Free galactose and galactosidase activity in the course of flax fiber development

Composition and content of free monosaccharides and β-galactosidase activity were determined in the course of development of the flax (Linum usitatissimum L.) tissues containing bast fibers. In the stem regions where the fibers were at the stage of formation of the secondary cell wall of gelatinous type, the content of free galactose was high (14 mM) and 13–20 times greater than in the upper part of the stem where the fibers were at the stage of intrusive growth. Pulse-chase experiments demonstrated the differences in the metabolism of individual low-molecular sugars. In respect to glucose and sucrose, all the examined characteristics (content, absolute and specific radioactivity, and the temporal changes of these indices) were identical in the stem regions wherein the fibers were at different stages of development. Labeled galactose was detected only in the stem regions where the fibers were at the stage of secondary cell wall formation. The specific radioactivity of glucose and sucrose reached the maximum immediately after photosynthesis in the presence of ¹⁴CO₂ and changed in the same way as in the primary products of photosynthesis. The time-course of label incorporation into galactose indicated that this monosaccharide arose as a result of hydrolytic processes. At the stage of secondary cell wall formation, high activity of β-galactosidase was observed, with tissue- and stage-specific fiber β-1,4-galactan as its substrate.     

Variability in the composition of tissue-specific galactan from flax fibers

Tissue-specific galactan of sclerenchyma fibers, with cell walls of the gelatinous type, was examined in flax plants (Linum usitatissimum L.) of 23 various genotypes. The content and average degree of polymerization of side chains of galactan were estimated before its deposition into the cell wall. The variability of the analyzed parameters of tissue-specific galactan from flax fibers was high; within the same genotype, the scope of paratypic variability between replicates and years of research was comparable to variability between different genotypes. The average length of side chains in the studied samples ranged from 5 to 41 galactose residues. The average degrees of polymerization of galactan side chains in flax fibers was found to be discrete, which could be explained by block assemblage of the polymer in the Golgi apparatus.     

Plant fiber intrusive growth characterized by NMR method

Intrusively growing plant cells insert themselves between surrounding cells, thus increasing the number of membranes on the tissue cross-section. This parameter can be assessed by spin echo NMR method with a magnetic field pulse gradient. Diffusion echo decay was measured for stem regions of long-fiber flax (Linum usitatissimum L.) differing in the stages of primary fiber development, which elongate thousand-fold during intrusive growth. Additionally, the number of fibers on stem cross-sections was counted under microscope. An increase in the slow component of the echo diffusion decay was correlated with an increase in the number of fibers on the stem cross-section in the zone of intrusive growth, while other stem-structure characteristics remained unchanged. Thus, NMR method can be used for characterization of intrusive fiber growth in situ.     

Structural characterization of tissue-specific galactan from flax fibers by 1H NMR and MALDI TOF mass spectrometry

A high-molecular-mass polysaccharide galactan (M 2000 kDa) was isolated from flax at the stage of cell wall thickening of the bast fiber development. The polymer structure was studied by ¹H NMR spectroscopy and MALDI TOF mass spectrometry. It is built up of Gal (59%), Rha (15%), GalA (23%), and Ara (3%) residues. The galactan backbone consists of successively alternating monomer disaccharide units (→ 4GalA1 → 2Rha1 →)_n and is similar in its structure to the backbone of rhamnogalacturonan-1 (RG-I). Rhamnose residues bear in position 4 β-(1 → 4)-galactose side chains of various lengths with a polymerization degree of up to 28 or higher. A part of the side chains have branchings.     

Secondary cell-wall assembly in flax phloem fibres: role of galactans

Non-lignified fibre cells (named gelatinous fibres) are present in tension wood and the stems of fibre crops (such as flax and hemp). These cells develop a very thick S2 layer within the secondary cell wall, which is characterised by (1) cellulose microfibrils largely parallel to the longitudinal axis of the cell, and (2) a high proportion of galactose-containing polymers among the non-cellulosic polysaccharides. In this review, we focus on the role of these polymers in the assembly of gelatinous fibres of flax. At the different stages of fibre development, we analyse in detail data based on sugar composition, linkages of pectic polymers, and immunolocalisation of the β-(1→4)-galactans. These data indicate that high molecular-mass gelatinous galactans accumulate in specialised Golgi-derived vesicles during fibre cell-wall thickening. They consist of RG-I-like polymers with side chains of β-(1→4)-linked galactose. Most of them are short, but there are also long chains containing up to 28 galactosyl residues. At fibre maturity, two types of cross-linked galactans are identified, a C–L structure that resembles the part of soluble galactan with long side chains and a C–S structure with short chains. Different possibilities for soluble galactan to give rise to C–L and C–S are analysed. In addition, we discuss the prospect for the soluble galactan in preventing the newly formed cellulose chains from completing immediate crystallisation. This leads to a hypothesis that firstly the secretion of soluble galactans plays a role in the axial orientation of cellulose microfibrils, and secondly the remodelling and cross-linking of pectic galactans are linked to the dehydration and the assembly of S2 layer.     

