The expansion of maize root-cap mucilage during hydration. 3. Changes in water potential and water content

Root-cap mucilage from aerial nodal roots of maize has been found to have water potential values of −11 MPa or lower when air dried. The value approaches 0 MPa within 2 min of hydration in distilled water. In this time the expanding gel absorbs only about 0.3% of the water content of fully expanded mucilage. It is concluded that the root-cap mucilage per se has almost no capacity to retain water in the rhizosphere. Any function that it may play in the slowing of root desiccation would be indirect. For example, mucilage might decrease pore size between and within soil aggregates by pulling the particles together in a cycle of nocturnal efflux of water from the root surface, and diumal dyring during transpiration.     

The expansion of maize root-cap mucilage during hydration. 1. Kinetics

The conventional view of root-cap mucilage as an expanded blob of mucilage is characteristic only of root tips in contact with free water. In soil, the mucilage is almost always a dry coating over the tip to which soil particles adhere. The kinetics of expansion of root-cap mucilage of Zea mays roots grown in field soil, in soil in pots, and axenically on agar, were determined when the mucilage was exposed to water. On the soil-grown roots the increase in mucilage volume was linear with time, sometimes reaching a constant volume during the 6 h of measurement, but sometimes not. This linear expansion is interpreted as limited by the rate at which the condensed mucilage in the periplasmic and intercellular spaces of the root cap passes to the exterior of the cap, expanding as fast as it arrives outside in the water. The height of the plateau is interpreted as a measure of the amount of mucilage initially present in the interior spaces. Because of the greater availability of water in the axenic roots grown on 1% agar, the mucilage was already outside the root cap, and it expanded more rapidly. It reached a final volume about 10-fold greater than that on the soil-grown roots. The volume increase was curvilinear with time. An analysis of these curves suggested that this swelling on axenic roots was a diffusion of mucilage outwards from the flanks of the root cap, and the diffusivity of the mucilage was estimated as 4 × 10^?8 cm² s^?1. The molecular radius derived from this diffusivity was 34 nm, and the estimated molecular weight was 1.6 × 10⁸ Da.     

Some water-related physical properties of maize root-cap mucilage

Abstract The dry weight (0.1%) and water potential -7 kPa) of root-cap mucilage from 3-d-old axenically grown maize seedlings have been determined. The results suggest strong gelling properties and weak water-holding capacity for the mucilage. Root tips from seedlings grown under low or high water stress were fixed by freeze-substitution. Micrographs showed that in both conditions, mucilage was secreted into the periplasmic space and extruded through the cell wall, though in dry conditions, the mucilage was tightly pressed against the root-cap surface. Histochemical and structural evidence is presented indicating chemical changes in the composition of the mucilage upon extrusion and a sharp increase in its hydration at increasing distance from the secretory cells. The possible functions of the root-cap mucilage in the rhizosphere are examined in light of these findings.     

Physical properties of axenic maize root mucilage

Root mucilage was collected from 3–4 day-old axenically-grown maize seedlings (Zea mays L. cv. Freya). The water potential of the hydrated mucilage was measured by thermocouple psychrometry and the rheology at low deformation rates was studied using an oscillating cone and plate rheometer which provides information on both the elastic and viscous components of its behaviour. Water potential decreased as mucilage solute concentration increased, reaching a value of −60kPa at 1.2 mg mL⁻¹. At the lowest oscillation rate, the mucilage had a dynamic viscosity of 145 mPa s and behaved as a weak viscoelastic gel. After filtration to remove suspended root cap cells and other solid plant material, mucilage viscosity was reduced to 5–10 mPa s at low oscillation rates and the behaviour was that of a viscous liquid. The decrease in viscosity which occurs on filtration indicates that the root cap cells form an integral part of the gel system, either by interacting directly with each other or via the polysaccharide. Our observations provide further support for the idea that mucilage plays a major role in maintaining root-soil contact in the rhizosphere. This revised version was published online in June 2006 with corrections to the Cover Date.     

