Xyloglucan Endotransglycosylase Activity, Microfibril Orientation and the Profiles of Cell Wall Properties Along Growing Regions of Maize Roots

A series of physical and chemical analyses were made on theexpanding zone of maize seedling roots grown in hydroponics.Comparison of longitudinal profiles of local relative elementalgrowth rate and turgor pressure indicated that cell walls becomelooser in the apical 5 mm and then tighten 5–10 mm fromthe root tip. Immersion of roots in 200 mol m^–3 mannitol(an osmotic stress of 0·48 MPa) rapidly and evenly reducedturgor pressure along the whole growing region. Growth was reducedto a greater extent in the region 5–10 mm from the roottip than in the apical region. This indicated rapid wall-looseningin the root tip, but not in the more basal regions. Following 24 h immersion in 400 mol m^–3 mannitol (an osmoticstress of 0·96 MPa) turgor had recovered to pre-stressedvalues. Under this stress treatment, growth was reduced in theregion 4–10 mm from the root tip, despite the recoveryof turgor, indicating a tightening of the wall. In the rootapex, local relative elemental growth rate was unchanged incomparison to control tissue, showing that wall properties herewere similar to the control values. Cellulose microfibrils on the inner face of cortical cell wallsbecame increasingly more parallel to the root axis along thegrowth profile of both unstressed and stressed roots. Orientationdid not correlate with the wall loosening in the apical regionof unstressed roots, or with the tightening in the region 5–10mm from the root tip following 24 h of osmotic stress. Longitudinal profiles of the possible wall-loosening enzymexyloglucan endotransglycosylase (XET) had good correspondencewith an increase in wall loosening during development. In thezone of wall tightening following osmotic stress, XET activitywas decreased per unit dry weight (compared with the unstressedcontrol), but not per unit fresh weight. Key words: Osmotic stress, turgor, growth, cell wall properties, microfibrils, XET     

Oligosaccharide side chains of wall molecules are essential for cell-wall lysis in Chlamydomonas reinhardtii

The glycoproteins of the cell walls of Chlamydomonas are lysed during the reproductive cycle by proteases (autolysins) which are specific for their substrates. The autolysin which digests the wall of sporangia to liberate the zoospore daughter cells in the vegetative life cycle is a collagenase-like enzyme which attacks only selected domains in its wall substrates containing (hydroxy)-proline clusters. Cell-wall fractions obtained by salt-extraction (NaClO₄) and oxidizing agents (NaClO₂) and the insoluble residue were tested as substrates. The most-crosslinked insoluble inner part of the wall is the best substrate for the sporangia autolysin. Oligosaccharides obtained from the insoluble cell-wall fraction of sporangia by hydrolysis with Ba(OH)₂ inhibit autolysin action. We conclude that the oligosaccharide side chains of wall substrates are essential for forming the reactive enzyme-substrate complex.Abbreviations CSW chlorite-soluble cell-wall fraction - ICW insoluble cell-wall fraction - PSW salt-soluble fraction - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

Extraction by lithium chloride of hydroxyproline-rich glycoproteins from intact cells of Chlamydomonas reinhardii

A procedure has been developed to isolate and analyse the cell-wall glycoproteins of Chlamydomonas reinhardii. Under appropriate conditions, cell-wall glycoproteins can be quantitatively extracted from intact cells by aqueous LiCl. Although proteins and glycoproteins, which are presumably not related to the cell wall, are coextracted with the cell-wall subunits, these components can be readily identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis as demonstrated by comparative analysis of LiCl-extracts from wild-type cells and the cell-wall-deficient mutant CW-15. Apart from the high-molecular-weight cell-wall components, two glycoproteins with apparent molecular weights (M_rs) of 36000 and 66000 were found to be present in LiCl-extracts of wild-type cells but absent in LiCl-extracts from the cell-wall-less mutant. Pulse-labeling experiments with [³H]proline and [³⁵S]methionine revealed that the LiCl-extracts contained — in addition to the well-known cell-wall subunits — proteins of lower molecular weight, which are also preferentially labeled with [³H]proline. Protein components with M_rs of 68000, 44000, 36000, 26000 and 22000 were found to be more strongly labeled with [³H]proline than with [³⁵S]methionine, whereas protein components with M_rs of 57000 and 52000 were more prominent after labeling with [³⁵S]methionine. The portion of cell-wall subunits within the total amount of proteins extracted by LiCl was calculated to be at least 10% on the basis of the amount of hydroxyproline. Self-assembly of cell walls could be demonstrated after dialysis against water of a mixture of crude LiCl-extract and purified, insoluble, inner wall layers. Cell-wall glycoproteins could be enriched by gel exclusion chromatography of crude LiCl-extracts on Sepharose CL-4B columns equilibrated with 1 mol l^-1 LiCl.Abbreviations EDTA ethylenediaminetetraacetic-acid - PAGE polyacrylamide gel electrophoresis - PAS periodic acid Schiff's reagent - SDS sodium dodecyl sulfate - TCA trichloroacetic acid - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol     

