starch concentration did not form current tubers. The carbohydrate concentration of rhizomes increased when the plants were transferred from 3% relative PFD to 100% irradiance conditions. After ca. 2 months of improved PFD, they had the same content of non-structural carbohydrates as the 100% PFD plants.

Carbohydrate specificity of partially purified lectins from 4 species of plants: horse-tail genus Equisetum (Equisetum arvense L., E. sylvaticum L., E. hyemale L. and E. tempatelia Ehrh.) has been studies. The obtained lectins have similar carbohydrate specificity. Among the tested carbohydrates the best inhibitor of activity is phenyl-2-acetamido-alpha-D-glucosaminopyranoside. Lectins poorly interact with yeasty mannan and galactomannan Trigonella foenum graecum seeds. Among glycoproteins the best inhibitor of activity is ovomucoid.