粘液繁殖体种子的粘液质形成、分泌及释放相关基因

种皮粘液质是在种皮外层细胞的高尔基体内产生并分泌到胞腔内或细胞壁层的一种果胶类多糖物质.当干燥种子遇水后,粘液质即刻被释放形成透明胶质并完全包被整个种子.粘液质对种子的扩散定居、种子萌发以及幼苗的存活和生长均具有重要作用.粘液质作为一种模型研究细胞壁的产生及其形成的分子机制已经成为植物种皮发育与环境变化相适应关系的研究...     

Comparison of maize root mucilages isolated from root exudates and root surface extracts by complementary cytological and biochemical investigations

Summary A strategy to obtain fractions enriched in mucilages secreted by root caps or produced by the rhizodermis of axenicallygrown maize seedlings is proposed. It involves a two-step procedure allowing the successive collection of root exudates and surface extracts from the same set of intact, sterile maize plants. Cytological controls were performed at each phase of collection. Whereas root cap mucilage is easily collected in water after one day's extraction, under conditions favouring secretory activity, rhizodermal mucilage remains tightly adherent to the root surface. It can be better extracted using neutral saline buffer assisted by gentle shaking at low temperature. Acidic saline buffer is unsuitable as it induces cell lysis and release of cell wall components.Biochemical analyses confirm that fractions enriched in root cap mucilage contain very high levels of fucose and galactose, high levels of arabinose, xylose and glucose and trace amounts of mannose. Fractions enriched in rhizodermal mucilage contain large amounts of glucose, moderate amounts of arabinose, xylose, mannose and galactose and trace levels of fucose. Isoelectric focusing and SDS-PAGE indicate that there are numerous similarities in the protein composition of materials enriched in root cap mucilages from root exudates or aqueous root surface extracts. However, specific protein bands that could be characteristic of rhizodermal mucilage are obtained using neutral saline buffer extracts. According to these biochemical data, the two-step procedure used in the present study appears to be useful for further biochemical characterization of both types of mucilages.Abbreviations BSA bovine serum albumin - BSTFA N,O-bis (trimethylsilyl)-trifluoroacetamide - DTT dithiothreitol - i. d. internal diameter - MW molecular weight - PATAg periodic acid-thiosemicarbazide-silver proteinate - PVPP polyvinylpolypyrrolidone - RE root exudates - RSE root surface extracts - TMCS trimethylchlorosilane - TMS trimethylsilyl     

A subtilisin-like serine protease essential for mucilage release from Arabidopsis seed coats

During Arabidopsis seed development large quantities of mucilage, composed of pectins, are deposited into the apoplast underneath the outer wall of the seed coat. Upon imbibition of mature seeds, the stored mucilage expands through hydration and breaks the outer cell wall that encapsulates the whole seed. Mutant seeds carrying loss-of-function alleles of AtSBT1.7 that encodes one of 56 Arabidopsis thaliana subtilisin-like serine proteases (subtilases) do not release mucilage upon hydration. Microscopic analysis of the mutant seed coat revealed no visible structural differences compared with wild-type seeds. Weakening of the outer primary wall using cation chelators triggered mucilage release from the seed coats of mutants. However, in contrast to mature wild-type seeds, the mutant's outer cell walls did not rupture at the radial walls of the seed coat epidermal cells, but instead opened at the chalazal end of the seed, and were released in one piece. In atsbt1.7, the total rhamnose and galacturonic acid contents, representing the backbone of mucilage, remained unchanged compared with wild-type seeds. Thus, extrusion and solubility, but not the initial deposition of mucilage, are affected in atsbt1.7 mutants. AtSBT1.7 is localized in the developing seed coat, indicating a role in testa development or maturation. The altered mode of rupture of the outer seed coat wall and mucilage release indicate that AtSBT1.7 triggers the accumulation, and/or activation, of cell wall modifying enzymes necessary either for the loosening of the outer primary cell wall, or to facilitate swelling of the mucilage, as indicated by elevated pectin methylesterase activity in developing atsbt1.7 mutant seeds.     