Cell wall peroxidase activity,hydrogen peroxide level and NaCl-inhibited root growth of rice seedlings

The changes in cell-wall peroxidase (POD) activity and H₂O₂ level in roots of NaCl-stressed rice seedlings and their correlation with root growth were investigated. Increasing concentrations of NaCl from 50 to 150 mM progressively reduced root growth and increased ionically bound cell-wall POD activity. NaCl had no effect on covalently bound cell-wall POD activities. The reduction of root growth by NaCl is closely correlated with the increase in H₂O₂ level. Exogenous H₂O₂ was found to inhibit root growth of rice seedlings. Since ammonium and proline accumulation are associated with root growth inhibition caused by NaCl, we determined the effects of NH₄Cl or proline on root growth, cell-wall POD activity and H₂O₂level in roots. External application of NH₄Cl or proline markedly inhibited root growth, increased cell-wall POD activity and increased H₂O₂ level in roots of rice seedlings in the absence of NaCl. An increase in cell-wall POD activity and H₂O₂ level preceded inhibition of root growth caused by NaCl, NH₄Cl or proline. NaCl or proline treatment also increased NADH-POD and diamine oxidase (DAO) activities in roots of rice seedlings, suggesting that NADH-POD and DAO contribute to the H₂O₂ generation in the cell wall of NaCl- or proline-treated roots. NH₄Cl treatment increased NADH-POD activity but had no effect on DAO activity, suggesting that NADH-POD but not DAO is responsible for H₂O₂ generation in cell wall of NH₄Cl-treated roots.     

Comparison of Various Procedures for Removing Proteins and Nucleic Acids from Cell Walls of Bacillus Subtilis

Several procedures were used i n an attempt to prepare clean cell walls from Bacillus subtilis. The results indicate that protein and nucleic acids are tightly bound tothe walls. cleanest wall preparations were found following trichloroacetic acid extraction at 60° or by extraction with 0.lN NaOH under a nitrogen atmosphere for 10 hrs. Protein denaturants, such as sodium dodecyl sulfate and concentrated guanidine hydrochloride were relatively ineffective in removing proteins and nucleic acids from the cell walls. Cell wall-bound DNA was biologically The active i n transformation assays.     

In-vivo measurement of cell-wall extensibility in maize coleoptiles: Effects of auxin and abscisic acid

Plastic and elastic in-vivo extensibilities (E_pl and E_el, respectively) of cell walls of growing maize (Zea mays L.) coleoptile segments were measured by stretching living tissue at constant force (creep test) in an extensiometer. The linear displacement transducer used as a measuring device permits the determination of load-induced extensions in the range of 0–1% of the segment's length, leading to a minimal disturbance of the hydraulic parameters of the tissue and allowing the measurement of unidirectional cell-wall creep at virtually unchanged turgor and metabolic activity. A rein-vestigation of the time-course of indole-3-acetic acid-promoted and abscisic acid-inhibited wall loo-sening revealed that the in-vivo creep test yields results very similar to those obtained previously with the in-vitro creep test [Kutschera and Schopfer, 1986, Planta 167, 527–535]. The hormones affect elongation rate and E_pl in a closely correlated manner both in step-up as well as step-down growth changes whereas E_el remains unaltered. It is argued that both hormones influence growth by modifying E_pl of the outer epidermis and that this effect can be quantitatively measured, in relative units, by either the in-vivo or the in-vitro creep test.Abbreviations ABA ±abscisic acid - E_el, E_pl elastic and plastic in-vivo cell-wall extensibility, respectively - E_tot E_el+E_pl - IAA indole-3-acetic acid; m, cell-wall yielding coefficient     

pH and expansin action on single suspension-cultured tomato (Lycopersicon esculentum) cells

