Short-term aluminium-induced changes in barley root tips

The short-term exposure of barley roots to low Al concentration caused significant root growth inhibition and radial swelling of roots. During Al treatment, the radial expansion of root cells occurred in root tissues representing elongation zone and meristem. Both low pH and Al treatments caused significant disruption of cell membranes in swollen roots. In contrast to Evans blue uptake callose formation was observed only at higher Al concentrations and was detected in both swollen and adjacent root areas. Similarly to Al, exogenous short-term application of indole-3-acetic acid, polar transport inhibitor triiodobenzoic acid, ethylene precursor 1-aminocyclopropane-1-carboxylic acid or H₂O₂ evoked root growth inhibition and radial cell expansion in barley root tip too.     

Enhanced lipoxygenase activity is involved in the stress response but not in the harmful lipid peroxidation and cell death of short-term cadmium-treated barley root tip

Root growth inhibition and radial root swelling were the characteristic symptoms of barley root tips after the short-term exposure of roots to 15 and 30 μM Cd. Higher Cd concentrations caused extensive cell death and root growth arrest. Enhanced lipid peroxidation was observed as early as 1 h after the short-term treatment in a Cd concentration-dependent manner. In contrast to lipid peroxidation, the induction of lipoxygenase activity was detected only 3 h after the exposure of roots to 15 or 30 μM Cd. In addition, it was not observed in 60 μM Cd-treated root tips. The highest lipoxygenase activity was detected 6 h after 15 μM Cd treatment in the meristematic and elongation zone of root tip and was probably associated with the radial expansion of cells. Our results indicate that the upregulation of lipoxygenase is an important component of stress response in barley roots to toxic Cd. It is probably involved in the morphological stress response of root tips or/and in the alleviation of Cd-induced toxic alterations in plant cell membranes, but it is not responsible for the Cd-induced harmful lipid peroxidation and cell death.     

Cadmium disrupts apoplastic ascorbate redox status in barley root tips

Using a short-term Cd treatment (5–30 min), we analysed the effect of Cd on apoplastic ascorbate redox status and their regeneration during the recovery period in barley root tips. Root growth inhibition induced by 15 μM Cd was detectable after 5 min of exposure and increased in a time-dependent manner up to 15 min of exposure. High 30 μM Cd concentration completely inhibited root growth during the first 6 h after short-term treatment. In parallel with Cd-induced root growth inhibition, a rapid decrease of apoplastic ascorbate dehydroascorbate ratio was observed immediately after short-term treatments. During the recovery from 15 μM Cd short-term treatment, apoplastic ascorbate was rapidly regenerated to the control level in the first root segment containing meristem and elongation zone. In contrast to 15 μM Cd treatment, in 30 μM Cd-treated roots apoplastic ascorbate level was sustained at a significantly lower level compared to control roots. We confirmed that a decrease of apoplastic ascorbate/dehydroascorbate ratio in the elongation zone was associated with root growth inhibition or arrest.     

Abiotic stress�Cinduced inhibition of root growth and ascorbic acid oxidase activity in barley root tip is associated with enhanced generation of hydrogen peroxide

Using short-term treatments, the aim of this study was to analyze the role of hydrogen peroxide in the regulation of AAO activity during Cd, Cu or IAA treatments in barley root tips. For analysis individual barley root segments were obtained by the gradual cutting of each root from the tip to the base 1, 2, 3 or 6 h after short-term treatments. Already a short 30 min exposure of barley roots to Cd induced significant root growth inhibition in a Cd concentration dependent manner, which was accompanied by a marked reduction of AAO activity. At Cu concentration which had no effect on the root growth a significant increase in AAO activity was observed. This increased AAO activity was detected only in ionically-bound CW fraction. In contrast, Cu at higher concentration and IAA inhibited both ionically-bound CW AAO isozymes. Prompt inhibition of AAO activity immediately after short-term treatment was observed only in the case of H₂O₂ treatment suggesting that H₂O₂ may act as an inhibitor of AAO. This was further supported by the observation that all Cd-, Cu- or IAA-induced root growth and AAO activity inhibition in barley roots was connected with an elevated production of H₂O₂.     

