Aluminum inhibits the H(+)-ATPase activity by permanently altering the plasma membrane surface potentials in squash roots

Although aluminum (AL) toxicity has been widely studied in monocotyledonous crop plants, the mechanism of Al impact on economically important dicotyledonous plants is poorly understood. Here, we report the spatial pattern of Al-induced root growth inhibition, which is closely associated with inhibition of H(+)-ATPase activity coupled with decreased surface negativity of plasma membrane (PM) vesicles isolated from apical 5-mm root segments of squash (Cucurbita pepo L. cv Tetsukabuto) plants. High-sensitivity growth measurements indicated that the central elongation zone, located 2 to 4 mm from the tip, was preferentially inhibited where high Al accumulation was found. The highest positive shifts (depolarization) in zeta potential of the isolated PM vesicles from 0- to 5-mm regions of Al-treated roots were corresponded to pronounced inhibition of H(+)-ATPase activity. The depolarization of PM vesicles isolated from Al-treated roots in response to added Al in vitro was less than that of control roots, suggesting, particularly in the first 5-mm root apex, a tight Al binding to PM target sites or irreversible alteration of PM properties upon Al treatment to intact plants. In line with these data, immunolocalization of H(+)-ATPase revealed decreases in tissue-specific H(+)-ATPase in the epidermal and cortex cells (2--3 mm from tip) following Al treatments. Our report provides the first circumstantial evidence for a zone-specific depolarization of PM surface potential coupled with inhibition of H(+)-ATPase activity. These effects may indicate a direct Al interaction with H(+)-ATPase from the cytoplasmic side of the PM.     

Apoplastic binding of aluminum is involved in silicon-induced amelioration of aluminum toxicity in maize

The alleviating effect of silicon (Si) supply on aluminum (Al) toxicity was suggested to be based on ex or in planta mechanisms. In our experiments with the Al-sensitive maize (Zea mays) cultivar Lixis, Si treatment but not Si pretreatment ameliorated Al-induced root injury as revealed by less root-growth inhibition and callose formation. Si treatment did not affect monomeric Al concentrations in the nutrient solution, suggesting an in planta effect of Si on Al resistance. A fractionated analysis of Si and Al in the 1-cm root apices revealed that more than 85% of the root-tip Al was bound in the cell wall. Al contents in the apoplastic sap, the symplastic sap, and the cell wall did not differ between -Si and +Si plants. Si did not affect the Al-induced exudation of organic acid anions and phenols from the root apices. However, Al treatment greatly enhanced Si accumulation in the cell wall fraction, reducing the mobility of apoplastic Al. From our data we conclude that Si treatment leads to the formation of hydroxyaluminumsilicates in the apoplast of the root apex, thus detoxifying Al.     

Aluminium rhizotoxicity in maize grown in solutions with Al3+ or Al(OH)-4 as predominant solution Al species

The rhizotoxicity of aluminium at low-pH with Al(3+) and at high pH with Al(OH)-(4) as the main Al species was studied. Aluminium reduced root growth to similar levels at pH 8.0 and pH 4.3, although the mononuclear Al concentration at pH 8.0 was three times lower than at pH 4.3. Al contents of root apices were much higher at pH 8 than at pH 4.3. Callose was induced only marginally at pH 8 and the formation was confined to the epidermis, whereas it proceeded through the cortex with time at pH 4.3. Well-documented genotypical differences in callose formation and Al accumulation could not be found at pH 8. The largest fraction of the root-tip Al was recovered in the cell-wall fraction independent of the solution pH. A sequential extraction of isolated cell walls suggests that most of the cell-wall Al was precipitated Al(OH)(3) at pH 8.0. This can be explained by a drastic pH reduction in the root apoplastic sap to 6.2, whereas at bulk solution pH 4.3 it rose to 5.6. Al precipitation was also confirmed by the microscopic localization of Al. At pH 8, Al could mostly be found in the epidermis, but in the apoplast of the outer cortex at pH 4.3. It is proposed here that at pH 4.3, Al(3+) inhibits root growth through binding to sensitive binding sites in the apoplast of the epidermis and the outer cortex. At pH 8, Al(OH)(3) precipitation in the epidermis causes a mechanical barrier thus impairing the root-growth control of the epidermis.     