The aim of this study was to measure key material properties of the cell walls of single suspension-cultured plant cells and relate these to cell-wall biochemistry. To this end, micromanipulation was used to compress single tomato cells between two flat surfaces until they ruptured, and force-deformation data were obtained. In addition to measuring the bursting force, we also determined the elastic (Young’s) modulus of the cell walls by matching low strain (≤20% deformation) experimental data with a cell compression model, assuming linear elastic cell walls. The walls were most elastic at pH 4.5, the pH optimum for expansin activity, with an elastic modulus of 2.0 ± 0.1 GPa. Following the addition of exogenous expansins, cell walls became more elastic at all pH values. Western blot analysis of proteins from walls of cultured cells revealed the presence of expansin epitopes, suggesting that the inherent pH dependence of elasticity and other compression phenomena is related to the presence of endogenous expansin proteins and their wall-loosening ability. Although strict application of the linear-elastic model could not be applied to large deformations—for example, up to cell bursting—because of irreversible behaviour, the deviation of the data from the model was generally small enough to allow estimation of the strain in the cell wall at failure. This strain was greater at pH 4.5 and when expansins were added to the suspension. The changes in elasticity are consistent with suggestions about the mode of expansin action. The estimated strains at failure are compatible with data on the failure of Acetobacter-derived cellulose–xyloglucan composites and proposed mechanisms of such failure. Through the measurement of cell-wall material properties using micromanipulation, it may be possible to understand more fully how cell-wall composition, structure and biochemistry lead to cell mechanical behaviour.     

Root hair-specific EXPANSIN B genes have been selected for graminaceae root hairs

Cell differentiation ultimately relies on the regulation of cell type-specific genes. For a root hair cell to undergo morphogenesis, diverse cellular processes including cell-wall loosening must occur in a root hair cell-specific manner. Previously, we identified and characterized root hairspecific cis-elements (RHE) from the genes encoding the cell wall-loosening protein EXPANSIN A (EXPA) which functions preferentially on dicot cell walls. This study reports two root hair-specific grass EXPB genes that contain RHEs. These genes are thought to encode proteins that function more efficiently on grass cell walls. The proximal promoter regions of two orthologous EXPB genes from rice (Oryza sativa; OsEXPB5) and barley (Hordeum vulgare; HvEXPB1) included RHE motifs. These promoters could direct root hair-specific expression of green fluorescent protein (GFP) in the roots of rice and Arabidopsis (Arabidopsis thaliana). Promoter deletion analyses demonstrated that the RHE motifs are necessary for root hairspecific expression of these EXPB promoters. Phylogenetic analysis of EXP protein sequences indicated that grass EXPBs are the only orthologs to these root hair-specific EXPBs, separating dicot EXPBs to distal branches of the tree. These results suggest that RHE-containing root hair-specific EXPB genes have evolved for grass-specific cell wall modification during root hair morphogenesis.     

Loss of capacity for acid-induced wall loosening as the principal cause of the cessation of cell enlargement in light-grown bean leaves

Cell enlargement in primary leaves of bean (Phaseolus vulgaris L.) can be induced, free of cell divisions, by exposure of 10-d-old, red-light-grown seedlings to white light. The absolute rate of leaf expansion increases until day 12, then decreases until the leaves reached mature size on day 18. The cause of the reduction in growth rate following day 12 has been investigated. Turgor calculated from measurements of leaf water and osmotic potential fell from 6.5 to 3.5 bar before day 12, but remained constant thereafter. The decline of growth after day 12 is not caused by a decrease in turgor. On the other hand, Instron-measured cell-wall extensibility decreased in parallel with growth rate after day 12. Two parameters influencing extensibility were examined. Light-induced acidification of cell walls, which has been shown to initiate wall extension, remained constant over the growth period (days 10–18). Furthermore, cells of any age could be stimulated to excrete H⁺ by fusicoccin. However, older tissue was not able to grow in response to fusicoccin or light. Measurements of acid-induced extension on preparations of isolated cell walls showed that as cells matured, the cell walls became less able to extend when acidified. These data indicate that it is a decline in the capacity for acid-induced wall loosening that reduces wall extensibility and thus cell enlargement in maturing leaves.Abbreviations and symbols FC fusicoccin - P turgor pressure - RL red light - WEx wall extensibility - WL white light - P _w leaf water potential - P _s osmotic potential     