Effects of 60-Hz electric fields on cellular elongation and radial expansion growth in cucurbit roots

Serial longitudinal and transverse sections were prepared from roots of Cucumis sativus and Cucurbita maxima that had been exposed/sham-exposed to 60-Hz electric fields for 0-2 days. Field exposures were selected to produce a 10-20% or a 70-80% growth inhibition in whole roots of both species. Cortical cell length and diameter were measured using a microscope and eyepiece micrometer; measurements were conducted "blind." In both species, inhibition of cellular elongation was associated with exposure to electric fields (EF). Cellular radial expansion was apparently unaffected by exposure to electric fields. The diameters of radially unexpanded or fully expanded C. sativus cortical cells were about 25-30% smaller than those of comparable cells in C. maxima roots. Previous studies of the relationship between rates of root growth and applied EF strength showed that the response thresholds of C. sativus and C. maxima differed by a similar relative amount. These results are consistent with the postulate that EF-induced effects in roots are elicited by induced transmembrane potentials.     

Low Cd concentration-activated morphogenic defence responses are inhibited by high Cd concentration-induced toxic superoxide generation in barley root tip

Exposure of roots to low Cd concentration induced morphogenic responses including the inhibition of root growth and the radial swelling of root tip. High Cd concentrations within a few minutes caused a robust induction of superoxide generation leading to the cell death and root growth arrest. This toxic superoxide generation blocked the development of low Cd concentration-activated morphogenic responses. While the morphogenic responses of roots to low Cd concentration are induced very rapidly and probably due to the interaction of Cd with the apoplast of root tissue, high Cd concentration-induced superoxide production required the entry of Cd into the symplast. Auxin signaling is involved in the activation of Cd-induced morphogenic defence responses but not in the Cd-induced toxic superoxide generation. These results suggest that oxidative stress is not a primary cause for the Cd-induced morphogenic responses such as growth reduction and radial cell expansion in barley root tips.     

Elevated oxalate oxidase activity is correlated with Al-induced plasma membrane injury and root growth inhibition in young barley roots

In order to characterise the possible mechanisms involved in Al toxicity some functional characteristics were analysed in young barley (Hordeum vulgare L.) seedlings cultivated between moistened filter paper. Transfer of germinated barley seeds into hydroponic culture system caused significant stress, which was manifested by root-growth inhibition and elevated Evans blue uptake of root tips. Hydroponics caused stress unabled the analysis of Al-induced stress in the young barley roots during the first day of cultivation. Several (3–4) days are required for adaptation of barley seedlings to hydroponics in spite of strong aeration of the medium. Using filter paper compared to cultivation in solution application of much higher Al concentrations were required to inhibit root growth. Al-induced root growth inhibition, Al uptake, damage of plasma-membrane (PM) permeability of root cells, as well as elevated oxalate oxidase - OxO (EC 1.2.3.4) activity were significantly correlated. While 1 mM Al concentration had no effect on barley roots growing on filter paper, 5 to 100 mM Al concentration inhibited root growth, enhanced cell death and induced oxalate oxidase activity with increasing intensity. The time course analysis of OxO gene expression and OxO activity showed that 10 mM Al increased OxO activity as soon as 3 h after exposure of roots to Al reaching its maximum at about 18 h after Al application. These results indicate that expression of OxO is activated very early after exposure of barley to Al, suggesting its role in oxidative stress and subsequent cell death caused by Al toxicity in plants.     

Superoxide production induced by short-term exposure of barley roots to cadmium, auxin, alloxan and sodium dodecyl sulfate

Key message

Abiotic stress-induced superoxide generation depending on its localization, level, duration and presumably also on the action of other signals may lead to different stress responses.

Abstract

The purpose of this study was to analyze the alterations in superoxide generation and morphogenesis following short-term Cd, IAA and alloxan treatments, during stress and recovery period in barley root tips. At low Cd concentration the transient accumulation of superoxide in the epidermal cells was accompanied by root growth inhibition and radial expansion of cortical cells in the elongation zone of root tips. These morphological changes were very similar to the externally applied IAA-induced responses. However, the role of superoxide generated in the epidermal cells by low concentration of Cd and IAA is probably alone not sufficient for the induction of these processes. SDS as an activator of NOX activity caused a strong accumulation of superoxide in the epidermal cells along the whole root apex but without any changes in root morphology and growth. On the other hand, higher Cd concentrations as well as alloxan stimulated the generation of superoxide in the cortical tissue of the elongation zone of root tip, which was accompanied by the induction of cell death. Our results suggest that enhanced superoxide generation, depending on its localization, level, duration and presumably also on the action of other signals, may lead to altered root morphology (15?μM Cd or IAA), root growth inhibition (alloxan), transient root growth cessation (30?μM Cd) or to the death of cells/root at higher (60?μM) Cd concentrations.     