The influence of ammoniates on 14CO2 assimilation in flax

A 1 μM solution of ammoniates [ZnCu(NH₃)_n]²⁺(CO₃)²⁻ was inserted into a cut shoot of flax with the transpiration stream of water. Analysis of the ¹⁴C content after ¹⁴CO₂ assimilation by the shoot showed that ammoniates increased radioactive label contents in the tissues (especially in the young leaves and stem). In the leaves the higher sucrose to hexoses ratio, an increased radioactivity of glycerate and malate and decreased incorporation of ¹⁴C into oligosaccharides and pigments were observed. These effects were more pronounced in the young leaves. Spraying of plants with 20 mM solution resulted in an increase of plant height and leaf number.     

Effect of a Bacillus subtilis strain on flax protection against Fusarium oxysporum and its impact on the root and stem cell walls

In Normandy, flax is a plant of important economic interest because of its fibres. Fusarium oxysporum, a telluric fungus, is responsible for the major losses in crop yield and fibre quality. Several methods are currently used to limit the use of phytochemicals on crops. One of them is the use of plant growth promoting rhizobacteria (PGPR) occurring naturally in the rhizosphere. PGPR are known to act as local antagonists to soil‐borne pathogens and to enhance plant resistance by eliciting the induced systemic resistance (ISR). In this study, we first investigated the cell wall modifications occurring in roots and stems after inoculation with the fungus in two flax varieties. First, we showed that both varieties displayed different cell wall organization and that rapid modifications occurred in roots and stems after inoculation. Then, we demonstrated the efficiency of a Bacillus subtilis strain to limit Fusarium wilt on both varieties with a better efficiency for one of them. Finally, thermo‐gravimetry was used to highlight that B. subtilis induced modifications of the stem properties, supporting a reinforcement of the cell walls. Our findings suggest that the efficiency and the mode of action of the PGPR B. subtilis is likely to be flax variety dependent.     

Accumulation of cadmium and selected elements in flax seed grown on a calcareous soil

Seed of flax (Linum usitatissimum L.) grown on calcareous and neutral soils sometimes accumulates relatively high concentrations of Cd. The influence of a post-flowering application of NH₄NO₃ (115 mg N kg^-1), CdSO₄ (1 mg Cd kg^-1), FeEDDHA (2 mg Fe kg^-1), NaH₂PO₄ (120 mg P kg^-1) and ZnSO₄ (8 mg Zn kg^-1) on seed accumulation of Cd, Fe, N, Mn, P and Zn by flax grown on a Calciaquoll was studied in two experiments under greenhouse conditions. Seed yields were increased by the N and Zn treatments, and the N×Zn interaction was positive. Zinc deficiency delayed flowering and boll formation by up to 20 days and reduced seed size. In the absence of added Cd, seed accumulated up to 0.33 mg Cd kg^-1. This Cd accumulation was reduced by approximately 50 and 17% by added Zn and Fe, respectively, but was little affected by P fertilizer and post-flowering N stress. In the presence of added Cd, seed Cd exceeded 3.3 mg Cd kg^-1, and the antagonistic effects of Fe and Zn on seed Cd were absent. Seed N, P, Fe and Zn concentrations were increased on average by 10, 45, 31 and 97% by the N, P, Fe and Zn fertilizer treatments, respectively. FeEDDHA reduced seed Mn concentration by approximately 58%. However, seed Mn concentration was much less than that found in vegetative tissue at flowering. Soil-applied Zn may reduce seed Cd concentration in flax under field conditions, and may increase marketability of flax for food use.     