Cellulose synthesis via the FEI2 RLK/SOS5 pathway and cellulose synthase 5 is required for the structure of seed coat mucilage in Arabidopsis

The seeds of Arabidopsis thaliana and many other plants are surrounded by a pectinaceous mucilage that aids in seed hydration and germination. Mucilage is synthesized during seed development within maternally derived seed coat mucilage secretory cells (MSCs), and is released to surround the seed upon imbibition. The FEI1/FEI2 receptor-like kinases and the SOS5 extracellular GPI-anchored protein were shown previously to act on a pathway that regulates the synthesis of cellulose in Arabidopsis roots. Here, we demonstrate that both FEI2 and SOS5 also play a role in the synthesis of seed mucilage. Disruption of FEI2 or SOS5 leads to a reduction in the rays of cellulose observed across the seed mucilage inner layer, which alters the structure of the mucilage in response to hydration. Mutations in CESA5, which disrupts an isoform of cellulose synthase involved in primary cell wall synthesis, result in a similar seed mucilage phenotype. The data indicate that CESA5-derived cellulose plays an important role in the synthesis and structure of seed coat mucilage and that the FEI2/SOS5 pathway plays a role in the regulation of cellulose synthesis in MSCs. Moreover, these results establish a novel structural role for cellulose in anchoring the pectic component of seed coat mucilage to the seed surface.     

Role of mucilage during achene germination and sprout growth of the endangered Tibetan medicinal herb Mirabilis himalaica (Nyctaginaceae) exposed to abiotic stresses

Aim Mirabilis himalaica (Nyctaginaceae) is an endangered medicinal plant mainly distributed in the plateau region of northern Tibet, China. The outer surface of M. himalaica achenes is covered by a pectinaceous mucilaginous layer upon hydration. However, the role of the achene mucilage is poorly understood. In this study, we investigated the effects of mucilage on achene germination and sprout growth under abiotic stress to explain how M. himalaica survive the alpine environment.Methods We investigated the effect of mucilage on achenes germination by contrast the capacity of water absorption, dehydration and respiration of intact achene and the achene with mucilage removal. We performed abiotic stresses experiments including drought stress, salt stress, cold stress and high temperature stress, and quantified the effects of mucilage removal on achene germination rate, root and shoot lengths of seedlings.Important findings Mucilage is extremely hydrophilic, and the mass of intact achenes can be 9-fold greater than that of demucilaged achenes. The removal of the mucilaginous layer did not significantly change final germination percentages under ideal conditions, but intact achenes (i.e. with mucilage) took longer to germinate. The mucilage significantly decreased seed respiration rates by acting as a physical barrier that prevented oxygen diffusion. Germination rates, shoot and root growth of intact achenes were higher than those of demucilaged ones during exposures to cold, heat, osmotic and salt stresses. Achene mucilage presumably plays an ecologically important role in the life cycle of M. himalaica by aiding the critical achene germination and early seedling growth in the stressful habitats of the plateau region of northern Tibet.     

Stimulation of polysaccharide formation by 2,4-dichlorophenoxyacetic acid in callus tissues of Arabidopsis thaliana

A relatively high concentration of 2,4-dichlorophenoxyacetic acid (45 μ M ) in solid culture medium stimulated the formation and secretion of mucilage polysaccharides by callus tissues of Arabidopsis thaliana L. Heynh. (line Estland). The mucilage was composed of at least two polysaccharides as revealed by gel chromatography on Sepharose 4B: the major component (87%) eluted in the void volume (molecular weight 2 × 10⁶ or greater) and the minor component (13%) eluted in the molecular weight range from 2 × 10⁴ to 4 × 10⁵. Both polysaccharide components contained small amounts of uronic acids. The major polysaccharide consisted mostly of galactose (49%), arabinose (28%) and fucose (10%), whereas the minor one consisted of galactose (44%), xylose (18%), arabinose (14%) and rhamnose (14%). One of the components of the secreted mucilage seems to be an arabinogalactan.     