Auxin signalling is involved in cadmium-induced glutathione-S-transferase activity in barley root

Transient exposure of barley roots to Cd, IAA or H₂O₂ for 30 min resulted in a significant root growth inhibition. Cd significantly increased the GST activity of roots 6 h after the end of short-term treatment. This increase was more relevant in root segment containing differentiation zone than in root segment just immediately behind the root apex. In contrast to Cd treatment, the short-term exposure of barley roots to IAA resulted in a significant increase of GST activity along the whole root tip and this increase was detectable already 3 h after the treatment with 10 μM IAA. Similarly to IAA, exogenously applied 10 mM H₂O₂ for 30 min caused significant increase of GST activity along the whole root tip 6 h after the treatment. This increase was already detectable 3 h after the exposure, but only in the differentiation zone of root tip. Auxin influx or signalling inhibitor considerable decreased the Cd- or IAA-induced GST activity in barley root tips. The strong activation of GST even after a brief exposure of barley roots to Cd support the crucial role of GST in the Cd-induced stress response in which presumably IAA and H₂O₂ play an important signalling role including the activation of GST.     

Gibberellin-ethylene interaction controls radial expansion in citrus roots

The involvement of gibberellins (GAs) and ethylene in the process of root radial expansion was studied in young seedlings of Carrizo citrange [Citrus sinensis (L.) Osb. × Poncirus trifoliata (L.) Raf.]. The GA inhibitors cycocel, paclobutrazol, and tetcyclacis enhanced radial expansion of the root tip (up to 2.3-fold) as a result of increases in stele diameter and inner cortex width. The GA deficiency increased cell number and width, and changed the polarity of growth, generating wider and shorter cortical cells in the elongation zone. In the presence or absence of GA inhibitors, GA₃ decreased root tip width and reduced all parameters related to radial expansion. The ethylene inhibitors (aminooxyacetic acid; cobalt ions, CoCl₂; silver thiosulfate) suppressed swelling induced by GA deficiency, generating thinner cells just as GA₃ did. In contrast to GA₃, ethylene inhibitors produced longer cells strongly resembling those of the untreated seedlings. Ethylene released by ethephon did not modify root tip width in control plants, while root diameter behind the root tip was increased. In the presence of low and ineffective concentrations of cycocel, the ethylene precursor 1-aminocyclopropane-1-carboxylic acid increased radial expansion of root tips (1.3-fold) and changed the polarity of growth, producing wider and shorter inner cortical cells as GA inhibitors did. These observations imply, first, that ethylene is the hormonal effector of the process of root radial expansion and, second, that the endogenous GAs modulate the promotive response of ethylene. Received: 4 October 1996 / Accepted: 25 December 1996     

ROOT CONTRACTION IN HYACINTH. I. EFFECTS OF IAA ON DIFFERENTIAL CELL EXPANSION

Contractile roots of Hyacinthus orientalis L. cv ‘Pink Pearl’ shorten as a result of growth of inner cortical cells which expand radially and contract longitudinally. Brief treatment with IAA (indole-3-acetic acid—0.5 and 1.0 mg/1) induces subapical swelling, root cap proliferation and decreased rates of elongation in potentially contractile roots. Growth resumes with removal of IAA from the culture medium and contraction subsequently occurs. The pattern of subsequent contraction is affected by prior IAA treatment; contraction occurs in the normal manner both acropetal and basipetal to the points of IAA-induced swelling, but does not occur in the swollen region itself. Microscopic examination of the swollen region reveals that cells of the middle and outer cortex are radially expanded and longitudinally shortened relative to middle and outer cortical cells of contracted and uncontracted portions of the same root and control roots. In contrast, inner cortical cells in swollen regions of IAA-treated roots show approximately 50% less radial expansion than inner cortical cells of control contracted roots. Middle and outer cortical cells in the swollen region of IAA-treated roots undergo radial expansion, while middle and outer cortical cells in adjacent contracting zones are compressed by radially expanding inner cortical cells. Average volumes of cortical cells in the IAA-induced swollen region increased approximately two-fold when contraction occurred in adjacent regions. These results suggest that in hyacinth roots, under certain circumstances, inner and outer cortical cells alike possess the ability for growth reorientation and expansion. However, during the usual course of contractile root development, cells of the outer cortex are restricted in this ability, through an as yet unknown mechanism, and are passively compressed by the radially expanding inner cortical cells.     