亚麻响应盐、碱胁迫的生理特征

利用中性盐NaCl、Na₂SO₄和碱性盐NaHCO₃、Na₂CO₃混合模拟不同强度的盐、碱胁迫条件, 对亚麻(Linum usitatissimum)进行14天胁迫处理, 测定其地上部分和根生长速率、光合特征、离子平衡及有机渗透调节物质积累, 以探讨亚麻对盐、碱两种胁迫的生理响应特点。研究表明: 亚麻生长对盐、碱胁迫的响应存在差异, 在相同盐浓度下, 碱胁迫对亚麻的伤害大于盐胁迫。碱胁迫使地上部分中Na⁺浓度急剧增高, 造成叶绿体破坏、光合色素含量下降, 光合能力及碳同化能力也急剧下降。亚麻中Na⁺含量随着胁迫强度的增加而升高, 而K⁺含量呈下降趋势, 碱胁迫下的变化明显大于盐胁迫。因此, 碱胁迫导致Na⁺过度积累可能是碱胁迫对植物伤害大于盐胁迫的最主要原因。碱胁迫下Ca²⁺和Mg²⁺在根中下降明显, 可见高pH值阻碍根对Ca²⁺和Mg²⁺的吸收。Fe²⁺和Zn²⁺对渗透调节的影响不大, 因为它们的离子含量较低。盐胁迫促进阴离子(Cl^-、H₂PO₄^-和SO₄^2-)的积累来平衡大量涌入的Na⁺, 但是碱胁迫明显减少无机阴离子含量, 可能造成严重营养胁迫(如P和S不足)。亚麻在盐胁迫下积累大量可溶性糖来平衡大量的Na⁺, 但碱胁迫下积累大量有机酸来维持细胞内离子平衡和pH值稳定, 碱胁迫大量积累的有机酸也可能被分泌到根外调节根外的pH值, 这说明亚麻对两种不同胁迫的响应方式不同。研究证明高pH值会直接影响植物根系的生长发育, 影响植物矿质元素的吸收, 阻碍离子稳态重建, 有机酸代谢是亚麻碱胁迫下的关键适应机制。     

Polysaccharides, tightly bound to cellulose in cell wall of flax bast fibre: Isolation and identification

Part of matrix polymers of flax bast fibre cell wall is tightly bound to cellulose and can not be extracted by conventional methods. To analyze these polymers, the residue, remaining after cell wall treatment with chelators and alkali, was dissolved in solution of lithium chloride in N,N-dimethylacetamide. Cellulose was precipitated by water and completely degraded by cellulase, giving the possibility to separate matrix polysaccharides, which remained in polymeric form. The obtained polymers were fractionated by gel permeation chromatography and characterized by monosaccharide analysis, staining with LM5 antibody and Yariv reagent, ¹H and ¹³C NMR. The total yield of the polysaccharides that are tightly bound to cellulose in flax fibre, was 4.6%. The major fractions (molecular mass 100–400 kDa) were composed of galactose, accompanied by two other significant monomers, GalA and Rha, with the ratio 1.1–1.4. Composition and structure of the cellulose bound galactan permit to consider it as fragment of the high-molecular mass (2000 kDa) galactan, synthesized by the developing fibres, while forming the secondary cell wall of gelatinous type.     

Biogenesis of plant fibers

The fiber (in terms of plant biology) is an individual cell characterized by spindle shape, length of up to several centimeters, well developed cell wall, and mechanical function. The review summarizes different, sometimes contradictory view points about duration, segregation and mechanisms of realization of individual stages of fiber biogenesis. Initiation and coordinated and intrusive growth are considered, as well as formation of secondary cell wall, including its gelatinous layers, and senescence. Biogenesis of fibers ontogenetically related to various tissues has been analyzed and the data about marker stage-specific characters of these cells. The data summarized in this review will allow not only deeper understanding the development of cells with such unique characters, but also interpret the growth mechanisms for much more cell types, in which it is more difficult to identify individual stages of biogenesis than in the sclerenchyma fibers.     

Plant Fiber Formation: State of the Art,Recent and Expected Progress,and Open Questions

Plant fibers are one of the most important renewable resources, used as raw material in the paper industry, and for various textiles and for composites. Fibers are structural components in timber and an energy-rich component of fuel-wood. For the plant itself, fibers are important in establishing plant architecture, as a source of mechanical support, in defence from herbivory, and in some cases as elements with contractile properties, resembling those of muscles. In addition, fibers may store ergastic carbon resources and water. Here, we review various aspects of fiber development such as initiation, elongation, cell wall formation and multinuclearity, discuss open questions and propose directions for further research. Most of the recent progress in fiber formation biology, especially in cell wall structure and chemistry, emerged from studies of only a few model plants including flax, Populus spp., Eucalyptus spp., Arabidopsis thaliana and hemp. Considering the enormous importance of fibers to humanity, it is surprising how little is known about the biology of fiber formation.     