Aluminum Inhibition of the Inositol 1,4,5-Trisphosphate Signal Transduction Pathway in Wheat Roots: A Role in Aluminum Toxicity?

In crop plants, aluminum (Al) rhizotoxicity is a major problem worldwide; however, the cause of Al toxicity remains elusive. The effects of Al on the inositol 1,4,5-trisphosphate (Ins[1,4,5]P3)-mediated signal transduction pathway were investigated in wheat roots. Exogenously applied Al (50 [mu]M) rapidly inhibited root growth (<2 hr) but did not affect general root metabolism. An Ins(1,4,5)P3 transient was generated in root tips, either before or after exposure to Al for 1 hr, by treating the roots with H2O2 (10 mM). Background (unstimulated) levels of Ins(1,4,5)P3 were similar in both Al-treated and Al-untreated root apices. However, H2O2-stimulated levels of Ins(1,4,5)P3 in root apices showed a significant (>50%) reduction after Al exposure in comparison with untreated controls, indicating that Al may be interfering with the phosphoinositide signaling pathway. When phospholipase C (PLC) was assayed directly in the presence of Al or other metal cations in microsomal membranes, AlCl3 and Al-citrate specifically inhibited PLC action in a dose-dependent manner and at physiologically relevant Al levels. Al exposure had no effect on inositol trisphosphate dephosphorylation or on a range of enzymes isolated from wheat roots, suggesting that Al exposure may specifically target PLC. Possible mechanisms of PLC inhibition by Al and the role of Ins(1,4,5)P3 in Al toxicity and growth are discussed. This study provides compelling evidence that the phytotoxic metal cation Al has an intracellular target site that may be integrally involved in root growth.     

Early events responsible for aluminum toxicity symptoms in suspension-cultured tobacco cells

We investigated the aluminum (Al)-induced alterations in zeta potential, plasma membrane (PM) potential and intracellular calcium levels to elucidate their interaction with callose production induced by Al toxicity. A noninvasive confocal laser microscopy has been used to analyse the live tobacco (Nicotiana tabacum) cell events by means of fluorescent probes Fluo-3 acetoxymethyl ester (intracellular calcium) and DiBAC4 (PM potential) as well as to monitor callose accumulation. Log-phase cells showed no detectable changes in the PM potential during the first 30 min of Al treatment, but sustained large depolarization from 60 min onwards. Measurement of zeta potential confirmed the depolarization effect of Al, but the kinetics were different. The Al-treated cells showed a moderate increase in intracellular Ca2+ levels and callose production in 1 h, which coincided with the time course of PM depolarization. Compared with the Al treatment, cyclopiazonic acid, an inhibitor of endoplasmic reticulum Ca(2+)-ATPase, facilitated a higher increase in intracellular Ca2+ levels, but resulted in accumulation of only moderate levels of callose. Calcium channel modulators and Al induced similar levels of callose in the initial 1 h of treatment. Callose production induced by Al toxicity is dependent on both depolarization of the PM and an increase in intracellular Ca2+ levels.     

Aluminum-induced deposition of (1,3)-β-glucans (callose) inTriticum aestivum L.