Radial and axial turgor pressure measurements in individual root cells of Mesembryanthemum crystallinum grown under various saline conditions

Abstract. Radial and axial turgor pressure profiles were measured with the pressure probe in untreated and salt-treated intact roots of Mesembryanthemum crystallinum. The microcapillary of the pressure probe was inserted step-wise into the root tissue 5, 25 and 50 mm away from the root cap. For evaluation of the data, only those recordings on a given root were used in which four discontinuous increases in turgor pressure occurred. These four turgor pressure increases could be related to the rhizodermal cells and to the cells in the three cortical layers. The measurements showed that a radial turgor pressure gradient of the same magnitude (directed from the third cortical layer to the external medium) existed along the root axis. The magnitude of this turgor pressure gradient decreased with increasing salinity (up to 400 mol m^-3 NaCl) in the growth medium. Addition of 10 mol m^-3 CaCl₂ to the 400 mol m^-3 NaCl medium partly reduced the salt-induced decrease in turgor pressure, but only in cells 25–50 mm away from the root tip. Combined with this effect, a small axial turgor pressure gradient was generated, therefore, in the cortex layers which was directed to the root tip. Measurements of the volumetric elastic modulus, ?, of the wall of the individual cells showed that the presence of salt considerably reduced the magnitude of this parameter and that addition of Ca²⁺ to the strongly saline medium partially diminished this decrease. This effect was strongest in cells 50 mm away from the root tip. The magnitude of ? of rhizodermal and cortical cells increased along the root axis both in untreated and in salt-treated roots. The ? value was significantly smaller for rhizodermal cells compared to the cortical cells, with the exception of cells 50 mm from the tip. In this tissue, rhizodermal and cortical cells exhibited nearly the same values. The decrease of the ?-values with salt and the increase along the root axis under the various growth conditions could be correlated with corresponding changes in cell volume. Diurnal changes in turgor pressure could not be detected in the individual root cells, with the notable exception of the rhizodermal and cortical cells located in the region 50 mm away from the root tip of the control plants. In these cells, an increase in turgor pressure was observed during the morning hours. Determination of the average osmotic pressure in tissue sections along the roots of control and salt-treated plants revealed that at 400 mol m^-3 NaCl the osmotic pressure gradient between the tissue and the medium is exo-directed, provided that the water is not (partly) immobilized.     

Exogenous nitric oxide enhances cadmium tolerance of rice by increasing pectin and hemicellulose contents in root cell wall

To study the mechanisms of exogenous NO contribution to alleviate the cadmium (Cd) toxicity in rice (Oryza sativa), rice plantlets subjected to 0.2-mM CdCl₂ exposure were treated with different concentrations of sodium nitroprusside (SNP, a NO donor), and Cd toxicity was evaluated by the decreases in plant length, biomass production and chlorophyll content. The results indicated that 0.1 mM SNP alleviated Cd toxicity most obviously. Atomic absorption spectrometry and fluorescence localization showed that treatment with 0.1 mM SNP decreased Cd accumulation in both cell walls and soluble fraction of leaves, although treatment with 0.1 mM SNP increased Cd accumulation in the cell wall of rice roots obviously. Treatment with 0.1 mM SNP in nutrient solution had little effect on the transpiration rate of rice leaves, but this treatment increased pectin and hemicellulose content and decreased cellulose content significantly in the cell walls of rice roots. Based on these results, we conclude that decreased distribution of Cd in the soluble fraction of leaves and roots and increased distribution of Cd in the cell walls of roots are responsible for the NO-induced increase of Cd tolerance in rice. It seems that exogenous NO enhances Cd tolerance of rice by increasing pectin and hemicellulose content in the cell wall of roots, increasing Cd accumulation in root cell wall and decreasing Cd accumulation in soluble fraction of leaves.     