GIGANTEA is a component of a regulatory pathway determining wall ingrowth deposition in phloem parenchyma transfer cells of Arabidopsis thaliana

Transfer cells are specialised transport cells containing invaginated wall ingrowths that generate an amplified plasma membrane surface area with high densities of transporter proteins. They trans‐differentiate from differentiated cells at sites at which enhanced rates of nutrient transport occur across apo/symplasmic boundaries. Despite their physiological importance, little is known of the molecular mechanisms regulating construction of their intricate wall ingrowths. We investigated the genetic control of wall ingrowth formation in phloem parenchyma transfer cells of leaf minor veins in Arabidopsis thaliana. Wall ingrowth development in these cells is substantially enhanced upon exposing plants to high‐light or cold treatments. A hierarchical bioinformatic analysis of public microarray datasets derived from the leaves of plants subjected to these treatments identified GIGANTEA (GI) as one of 46 genes that are commonly up‐regulated twofold or more under both high‐light and cold conditions. Histological analysis of the GI mutants gi‐2 and gi‐3 showed that the amount of phloem parenchyma containing wall ingrowths was reduced 15‐fold compared with wild‐type. Discrete papillate wall ingrowths were formed in gi‐2 plants but failed to develop into branched networks. Wall ingrowth development in gi‐2 was not rescued by exposing these plants to high‐light or cold conditions. In contrast, over‐expression of GI in the gi‐2 background restored wall ingrowth deposition to wild‐type levels. These results indicate that GI regulates the ongoing development of wall ingrowth networks at a point downstream of inputs from environmental signals.     

Impact of interspecific competition and drought on the allocation of new assimilates in trees

In trees, the interplay between reduced carbon assimilation and the inability to transport carbohydrates to the sites of demand under drought might be one of the mechanisms leading to carbon starvation. However, we largely lack knowledge on how drought effects on new assimilate allocation differ between species with different drought sensitivities and how these effects are modified by interspecific competition. We assessed the fate of ¹³C labelled assimilates in above‐ and belowground plant organs and in root/rhizosphere respired CO₂ in saplings of drought‐tolerant Norway maple (Acer platanoides) and drought‐sensitive European beech (Fagus sylvatica) exposed to moderate drought, either in mono‐ or mixed culture. While drought reduced stomatal conductance and photosynthesis rates in both species, both maintained assimilate transport belowground. Beech even allocated more new assimilate to the roots under moderate drought compared to non‐limited water supply conditions, and this pattern was even more pronounced under interspecific competition. Even though maple was a superior competitor compared to beech under non‐limited soil water conditions, as indicated by the changes in above‐ and belowground biomass of both species in the interspecific competition treatments, we can state that beech was still able to efficiently allocate new assimilate belowground under combined drought and interspecific competition. This might be seen as a strategy to maintain root osmotic potential and to prioritise root functioning. Our results thus show that beech tolerates moderate drought stress plus competition without losing its ability to supply belowground tissues. It remains to be explored in future work if this strategy is also valid during long‐term drought exposure.     

Ascorbic acid and development of xylem and phloem cells in the pine trunk

Changes in the levels of ascorbic acid (AA), its oxidized form, dehydroascorbic acid (DHA), and uronic acids as initial precursors for the AA synthesis were studied as related to the degree of xylem and phloem cell development in the course of early and late wood formation in the trunks of Scots pine (Pinus sylvestris L.). The cells of mature and conducting phloem, cambial zone, differently developed cells in the zones of cell enlargement and maturation were obtained by successive scraping tissue layers from trunk segments of 20–25-year-old trees; tissue identification was checked anatomically and histochemically. The contents of compounds tested were calculated per dry weight and per cell basis. We found great differences in the contents of AA and DHA and also in their ratio in dependence of the wood type developing in the pine trunks during growth period and on the stage of differentiation of xylem and phloem cells. Changes in the AA content during xylem cell differentiation were accompanied by changes in the content of uronic acids. The amounts of AA, DHA, and uronic acids were the highest at the stage of early lignification and reduced with tracheid maturation. The AA to DHA ratio changed differently in the course of early and late xylem lignification. It reduced from the start of lignification to the formation of early mature xylem and, in contrast, increased in mature late wood; this indicates a difference in the level of redox processes in these tissues.     