Aluminum (Al)-induced damage to leaves and roots of two Al-resistant (cv. Atlas 66, experimental line PT741) and two Al-sensitive (cv. Scout 66, cv. Katepwa) lines ofTriticum aestivum L. was estimated using the deposition of (1, 3)--glucans (callose) as a marker for injury. Two-day-old seedlings were grown for forty hours in nutrient solutions with or without added Al, and callose deposition was quantified by spectrofluorometry (0–1000 µM Al) and localized by fluorescence microscopy (0 and 400 µM Al). Results suggested that Al caused little damage to leaves. No callose was observed in leaves with up to 400 µM Al treatment. In contrast, root callose concentration increased with Al treatment, especially in the Al-sensitive lines. At 400 µM Al, root callose concentration of Al-sensitive Scout 66 was nearly four-fold that of Al-resistant Atlas 66. After Al treatment, large callose deposits were observed in the root cap, epidermis and outer cortex of root tips of Scout 66, but not Atlas 66. The identity of callose was confirmed by a reduced fluorescence in Al-treated roots: firstly, after adding an inhibitor of callose synthesis (2-deoxy-D-glucose) to the nutrient solution, and secondly, after incubating root sections with the callosedegrading enzyme -D-glucoside glucohydrolase [EC 3.2.1.21]. Root callose deposition may be a good marker for Al-induced injury due to its early detection by spectrofluorometry and its close association with stress perception.Abbreviations DDG 2-deoxy-D-glucose - PAS periodic acid - Schiffs reagent - PE pachyman equivalents     

Sorption of Aluminum to Plasma Membrane Vesicles Isolated from Roots of Scout 66 and Atlas 66 Cultivars of Wheat

To further elucidate the mechanisms of differential genotypic tolerance to Al, plasma membrane (PM) vesicles were isolated from whole roots, root tips, and tipless roots of Al3+-sensitive and Al3+-tolerant cultivars (cv) of wheat (Triticum aestivum L. cv Scout 66 and cv Atlas 66, respectively). Vesicles from cv Scout root tips sorbed more Al than vesicles prepared from any other source. The intrinsic surface-charge density of vesicles isolated from cv Scout was 26% more negative than vesicles from cv Atlas (-37.2 versus -29.5 millicoulombs m-2). Growth experiments indicated that cv Scout is slightly more sensitive to La3+ than is cv Atlas, that the cultivars are equally sensitive to H+, and that cv Atlas is slightly more sensitive to SeO42-. The difference in sensitivity to Al3+ was very large; for a 50% inhibition, a 16-fold greater activity of Al3+ was required for cv Atlas. Using a newly developed Gouy-Chapman-Stern model for ion sorption to the PM together with growth-response curves, we estimate that the difference in surface-charge density can account for the slightly greater sensitivity of cv Scout to cationic toxicants and the slightly greater sensitivity of cv Atlas to anionic toxicants. According to our estimates the differences in PM surface negativity and Al sorptive capacity probably account for some of the difference in sensitivity to Al3+, but the greater part of the difference probably arises from other tolerance mechanisms expressed in cv Atlas root tips that reduce the amount of Al3+ that can reach the PM.     

Quantitative determination of callose in tree roots

The formation of callose in tree roots has been suggested as a physiological indicator of aluminum (Al) toxicity. Quantifying callose in the roots in forest soils, however, is hampered by the presence of autofluorescent materials in the roots that disturb the measurement of callose by fluorescence spectrophotometry. Tannins in the roots cause these measurement problems. Here we report on the measurement of callose in the root apices of European chestnut (Castanea sativa) seedlings collected in an acidified forest soil. The callose was quantified with a modified protocol which included three washing steps with polyvinylpolypyrrolidone (PVPP) before the callose was extracted. This procedure reduced the autofluorescence by about 50%. With the use of water or ethanol alone, callose could be measured in only about 15% of the root samples, whereas with the use of PVPP callose could be determined in 95% of the samples. This improved method could help to evaluate the effects of Al toxicity on tree roots grown in forest soils, where callose is detected as a physiological indicator.     