BOTERO1 is required for normal orientation of cortical microtubules and anisotropic cell expansion in Arabidopsis

Mutants at the BOTERO1 locus are affected in anisotropic growth in all non-tip-growing cell types examined. Mutant cells are shorter and broader than those of the wild type. Mutant inflorescence stems show a dramatically reduced bending modulus and maximum stress at yield. Our observations of root epidermis cells show that the cell expansion defect in bot1 is correlated with a defect in the orientation of the cortical microtubules. We found that in cells within the apical portion of the root, which roughly corresponds to the meristem, microtubules were loosely organized and became much more highly aligned in transverse arrays with increasing distance from the tip. Such a transition was not observed in bot1. No defect in microtubule organization was observed in kor-1, another mutant with a radial cell expansion defect. We also found that in wild-type root epidermal cells, cessation of radial expansion precedes the increased alignment of cortical microtubules into transverse arrays. Bot1 roots still show a gravitropic response, which indicates that ordered cortical microtubules are not required for differential growth during gravitropism. Interestingly, the fact that in the mutant, these major changes in microtubule organization cause relatively subtle changes in cell morphology, suggest that other levels of control of growth anisotropy remain to be discovered. Together, these observations suggest that BOT1 is required for organizing cortical microtubules into transverse arrays in interphase cells, and that this organization is required for consolidating, rather than initiating, changes in the direction of cell expansion.     

Tissue-specific expression of the ethylene biosynthetic machinery regulates root growth in maize

Although the hormonal control of root growth and development has been extensively studied, relatively little is known about the role that ethylene plays in cereal root development. In this work, we have investigated how the ethylene biosynthetic machinery is spatially regulated in maize roots and how changes in its expression alter root growth. ACC synthase (ZmACS) expression was observed in the root cap and in cortical cells whereas ACC oxidase (ZmACO) expression was detected in the root cap, protophloem sieve elements, and the companion cells associated with metaphloem sieve elements. Roots from Zmacs6 mutants exhibited significantly reduced ethylene production, a smaller root cap of increased cell number but smaller cell size, accelerated elongation of metaxylem, cortical, and epidermal cells, and increased vacuolation of cells in the calyptrogen of the root cap, phenotypes that were complemented by exogenous ACC. Zmacs6 mutant roots exhibited increased growth when largely unimpeded, a phenotype complemented by exogenous ACC, whereas loss of ZmACS2 expression had less of an effect. In contrast, Zmacs6 plants exhibited reduced root growth in soil. These results suggest that expression of ZmACS6 is important in regulating growth of maize roots in response to physical resistance.     

Effect of cadmium on growth and respiration of barley plants grown under two temperature regimes

We studied cadmium effect on the respiratory pathways ratio in the organs of barley (Hordeum distichum L., cv. Novichok) plants grown in water culture at two temperature regimes. Mineral nutrients were supplied daily in exponentially increasing amounts in order to provide for steady-state growth. CdSO₄ (30, 60, or 100 μmol/l) was added to nutrient solution at a single time in the beginning of the exponential growth period (19 days after germination). In further 6 days, the relative growth rate and biomass accumulation declined stronger with the increase in the cadmium concentration in plants grown at 13/8°C (day/night) than at 21/17°C (day/night). Cadmium suppressed root respiration (down to 60% of control) stronger than leaf respiration, and the roots manifested a higher sensitivity to the inhibitor of alternative oxidase, salicylhydroxamic acid. The respiratory pathways ratio in the roots occurred against the background of activated lipid peroxidation (POL). The highest POL activity and the highest proportion of alternative respiration pathway (AP) (up to 46% of total respiration) were observed in the roots in the presence of the highest cadmium concentration (100 μM) under lower temperature (13/8°C). Thus, high cadmium concentrations affected strongly the total rate of respiration and respiratory pathways ratio. Growth temperature modulated Cd effects on respiration. AP activation is one of the mechanisms for maintenance of root cell homeostasis under cadmium-induced stress.     