Hardening of root cell walls: a growth inhibitory response to salinity stress

Primary roots of intact maize plants (Zea mays L.) grown for several days in nutrient solutions containing 100 mol m⁻³ NaCl and additional calcium, had relatively inhibited rates of elongation. Possible physical restraints underlying this salt induced inhibition were investigated. The inhibition did not involve reductions in osmotic potential gradients and turgor in the tip tissues responsible for root elongation growth. The apparent yield threshold pressure, which is related to capacity of cell walls to undergo loosening by stress relaxation, was estimated psychrometrically in excised root tips. Salinity increased yield threshold values. Comparative root extensibility values were obtained for intact plants by determining the initial (1 min) increase in root elongation rate induced by an 0.1 MPa osmotic jump. Comparative extensibility was significantly reduced in the salinized root tips. Salinity did not reduce capacities for water efflux and associated elastic contraction in root tip tissues of intact plants exposed to hypertonic mannitol. We conclude that cell wall hardening in the elongating root tips is an important component of root growth inhibition induced by long-term salinization.     

Rheological analysis of viscoelastic cell wall changes in maize coleoptiles as affected by auxin and osmotic stress

The effects of auxin and osmotic stress on elongation growth of maize (Zea mays L.) coleoptile segments are accompanied by characteristic changes in the extensibility of the growth-limiting cell walls. At full turgor auxin causes growth by an increase in wall extensibility (wall looseining). Growth can be stopped by an osmotically produced step-down in turgor of 0.45 MPa. Under these conditions auxin causes the accumulation of a potential for future wall extension which is released after restoration of full turgor. Turgor reduction causes a reversible decrease in wall extensibility (wall stiffening) both in the presence and absence of auxin. These changes in vivo are correlated with corresponding changes in the rheological properties of the cell walls in vitro which can be traced back to specific modifications in the shape of the hysteretic stress-strain relationship. The longitudinally load-bearing walls of the coleoptile demonstrate almost perfect viscoelasticity as documented by a nearly closed hysteresis loop. Auxin-mediated wall loosening causes an increase of loop width and thus affects primarily the amount of hysteresis in the isolated wall. In contrast, turgor reduction by osmotic stress reduces loop length and thus affects primarily the amount of viscoelastic wall extensibility. Pretreatment of segments with anoxia and H₂O₂ modify the hysteresis loop in agreement with the conclusion that the wall-stiffening reaction visualized under osmotic stress in vivo is an O₂-dependent process in which O₂ can be substituted by H₂O₂. Cycloheximide specifically inhibits auxin-mediated wall loosening without affecting wall stiffening, and this is mirrored in specific changes of the hysteresis loop. Corroborating a previous in vivo study (Hohl et al. 1995, Physiol. Plant. 94: 491–498) these results show that cell wall stiffening in vivo can also be demonstrated by Theological measurements with the isolated cell wall and that this process can be separated from cell wall loosening by specific changes in the shape of the hysteresis loop.     

Acid-induced wall loosening is confined to the accelerating region of the root growing zone

The aims of this study were to quantify developmental differences in acid growth along the root axis and to determine whether these differences were due to alterations in cell turgor or cell wall properties. The apoplast pH of maize roots growing in hydroponics was altered from pH 7.0 to pH 3.4 using 2 mol m^-3 citrate-phosphate buffer or unbuffered solutions. Whole root elongation rate rapidly increased and measurement of the local growth profile indicated that this increase in growth occurred in young cells in the accelerating zone (apical 0-4 mm) while more proximal growing cells were unaffected. Unbuffered solutions of identical pH produced qualitatively similar results. Single cell turgor pressures were unchanged between pH treatments both longitudinally and radially in the root tip. This suggests that the rapid acid-induced changes in growth rate were due to an increase in cell wall loosening. Single cell osmotic pressure and water potential were not significantly different between pH treatments. Acid pH caused net solute import at the root tip to increase 3- to 4-fold, which, coupled with the maintenance of turgor and osmotic pressure, indicated that solute import was not limiting expansion. Thus, acidic solutions cause an increase in growth in accelerating but not decelerating regions. It has been shown for the first time that acid growth in intact, growing roots is not due to differences in turgor, assigning these changes to cell wall properties. Possible cell wall biochemical alterations are discussed.     