Transgenic Arabidopsis thaliana plants expressing a β‐1,3‐glucanase from sweet sorghum (Sorghum bicolor L.) show reduced callose deposition and increased tolerance to aluminium toxicity

Seventy‐one cultivars of sweet sorghum (Sorghum bicolor L.) were screened for aluminium (Al) tolerance by measuring relative root growth (RRG). Two contrasting cultivars, ROMA (Al tolerant) and POTCHETSTRM (Al sensitive), were selected to study shorter term responses to Al stress. POTCHETSTRM had higher callose synthase activity, lower β‐1,3‐glucanase activity and more callose deposition in the root apices during Al treatment compared with ROMA. We monitored the expression of 12 genes involved in callose synthesis and degradation and found that one of these, SbGlu1 (Sb03g045630.1), which encodes a β‐1,3‐glucanase enzyme, best explained the contrasting deposition of callose in ROMA and POTCHETSTRM during Al treatment. Full‐length cDNAs of SbGlu1 was prepared from ROMA and POTCHETSTRM and expressed in Arabidopsis thaliana using the constitutive cauliflower mosaic virus (CaMV) 35S promoter. Independent transgenic lines displayed significantly greater Al tolerance than wild‐type plants and vector‐only controls. This phenotype was associated with greater total β‐1,3‐glucanase activity, less Al accumulation and reduced callose deposition in the roots. These results suggest that callose production is not just an early indicator of Al stress in plants but likely to be part of the toxicity pathway that leads to the inhibition of root growth.     

Isolation of aluminum-tolerant cell lines of tobacco in a simple calcium medium and their responses to aluminum

Aluminum (Al)-tolerant cell lines (ALT301 and ALT401) of tobacco were isolated in a simple calcium (Ca) solution from ethyl methane sulfonate (EMS)-treated suspension cultured tobacco cells ( Nicotiana tabacum L. cv. Samsun, a cell line SL) at the logarithmic phase of growth. A highly tolerant cell line ALT301 exhibited the accumulation of Al and the deposition of callose to the same extent as the parental SL cells during the exposure to Al. However, the Al-treated ALT301 cells grew much better than the Al-treated SL cells after transfer to Al-free growth medium. Compared to SL cells, ALT301 cells were more tolerant to toxicity of copper and iron, but not to that of lanthanum. These results suggest that ALT301 cells have an internal tolerance mechanism, which makes cells grow normally in spite of Al accumulation and Al-induced lesion represented by the deposition of callose. This tolerance mechanism seems also to be effective against copper and iron toxicity. A slightly tolerant cell line ALT401 also accumulated Al to the same degree as SL cells, but deposited significantly less callose than did SL cells (43% of SL). The growth of ALT401 cells after Al treatment was only slightly better than that of SL cells. Thus, it seems that ALT401 cells have a mechanism to protect cells only from the Al-induced deposition of callose, which is not enough to overcome the Al-induced inhibition of growth. The different phenotypes exhibited by these Al-tolerant cell lines suggest that the deposition of callose is not directly related to the inhibition of growth in Al-treated cells.     

Effect of aluminium on membrane properties of soybean (Glycine max) cells in suspension culture

Short-term responses of soybean (Glycine max) cells to aluminium (Al) were studied in suspension culture. Formation of callose was the most sensitive indicator of Al effects. As low as 5 µM Al induced callose formation and an increase in callose concentration could be measured as early as 15 min after beginning the Al treatment. Also membrane permeability was rapidly affected by Al. Potassium net-efflux was reduced by increasing Al concentrations up to 300 µM Al. Increasing the pH of the external solution from 4.3 to 5.3 enhanced callose formation, indicating more severe Al damage at pH 5.3, which is in agreement with a model on H⁺ amelioration of Al toxicity. Al did not initiate or enhance ferrous sulfate (FeSO₄)-promoted lipid peroxidation. The results indicate that the plasma membrane is a primary target of Al and that cell suspension culture is a powerful tool to study effects of Al on plant roots.     