Growth responses of barley and wheat seedlings to lead and cadmium

The responses of barley and wheat seedlings to lead and cadmium ions in relation to the concentration and duration of treatment were studied. Both metals inhibited seed germination and growth of roots and shoots, but the toxic effect of cadmium was observed at lower concentrations. Inhibition of seedling growth was already recorded already within a day after the beginning of the treatment, and then increased further. The sensitivity of the processes studied to both the metals decreased in the order: root growth, shoot growth and seed germination. The resistance of barley and wheat to lead was similar, whereas the resistance to cadmium was higher in barley     

Cadmium-induced microsomal membrane-bound peroxidases mediated hydrogen peroxide production in barley roots

The effect of cadmium on microsomal membrane-bound peroxidases and their involvement in hydrogen peroxide production was studied in barley roots. One anionic and two cationic peroxidases were detected, which were strongly activated by Cd treatment. Positive correlation was found between root growth inhibition and increased peroxidase, NADH oxidase activity and H₂O₂ generation in root microsomal membrane fraction of Cd-treated barley roots.     

Cadmium-Induced Inhibition of Apoplastic Ascorbate Oxidase in Barley Roots

The effect of excess cadmium (Cd: 0.0, 0.25, 0.5, 1.0 and 2.0 mM) on growth and ascorbate oxidase (AO) activity was investigated in barley (Hordeum vulgare L. cv. Jubilant) roots. The study employed a filter-paper technique to germinate and grow the germinating seeds following imbibition with respective Cd treatments for 4 h at 25 °C in darkness. Cd was required at 1.0 mM to affect 50% root growth inhibition 72 h after the treatment. This Cd-induced root growth inhibition was accompanied by a corresponding loss of plasma membrane integrity in root cells as evaluated by Evans blue uptake. Excess Cd (1.0 and 2.0 mM) significantly inhibited the AO activity in all the analysed fractions of barley roots such as extracellular, soluble, cell wall (CW)- and membrane-bound fractions. AO was localized in the apoplast, and its highest specific activity was detected in the CW II fraction obtained by extraction with 1.0 M NaCl from purified cell walls. The analysis of AO isozyme profile showed that besides the reduced activity of two anionic and two cationic isozymes, one cationic AO isozyme was activated during excess Cd treatment, which could be detected in cell wall fractions CW II, III and IV.     

Ethylene enhances water transport in hypoxic aspen

Water transport was examined in solution culture grown seedlings of aspen (Populus tremuloides) after short-term exposures of roots to exogenous ethylene. Ethylene significantly increased stomatal conductance, root hydraulic conductivity (L(p)), and root oxygen uptake in hypoxic seedlings. Aerated roots that were exposed to ethylene also showed enhanced L(p). An ethylene action inhibitor, silver thiosulphate, significantly reversed the enhancement of L(p) by ethylene. A short-term exposure of excised roots to ethylene significantly enhanced the root water flow (Q(v)), measured by pressurizing the roots at 0.3 MPa. The Q(v) values in ethylene-treated roots declined significantly when 50 microM HgCl(2) was added to the root medium and this decline was reversed by the addition of 20 mM 2-mercaptoethanol. The results suggest that the response of Q(v) to ethylene involves mercury-sensitive water channels and that root-absorbed ethylene enhanced water permeation through roots, resulting in an increase in root water transport and stomatal opening in hypoxic seedlings.     

Ethephon and auxin induce mycorrhiza-like changes in the morphology of root organ cultures of Mugo pine

The effects of ethylene and auxin on the morphology and anatomy of root organ cultures of Pinus mugo Turra var. mugo were investigated to test the hypothesis that changes in root morphology associated with formation of ectomycorrhizae may be related to ethylene produced by ectomycorrhizal fungi or by host plant roots in response to fungus-produced auxin. Morphological changes characteristic of mycorrhizal infection include dichotomous branching of lateral roots, inhibition of root hair formation and enlargement of cortical cells. Lateral roots on non-mycorrhizal root organ cultures, grown in a defined medium, underwent dichtotomous branching while root hair formation was inhibited in response to the ethylene released by 50 and 100 μ M ethephon (2-chloroethylphosphonic acid), but no effect on cortical cell dimensions was observed. The auxin, naphthaleneacetic acid (1 and 10 μ M ) also stimulated dichotomous branching and inhibited root hair formation, but to a lesser extent and with a greater lag time than ethephon. Auxin-stimulated ethylene production by root organ cultures was demonstrated. This appeared to be responsible, at least in part, for the auxin-induced dichotomous branching since the ethylene action inhibitor, silver thiosulfate (0.1 m M ) inhibited the response to auxin by 35%.