Silicon-induced changes in viscoelastic properties of sorghum root cell walls

Silicon is deposited in the endodermal tissue in sorghum (Sorghum bicolor L. Moench) roots. Its deposition is thought to protect vascular tissues in the stele against invasion by parasites and drying soil via hardening of endodermal cells. We studied the silicon-induced changes in mechanical properties of cell walls to clarify the role of silicon in sorghum root. Sorghum seedlings were grown in nutrient solution with or without silicon. The mechanical properties of cell walls were measured in three separated root zones: basal, apical and subapical. Silicon treatment decreased cell-wall extensibility in the basal zone of isolated stele tissues covered by endodermal inner tangential walls. The silicon-induced hardening of cell walls was also measured with increases in elastic moduli (E) and viscosity coefficients (eta). These results provided new evidence that silicon deposition might protect the stele as a mechanical barrier by hardening the cell walls of stele and endodermal tissues. In contrast to the basal zone, silicon treatment increased cell-wall extensibility in the apical and subapical zones with concomitant decrease in E and eta. Simultaneously, silicon promoted root elongation. When root elongation is promoted by silicon, one of the causal factors maybe the silicon-enhanced extensibility of cell walls in the growing zone.     

Chemical analysis and immunolocalisation of lignin and suberin in endodermal and hypodermal/rhizodermal cell walls of developing maize (Zea mays L.) primary roots

The composition of suberin and lignin in endodermal cell walls (ECWs) and in rhizodermal/hypodermal cell walls (RHCWs) of developing primary maize (Zea mays L.) roots was analysed after depolymerisation of enzymatically isolated cell wall material. Absolute suberin amounts related to root length significantly increased from primary ECWs (Casparian strips) to secondary ECWs (suberin lamella). During further maturation of the endodermis, reaching the final tertiary developmental state characterised by the deposition of lignified secondary cell walls (u-shaped cell wall deposits), suberin amounts remained constant. Absolute amounts of lignin related to root length constantly increased throughout the change from primary to tertiary ECWs. The suberin of Casparian strips contained high amounts of carboxylic and 2-hydroxy acids, and differed substantially from the suberin of secondary and tertiary ECWs, which was dominated by high contents of ω-hydroxycarboxylic and 1,ω-dicarboxylic acids. Furthermore, the chain-length distribution of suberin monomers in primary ECWs ranged from C₁₆ to C₂₄, whereas in secondary and tertiary ECWs a shift towards higher chain lengths (C₁₆ to C₂₈) was observed. The lignin composition of Casparian strips (primary ECWs) showed a high syringyl content and was similar to lignin in secondary cell walls of the tertiary ECWs, whereas lignin in secondary ECWs contained higher amounts of p-hydroxyphenyl units. The suberin and lignin compositions of RHCWs rarely changed with increasing root age. However, compared to the suberin in ECWs, where C₁₆ and C₁₈ were the most prominent chain lengths, the suberin of RHCWs was dominated by the higher chain lengths (C₂₄ and C₂₆). The composition of RHCW lignin was similar to that of secondary-ECW lignin. Using lignin-specific antibodies, lignin epitopes were indeed found to be located in the Casparian strip. Surprisingly, the mature suberin layers of tertiary ECWs contained comparable amounts of lignin-like epitopes. Received: 19 August 1998 / Accepted: 3 February 1999     

Osmotic stress-induced changes in cell wall peroxidase activity and hydrogen peroxide level in roots of rice seedlings

The changes in activity of peroxidase (POD) extracted from the cellwalls and the level of H₂O₂ in rice seedling rootstreatedwith mannitol and their correlation with root growth were investigated.Increasing concentrations of mannitol from 92 to 276 mM, which isiso-osmotic with 50 to 150 mM NaCl, progressively reduced rootgrowth and increased POD activities extracted from the cell walls of riceroots.The reduction of growth was also correlated with an increase inH₂O₂ level. Both diamine oxidase (DAO) and NADHperoxidase(NADH-POD) are known to be responsible for the generation ofH₂O₂. Mannitol treatment increased DAO but not NADH-PODactivities in roots of rice seedlings, suggesting that DAO contributes to thegeneration of H₂O₂ in the cell walls of mannitol-treatedroots. An increase in the level of H₂O₂ and the activityof POD extracted from the cell walls of rice roots preceded root growthreduction caused by mannitol. An increase in DAO activity coincided with anincrease in H₂O₂ in roots caused by mannitol. Since DAOcatalyses the oxidation of putrescine, the demonstration that mannitolincreasesthe activity of DAO in roots is consistent with those that mannitol decreasesthe level of putrescine. In conclusion, cell-wall stiffening catalysed by PODispossibly involved in the regulation of root growth reduction caused bymannitol.     