Response of the plant root to aluminum stress: Analysis of the inhibition of the root elongation and changes in membrane function

Pea root elongation was strongly inhibited in the presence of a low concentration of Al (5 μM). In Al-treated root, the epidermis was markedly injured and characterized by an irregular layer of cells of the root surface. Approximately 30% of total absorbed Al accumulated in the root tip and Al therein was found to cause the inhibition of whole root elongation. Increasing concentrations of Ca²⁺ effectively ameliorated the inhibition of root elongation by Al and 1 mM of CaCl₂ completely repressed the inhibition of root elongation by 50 μM Al. The ameliorating effect of Ca²⁺ was due to the reduction of Al uptake. H⁺-ATPase and H⁺-PPase activity as well as ATP and PPidependent H⁺ transport activity of vacuolar membrane vesicles prepared from barley roots increased to a similar extent by the treatment with 50 μM AlCl₃. The rate of increase of the amount of H⁺-ATPase and H⁺-PPase was proportional to that of protein content measured by immunoblot analysis with antibodies against the catalytic subunit of the vacuolar H⁺-ATPase and H⁺-PPase of mung bean. The increase of both activities was discussed in relation to the physiological tolerance mechanism of barley root against Al stress.     

Does aluminium affect root growth of maize through interaction with the cell wall – plasma membrane – cytoskeleton continuum?

The mechanism of aluminium-induced inhibition of root elongation is still not well understood. It is a matter of debate whether the primary lesions of Al toxicity are apoplastic or symplastic. The present paper summarises experimental evidence which offers new avenues in the understanding of Al toxicity and resistance in maize. Application of Al for 1 h to individual 1 mm sections of the root apex only inhibited root elongation if applied to the first 3 apical mm. The most Al-sensitive apical root zone appeared to be the 1–2 mm segment. Aluminium-induced prominent alterations in both the microtubular (disintegration) and the actin cytoskeleton (altered polymerisation patterns) were found especially in the apical 1–2 mm zone using monoclonal antibodies. Since accumulation of Al in the root apoplast is dependent on the properties of the pectic matrix, we investigated whether Al uptake and toxicity could be modulated by changing the pectin content of the cell walls through pre-treatment of intact maize plants with 150 mM NaCl for 5 days. NaCl-adapted plants with higher pectin content accumulated more Al in their root apices and they were more Al-sensitive as indicated by more severe inhibition of root elongation and enhanced callose induction by Al. This special role of the pectic matrix of the cell walls in the modulation of Al toxicity is also indicated by a close positive correlation between pectin, Al, and Al-induced callose contents of 1 mm root segments along the 5 mm root apex. On the basis of the presented data we suggest that the rapid disorganisation of the cytoskeleton leading to root growth inhibition may be mediated by interaction of Al with the apoplastic side of the cell wall – plasma membrane – cytoskeleton continuum. This revised version was published online in June 2006 with corrections to the Cover Date.     

Genotypical differences in aluminum resistance of maize are expressed in the distal part of the transition zone. Is reduced basipetal auxin flow involved in inhibition of root elongation by aluminum?

Short-term Al treatment (90 microM Al at pH 4.5 for 1 h) of the distal transition zone (DTZ; 1-2 mm from the root tip), which does not contribute significantly to root elongation, inhibited root elongation in the main elongation zone (EZ; 2.5-5 mm from the root tip) to the same extent as treatment of the entire maize (Zea mays) root apex. Application of Al to the EZ had no effect on root elongation. Higher genotypical resistance to Al applied to the entire root apex, and specifically to the DTZ, was expressed by less inhibition of root elongation, Al accumulation, and Al-induced callose formation, primarily in the DTZ. A characteristic pH profile along the surface of the root apex with a maximum of pH 5.3 in the DTZ was demonstrated. Al application induced a substantial flattening of the pH profile moreso in the Al-sensitive than in the Al-resistant cultivar. Application of indole-3-acetic acid to the EZ but not to the meristematic zone significantly alleviated the inhibition of root elongation induced by the application of Al to the DTZ. Basipetal transport of exogenously applied [(3)H]indole-3-acetic acid to the meristematic zone was significantly inhibited by Al application to the DTZ in the Al-sensitive maize cv Lixis. Our results provide evidence that the primary mechanisms of genotypical differences in Al resistance are located within the DTZ, and suggest a signaling pathway in the root apex mediating the Al signal between the DTZ and the EZ through basipetal auxin transport.     