Effects of mucilage on seed germination of the desert ephemeral plant Plantago minuta Pall. under osmotic stress and cycles of wet and dry conditions

To test the role of the seed mucilage of Plantago minuta Pall. in regulating germination under osmotic stress and cycles of hydration and dehydration, two experiments were carried out using seeds with intact mucilage and mucilage‐free seeds. In Experiment 1 seeds were immersed in a range of iso‐osmotic polyethylene glycol solutions (?1.15 to 0 MPa) for 14 days; any ungerminated seeds were transferred to deionized water to investigate the recovery germination. In Experiment 2 seeds were immersed in deionized water for 24 h, and were then incubated on filter paper for an additional 13 days to ensure complete desiccation before reimbibition to test the germination recovery percentage. Under mild osmotic stress (?0.73 to 0 MPa), the intact seeds with mucilage were shown to have higher germination rates than the mucilage‐free seeds, indicating that the mucilage led to a “fast sprouting” germination strategy under mild osmotic stress. However, when seeds were exposed to high osmotic stress (?1.15 MPa), the mucilage apparently slowed the germination rate, resulting in a “risk‐balancing” germination strategy. Extreme drought induced by polyethylene glycol solution and the desiccation pretreatment accelerated germination rates compared to non‐pretreated seeds; both germination potential and recovery percentage of the mucilage seeds were significantly higher than that of the mucilage‐free seeds. Our results revealed that the seed mucilage of P. minuta plays a crucial role in regulating seed germination rates and the germination strategies adopted by controlling seed water absorption when the seeds experience different osmotic stresses or alternating wet and dry conditions.     

Detection of root mucilage using an anti-fucose antibody

Plant root mucilage is known to enhance soil quality by contributing towards the soil carbon pool, soil aggregation, detoxification of heavy metal ions and interactions with rhizospheric microflora. Mucilage consists of many monosaccharide units, including fucose which can be used as an indicator for plant root based polysaccharides. This is the first report of an immunological technique developed to use anti-fucose antibodies as markers for probing and localizing fucosyl residues in mucilage polysaccharide and, in turn, for localization of plant root mucilage. Fucose was complexed with bovine serum albumin to raise antibodies against fucose. A fucose-directed antibody was shown to cross-react with root cap mucilages from grasses. This antibody was used to localize root mucilage polysaccharide in maize and wheat root caps using immunogold electron microscopy. Abundant labelling could be localized on the cell wall, and in the intercellular matrix and vesicles of the peripheral root cap cells. Labelling was less intense in cells towards the centre of the root cap tissue. Control experiments confirmed that immunogold localization of fucose was specific and reliable.     

DEVELOPMENT OF MUCILAGINOUS SURFACES IN EUGLENOIDS. II. FLAGELLATED,CREEPING AND PALMELLOID CELLS OF EUGLENA1

Critical-point dried (CPD) cells from clonal cultures of Euglena gracilis Klebs (Z strain), E. deses Ehrb., E. tripteris (Duj.) Klebs and E. myxocylindracea Bold & MacEntee were examined by scanning electron microscopy. Flagellated motile cells of E. gracilis are naked except for a few strands of mucilage on the posterior tip. Flagellated cells of E. tripteris have a permanent mucilage coating often of uneven distribution and usually not as well developed as that of nonflagellated creeping cells which have a distinctive mucilage. In E. deses the coating appears rough due to the aggregation of isolated groups of strands above the cell surface. In E. tripteris the coating appears smooth except for breaks near the articulation of the pellicular strips where the mucilage may rise above the surface to form waves. At high magnification this mucilage consists of a network of strands generally lying parallel to the cell surface; the strands become obscure in some specimens. In E. myxocylindracea elongated, mucilage-coated cells contract to form spheres which undergo further mucilage deposition producing the mucilage covering of palmellae. As palmellae mature, the mucilage surface becomes less porous and the individuality of most mucilage strands is lost.     

Cholesterol is not crucial for the existence of microdomains in kidney brush-border membrane models.