Identification and partial characterization of proteins and proteoglycans encrusting the secondary cell walls of flax fibres

 Four proteins were isolated from depectinised elementary fibres of flax (Linum usitatissimum L.), using either alkali or cellulase digestion treatments. All the four proteins were characterized by a deficiency or low contents of hydroxyproline and by high levels of glutamic acid/glutamine and/or aspartic acid/asparagine. The two proteoglycans solubilized with cellulase strongly reacted with β-glucosyl Yariv reagent but not with α-glucosyl Yariv reagent and contained appreciable amounts of alanine, glycine, serine and threonine, suggesting a relationship with cell wall hydroxyproline-deficient arabinogalactan-proteins. The two alkali-extracted proteins did not show any reaction with β-glucosyl Yariv dye. Due to the harsh treatment, they might only partially represent the original proteins. Due to its high level of glycine (41%), one of these proteins might be classified as a glycine-rich protein. The latter polypeptide, of low molecular molar mass, contained 14.6% leucine and might consist of a domain related to leucine-rich proteins. The data show that these proteins and arabinogalactan-protein-like proteoglycans were strongly associated with the secondary walls of flax fibres. Their presence in small amounts (0.1–0.4%), raises the problem of their putative structural role. Received: 22 October 1999 / Accepted: 17 January 2000     

Seasonal dynamics of phloem and xylem formation in silver fir and Norway spruce as affected by drought

The dynamics of phloem growth ring formation in silver fir (Abies alba Mill.) and Norway spruce (Picea abies Karst.) at different sites in Slovenia during the droughty growing season of 2003 was studied. We also determined the timing of cambial activity, xylem and phloem formation, and counted the number of cells in the completed phloem and xylem growth rings. Light microscopy of cross-sections revealed that cambial activity started on the phloem and xylem side simultaneously at all three plots. However, prior to this, 1–2 layers of phloem derivatives near the cambium were differentiated without previous divisions. The structure of the early phloem was similar in silver fir and Norway spruce. Differences in the number of late phloem cells were found among sites. Phloem growth rings were the widest in Norway spruce growing at the lowland site. In all investigated trees, the cambium produced 5–12 times more xylem cells than phloem ones. In addition, the variability in the number of cells in the 2003 growth ring around the stem circumference of the same tree and among different trees was higher on the xylem side than on the phloem side. Phloem formation is presumably less dependent on environmental factors but is more internally driven than xylem formation.     

Gelling agent influences the detrimental effect of kanamycin on adventitious budding in flax

Flax (Linum usitatissimum L.) hypocotyls were cultivated on regeneration media containing various concentrations of kanamycin (an aminoglycoside antibiotic commonly used to select transgenic plant material) solidified with three different gelling agents: gellan gum, agar and a mixture of both. The inhibitory effect of kanamycin on bud regeneration was analyzed. A significant interaction was observed between the nature of the gelling agent and the kanamycin concentration. The antibiotic concentration needed to strongly inhibit bud production varied greatly with the nature of the gelling agent. Gellan gum lowered the inhibitory effect of kanamycin. This revised version was published online in June 2006 with corrections to the Cover Date.     

Ultracytochemical localization of Ca2+ during the phloem ganglion development in Phyllostachys edulis

Ultracytochemical localization of Ca2+ was investigated using the potassium pyroantimonate precipitation method during the development of phloem ganglion.The result showed that Ca2+ was mainly localized in the cell wall and intercellular spaces in the initiating phase.With the development of the phloem ganglion,the distribution of Ca2+ transferred to the vacuole,and the Ca2+ deposits in the cell wall and intercellular space decreased.At the later stage of the developmental phase.Ca2+ was distributed in the tonoplast and vacuole phagocytosis,and the vacuole became the main calcium storage in this phase.At the early stage of maturation of the phloem ganglion,most of the phloem ganglion cells'vacuoles cracked,and the cytoplastic Ca2+ content increased in large number.In the mature phloem ganglion,not only were there a few Ca2+ localized in the cytoplast of mature cells,but also in the differentiating cells in the vacuoles.Ca2+ was distributed in the tonoplast and vacuole contents;initiating cells almost had no Ca2+.In general,Ca2+ concentration in mature phloem ganglion cells was at a low level.The results indicated that the changes in Ca2+ distribution evoked the phloem ganglion generation,and Ca2+ regulated the physiological function of the phloem ganglion.