Aluminum-induced cell death in root-tip cells of barley

Aluminum-induced cell death was investigated in root-tip cells of barley (Hordeum vulgare). The growth of roots in 0.1-50 mM Al treatments was inhibited after 8 h treatments, and could not be recovered after 24 h recovery culture without Al. Viable detection with fluorescein diacetate-propidium iodide (FDA-PI) staining shows that most of the root-tip cells have lost viability. These results suggest that the irreversible inhibition of root growth after 8 h Al treatments or 24 h recovery culture is mainly caused by cell death. DNA ladders occurred in root tips only after 8 h Al treatments (0.1-1.0 mM), but no apoptotic bodies in root tips were observed. Thus, the cell death caused by Al stress is likely to be Al-induced programmed cell death (PCD). The reactive oxygen species (ROS) in root-tip cells measured by ultraweak luminescence indicated that the oxidation status in root-tip cells basically ceased after exposure to 10-50 mM Al for 24 h, but was very violent in the root-tip cells treated with 0.1-1.0 mM for 24 h. Exposure to 0.1-1.0 mM Al for 3-12 h led to ROS burst. Therefore, our results suggest that 0.1-1.0 mM Al treatments for 8 h induce cell death (Al-induced PCD) possibly via a ROS-activated signal transduction pathway, whereas 10-50 mM Al treatments may cause necrosis in the root-tip cells. These results have an important role for further studies on the mechanism of Al toxicity in plants.     

Induction of callose formation is a sensitive marker for genotypic aluminium sensitivity in maize

The screening of 37 Zea mays L. cultivars in nutrient solution using root elongation (24 h) as a parameter showed large genotypic differences in Al resistance among the genetic material evaluated.Callose concentrations in root tips were closely and positively related to Al-induced inhibition of root elongation. Therefore, Al-induced callose formation in root tips appears to be an excellent indicator of Al injury and can be used as a selection criteria for Al sensitivity. In contrast, aluminium concentrations in root tips were not related to Al-induced inhibition of root elongation, nor to Al-induced callose formation. Callose formation was also induced by short-term A1 treatment in root tip protoplasts, and the response of protoplasts clearly reflected the cultivar-specific response to Al of intact roots. This indicates that in maize, Al sensitivity is expressed on the protoplast level.     

Magnesium enhances aluminum-induced citrate secretion in rice bean roots (Vigna umbellata) by restoring plasma membrane H+-ATPase activity

We demonstrated that magnesium (Mg) can alleviate aluminum (Al) toxicity in rice bean [Vigna umbellata (Thunb.) Ohwi & Ohashi] more effectively than is expected from a non-specific cation response. Micromolar concentrations of Mg alleviated the inhibition of root growth by Al but not by lanthanum, and neither strontium nor barium at the micromolar level alleviates Al toxicity. Aluminum also induced citrate efflux from rice bean roots, and this response was stimulated by inclusion of 10 microM Mg in the treatment solution. The increase in the Al-induced citrate efflux by Mg paralleled the improvement in root growth, suggesting that the ameliorative effect of Mg might be related to greater citrate efflux. Vanadate (an effective H+-ATPase inhibitor) decreased the Al-induced citrate efflux, while addition of Mg partly restored the efflux. Mg addition also increased the activity of Al-reduced plasma membrane H+-ATPase, as well as helping to maintain the Mg and calcium contents in root apices. We propose that the addition of Mg to the toxic Al treatment helps maintain the tissue Mg content and the activity of the plasma membrane H+-ATPase. These changes enhanced the Al-dependent efflux of citrate which provided extra protection from Al stress.     