The external membrane leaflet plays a key role in the organization of the cell plasma membrane as a mosaic of ordered microdomains enriched in sphingolipids and cholesterol and of fluid domains. In this study, the thermotropic behavior and the topology of bilayers made of a phosphatidylcholine/sphingomyelin mixture, which mimicks the lipid composition of the external leaflet of renal brush-border membranes, were examined by differential scanning calorimetry and atomic force microscopy. In the absence of cholesterol, a broad phase separation process occurred where ordered gel phase domains of size varying from the mesoscopic to the microscopic scale, enriched in sphingomyelin, occupied half of the bilayer surface at room temperature. Increasing amounts of cholesterol progressively decreased the enthalpy of the transition and modified the topology of membranes domains up to a concentration of 33 mol % for which no membrane domains were detected. These results strongly suggest that, in membranes highly enriched in sphingolipids like renal and intestinal brush borders, there is a threshold close to the physiological concentration above which cholesterol acts as a suppressor rather than as a promoter of membrane domains. They also suggest that cholesterol depletion does not abolish the lateral heterogenity in brush-border membranes.     

Simulation of gel phase formation and melting in lipid bilayers using a coarse grained model

The transformation between a gel and a fluid phase in dipalmitoyl-phosphatidylcholine (DPPC) bilayers has been simulated using a coarse grained (CG) model by cooling bilayer patches composed of up to 8000 lipids. The critical step in the transformation process is the nucleation of a gel cluster consisting of 20-80 lipids, spanning both monolayers. After the formation of the critical cluster, a fast growth regime is entered. Growth slows when multiple gel domains start interacting, forming a percolating network. Long-lived fluid domains remain trapped and can be metastable on a microsecond time scale. From the temperature dependence of the rate of cluster growth, the line tension of the fluid-gel interface was estimated to be 3+/-2 pN. The reverse process is observed when heating the gel phase. No evidence is found for a hexatic phase as an intermediate stage of melting. The hysteresis observed in the freezing and melting transformation is found to depend both on the system size and on the time scale of the simulation. Extrapolating to macroscopic length and time scales, the transition temperature for heating and cooling converges to 295+/-5 K, in semi-quantitative agreement with the experimental value for DPPC (315 K). The phase transformation is associated with a drop in lateral mobility of the lipids by two orders of magnitude, and an increase in the rotational correlation time of the same order of magnitude. The lipid headgroups, however, remain fluid. These observations are in agreement with experimental findings, and show that the nature of the ordered phase obtained with the CG model is indeed a gel rather than a crystalline phase. Simulations performed at different levels of hydration furthermore show that the gel phase is stabilized at low hydration. A simulation of a small DPPC vesicle reveals that curvature has the opposite effect.     

Influence of maize root mucilage on soil aggregate stability

This study was undertaken to determine the effects of root exudates on soil aggregate stability. Root mucilage was collected from two-month old maize plants (Zea mays L.) Mucilage and glucose solutions were added at a rate of 2.45 g C kg⁻¹ dry soil to silty clay and silt loam soils. Amended soils, placed in serum flasks, were incubated for 42 d with a drying-wetting cycle after 21 d. Evolved CO₂ was measured periodically as well as the water-stable aggregates and soluble sugar and polysaccharide content of the soil. In mucilage-amended soils CO₂ evolution started with a lag phase of 2–3 days, which was not observed in glucose-amended soils. There was then a sharp increase in evolved CO₂ up to day 7. During the second incubation period there were only small differences in evolved C between treatments. Incorporation of mucilage in both soils resulted in a spectacular and immediate increase in soil aggregate stability. Thereafter, the percent of water-stable aggregates quickly decreased parallel to microbial degradation. On completion of the incubation, aggregate stability in the silty clay soil was still significantly higher in the presence of mucilage than in the control. This work supports the assumption that freshly released mucilage is able to stick very rapidly to soil particles and may protect the newly formed aggregates against water destruction. On the silty clay, microbial activity contributes to a stabilization of these established organo-mineral bounds.     