The lithium-chloride-soluble cell-wall layers of Chlamydomonas reinhardii contain several immunologically related glycoproteins

Cell-wall glycoproteins of the unicellular green alga Chlamydomonas reinhardii have been purified from LiCl extracts of intact cells by gel exclusion chromatography and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Antibodies were raised against several polypeptide components isolated from the LiCl extracts. All these antibodies specifically reacted with the cell surface of formaldehyde-fixed cells. They showed cross-reactivity with the different antigens and were also reactive against some other polypeptides present in the LiCl extracts of intact wild-type cells as shown by double-diffusion assays and immunoblot analyses. These antigens were largely missing in LiCl extracts from the cell-wall-deficient mutant CW-15. The pattern of immunologically related cell-wall polypeptides of C. reinhardii varied during the vegetative cell cycle and was found to be also dependent on the growth conditions. Dot-immunobinding assays on chemically modified cell-wall glycoproteins demonstrated differences between the various antibodies with respect to their specificities. Differences were observed especially with respect to their reactivities against chemically deglycosylated cell-wall polypeptides. Chemical deglycosylation generally reduced the binding of the different antibodies indicating that all these antibodies recognize carbohydrate side chains. Only two of these antibody preparations, raised against cell-wall glycoproteins of relative molecular mass 35 and 150 kilodaltons, were found to be strongly reactive against deglycosylated cell-wall polypeptides. When these antibodies were saturated with cell-wall-derived glycopeptides in order to abolish the binding to carbohydrate side chains, they still recognized the same cell-wall polypeptides as did the untreated antibodies. These findings indicate that the cross-reactivity of the different cell-wall polypeptides with the antibodies is not exclusively the consequence of similar glycosylation patterns but is also the result of the presence of similar structures within the non-glycosylated stretches of the polypeptide backbones. Cell walls isolated from growing tobacco pollen tubes contained a single polypeptide component which showed crossreactivity with the antibodies to the cell-wall glycoproteins of C. reinhardii.Abbreviations BSA bovine serum albumin - IgG immunoglobulin G - kDa kilodalton - M_r relative molecular mass - PBS phosphate-buffered saline - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol     

Cell wall peroxidase against ferulic acid, lignin, and NaCl-reduced root growth of rice seedlings

The changes in cell wall peroxidase activity against ferulic acid (FPOD) and lignin level in roots of NaCl-stressed rice seedlings and their correlation with root growth were investigated. Increasing concentrations of NaCl from 50 to 150 mmol L⁻¹ progressively decreases root growth. The reduction of root growth by NaCl is closely correlated with the increase in FPOD activity extracted from the cell wall. In contrast, lignin level was reduced by NaCl. Since proline and ammonium accumulations are associated with root growth inhibition caused by NaCl, we determined the effect of proline or NH₄Cl on root growth and FPOD in roots. Exogenous application of NH₄Cl or proline markedly inhibited root growth and increased FPOD activity in rice seedlings in the absence of NaCl. An increase in FPOD activity in roots preceded inhibition of root growth caused either by NaCl, NH₄Cl, or proline. Our results suggest that cell-wall stiffening catalyzed by FPOD may participate in the regulation of root growth reduction of rice seedlings caused by NaCl.     

The kinetics of aluminum adsorption and desorption by root cell walls of an aluminum resistant wheat (Triticum aestivum L.) cultivar

In the present study, root cell walls were extracted from an Al-resistant wheat (Triticum aestivum L.) cultivar, Atlas 66 to investigate the effect of cell-wall properties on the kinetics of Al adsorption and desorption. Nearly all the Al adsorbed was desorbed by 2.5 mM CaCl₂ at pH 4.5, indicating that most of Al ions were electrically bound to cell wall materials. After the cell walls were treated with 1% pectinase for 30 min to degrade part of pectin, the total amount of Al absorbed was decreased by about 50%, indicating that pectin in the cell walls played an important role in binding Al. When the cell walls were preincubated in 1 and 10 mM malate solution overnight to mimic the organic acid secretion by the roots of wheat, the total amount of Al adsorbed was decreased by 60 and 80%, respectively, suggesting that the malate secreted in response to Al stress not only detoxifies Al by its chelating effect, but also reduce the cell walls' capacity to bind Al.