Callose in roots of Norway spruce (Picea abies (L.) Karst.) is a sensitive parameter for aluminium supply at a forest site (Höglwald)

Under controlled environmental conditions in nutrient solution experiments induction of non-constitutive callose in roots has been shown to be a symptom of aluminium (Al) toxicity. In the present study roots of Norway spruce were sampled from a forest site where soil conditions had been modified by acidic irrigation and liming (Höglwald Experiment in Bavaria, Germany). A significant positive relationship was found between the callose content in short roots and the Al concentration in the soil solution, particularly if free Al, rather than total concentrations of soluble Al, were used for prediction. At the same sites root growth of Norway spruce was not affected by free Al concentrations in the range of 2.5 to 199 µM Al. The results show that also under field conditions a positive relationship between Al supply and callose content can be established. In Norway spruce callose content in roots is a much more sensitive parameter for Al supply than root growth.     

Zonal responses of sensitive vs. tolerant wheat roots during Al exposure and recovery

Aluminium (Al) irreversibly inhibits root growth in sensitive, but not in some tolerant genotypes. To better understand tolerance mechanisms, seedlings from tolerant ('Barbela 7/72' line) and sensitive ('Anahuac') Triticum aestivum L. genotypes were exposed to AlCl(3) 185 μM for: (a) 24 h followed by 48 h without Al (recovery); (b) 72 h of continuous exposure. Three root zones were analyzed (meristematic (MZ), elongation (EZ) and hairy (HZ)) for callose deposition, reserves (starch and lipids) accumulation, endodermis differentiation and tissue architecture. Putative Al-induced genotoxic or cytostatic/mytogenic effects were assessed by flow cytometry in root apices. Tolerant plants accumulated less Al, presented less root damage and a less generalized callose distribution than sensitive ones. Starch and lipid reserves remained constant in tolerant roots but drastically decreased in sensitive ones. Al induced different profiles of endodermis differentiation: differentiation was promoted in EZ and HZ, respectively, in sensitive and tolerant genotypes. No ploidy changes or clastogenicity were observed. However, differences in cell cycle blockage profiles were detected, being less severe in tolerant roots. After Al removal, only the 'Barbela 7/72' line reversed Al-induced effects to values closer to the control, mostly with respect to callose deposition and cell cycle progression. We demonstrate for the first time that: (a) cell cycle progression is differently regulated by Al-tolerant and Al-sensitive genotypes; (b) Al induces callose deposition >3 cm above root apex (in HZ); (c) callose deposition is a transient Al-induced effect in tolerant plants; and (d) in HZ, endodermis differentiation is also stimulated only in tolerant plants, probably functioning in tolerant genotypes as a protective mechanism in addition to callose.     

Aluminium causes variable responses in actin filament cytoskeleton of the root tip cells of Triticum turgidum

Summary. The effects of aluminium on the actin filament (AF) cytoskeleton of Triticum turgidum meristematic root tip cells were examined. In short treatments (up to 2 h) with 50–1000 μM AlCl₃·6H₂O, interphase cells displayed numerous AFs arrayed in thick bundles that lined the plasmalemma and traversed the endoplasm in different directions. Measurements using digital image analysis and assessment of the overall AF fluorescence revealed that, in short treatments, the affected cells possessed 25–30% more AFs than the untreated ones. The thick AF bundles were not formed in the Al-treated cells in the presence of the myosin inhibitors 2,3-butanedione monoxime (BDM) and 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-7), a fact suggesting that myosins are involved in AF bundling. In longer Al treatments, the AF bundles were disorganised, forming granular actin accumulations, a process that was completed after 4 h of treatment. In the Al-treated cells, increased amounts of callose were uniformly deposited along the whole surface of the cell walls. In contrast, callose formed local deposits in the Al-treated cells in the presence of cytochalasin B, BDM, or ML-7. These results favour the hypothesis that the actomyosin system in the Al-treated cells, among other roles, participates in the mechanism controlling callose deposition. Correspondence and reprints: Department of Botany, Faculty of Biology, University of Athens, Athens 157 84, Greece.