The effect of dolichol on the structure and phase behaviour of phospholipid model membranes

The effect of dolichol C(95) on the structure and thermotropic phase behaviour of dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylethanolamine and stearoyloleoylphosphatidylethanolamine has been examined by synchrotron X-ray diffraction and differential scanning calorimetry. The presence of dolichol C(95) had no detectable effects on the temperature of either the gel to ripple or the ripple to liquid-crystal phase transition of dipalmitoylphosphatidylcholine. A proportionate increase of a few degrees in the temperature of the gel to lamellar liquid-crystal phase transition is observed in dispersions of dipalmitoylphosphatidylethanolamine and significantly there is a decrease in the temperature of the lamellar to non-lamellar phase transition of stearoyloleoylphosphatidylethanolamine. There was no significant change in the bilayer repeat spacing of all three mixed dispersions in gel phase in the presence of up to 20 mol% dolichol C(95). Electron density calculations showed that there was no change of bilayer thickness of dipalmitoylphosphatidylcholine with incorporation of up to 7.5 mol% dolichol C(95). These data suggest that effect of dolichol on the phospholipid model membranes depend on both the head group and the hydrocarbon chains of the phospholipid molecules. The presence of dolichol in phosphatidylcholine bilayers conforms to a model in which the polyisoprene compound is phase separated into a central domain sandwiched between the two monolayers in gel phase. In bilayers of phosphatidylethanolamines dolichol tends to stabilize the bilayers in gel phase at low temperatures and destabilize the bilayers in lamellar disordered structure at high temperatures. Non-lamellar structures coexist with lamellar disordered phase over a wide temperature range suggesting that dolichol is enriched in domains of non-lamellar structure and depleted from lamellar phase. These findings are useful to understand the function of dolichol in cell membranes.     

Root Cap Mucilage and Extracellular Calcium as Modulators of Cellular Growth in Postmitotic Growth Zones of the Maize Root Apex*

Abstract: The control of maize root growth by root cap mucilage and extracellular calcium (Ca) was examined. Special attention was paid to the influence of these factors on cellular aspects of root growth, such as cell shape and organization of the microtubular (MT) cytoskeleton. Externally supplied Ca impaired the transition of early post-mitotic cells from a more-or-less apolar mode of expansion to a strictly anisotropic mode of elongation accompanied by their more rapid growth. However, this inhibitory effect of Ca was not associated with any re-arrangement of the cortical MTs, their transverse arrays, with respect to the root axis, being maintained under these conditions. Root mucilage, collected from donor root caps and placed around root tips, exerted a similar effect on cell shapes as did externally supplied Ca. In contrast, roots grown in a medium of low Ca content, or from which the root cap mucilage was continually removed, had more elongated cell shapes in their post-mitotic growth regions when compared to the control roots. These findings are consistent with a notion that Ca is present in the root cap mucilage in physiologically relevant amounts and can mediate growth responses in both the PIG region and the apical part of the elongation zone. Integrating several known effects of Ca ions on growth at the root apex, a hypothesis is proposed that a Ca-mediated and MT-independent control of cell growth in the PIG region might be involved in morphogenetic root movements (e.g. gravitropism), and that root growth responses could be initiated by an asymmetric distribution of extracellular calcium, or root cap slime, around the growing root tip.     

Characterization of root mucilage from Melastoma malabathricum, with emphasis on its roles in aluminum accumulation

Plant roots exude viscous polysaccharides, called mucilage. One of the suggested roles of mucilage is immobilization of toxic metal cations, including aluminum (Al), in the rhizosphere. Mucilage exuded from roots of Melastoma malabathricum (Al accumulator) was characterized in comparison with that of Zea mays (maize; Al nonaccumulator). Removal of mucilage significantly reduced Al accumulation in M. malabathricum. The cation adsorption affinity of M. malabathricum mucilage was higher for Al and lanthanum (La) than for barium (Ba), whereas that of maize mucilage was in the order Ba > La > Al. A (27)Al nuclear magnetic resonance (NMR) spectrum of the Al-adsorbed mucilage and bioassay with alfalfa seedlings indicated that the concentrated Al in the mucilage of M. malabathricum, unlike that of maize, bound very weakly to cation exchange sites of mucilage. The higher charge density in M. malabathricum mucilage, derived from unmethylated uronic acid, is inferred to be related to preferential adsorption of trivalent cation. Not only a higher degree of methylation in the uronic acid (glucuronic acid) but also H(+) release from roots to the mucilage appears to be responsible for the loose binding of Al in M. malabathricum mucilage. These characteristics of mucilage may help Al hyperaccumulation in M. malabathricum.