A comparative study on the aluminium- and copper-induced organic acid exudation from wheat roots

It is well established that aluminium (Al) and some heavy metals can elicit organic acid exudation from a range of species. In the present research we found that copper (Cu) can also induce organic acid exudation from the roots of wheat, rye, triticale, maize and soybean. Using intact wheat plants, we made a comparative study of Al- and Cu- induced organic acid exudation. In 5-day-old wheat seedlings, severe Cu stress (40 µ M CuCl₂) mainly induced the exudation of malate and citrate, and Al-tolerant genotypes could release significantly greater amounts of malate than Al-sensitive genotypes. The time course of the exudation of malate and citrate from the roots of 5-day-old seedlings of wheat (cv. Atlas) in 200 µ M AlCl₃ was similar to that in 40 µ M CuCl₂. In older wheat plants (15-day-old), moderate Cu stress (12 µ M CuCl₂) induced the exudation of large amounts of citrate and addition of Al or La sharply reduced Cu-induced citrate exudation, while Cu or La did not affect Al-induced malate efflux. When half of the root system of Atlas wheat was immersed in Al- or Cu-containing solution and the remaining half in Al- or Cu-free solution, organic acids were only exuded into the solution containing Al or Cu. This suggests that no long distance signal transport is involved in organic acid exudation induced by Al or Cu, and that direct contact of Al or Cu with plant roots is a prerequisite for the induction of organic acid exudation. The anion-channel inhibitor niflumic acid (NIF) significantly stimulated the exudation of both citrate and malate from 5-day-old wheat seedlings under severe Al or Cu stress. Our results suggest that Cu-induced organic acid efflux may be a common response, which may play a role in alleviating Cu toxicity in plants.     

Adenosine 5′-monophosphate decreases citrate exudation and aluminium resistance in Tamba black soybean by inhibiting the interaction between 14-3-3 proteins and plasma membrane H<Superscript>+</Superscript>-ATPase

Our previous study suggested that aluminium (Al) stress increased plasma membrane (PM) H⁺-ATPase activity and citrate secretion and simultaneously enhanced the interaction between 14-3-3 proteins and phosphorylated PM H⁺-ATPase in Al-resistant Tamba black soybean (RB). Adenosine 5′-monophosphate (AMP) is known as an inhibitor of the interaction between 14-3-3 proteins and PM H⁺-ATPases. To investigate the effects of AMP on Al resistance, PM H⁺-ATPase activity and citrate exudation, AMP was used to treat Al-stressed RB. The results showed that after treatment with either 100 μM AMP or 50 μM Al for 8 h, RB root growth was inhibited by approximately 50 and 30%, respectively. However, simultaneous treatment with 100 μM AMP and 50 μM Al for 8 h resulted in a 60% inhibition of RB root growth, indicating that the presence of AMP reduced Al tolerance in RB. The interaction of PM H⁺-ATPase and 14-3-3 proteins in the root tips of Al-treated RB was stronger than that in the untreated control. However, the interaction of the two proteins was greatly reduced (lower than that in the control) after co-treatment with Al and AMP, suggesting that the presence of AMP under Al stress reduced the Al-enhanced interaction between PM H⁺-ATPase and 14-3-3 proteins. Consequently, PM H⁺-ATPase activity decreased by approximately 50%, which led to a significant decrease in H⁺ efflux and citrate secretion in RB roots under Al stress. Collectively, these results indicate that AMP reduced citrate exudation and Al resistance in RB by inhibiting the interaction between 14-3-3 proteins and PM H⁺-ATPases under Al stress.     

Jasmonates modulate the promotion effects induced by SNP on root development of wheat under osmotic stress through lipoxygenase activation

We investigated promotion effects of exogenous sodium nitroprusside (SNP) on wheat seedling (Triticum aestivum L.) lateral root (LR) and root hair development, and the relationship between endogenous jasmonate (JA) production and activity changes of lipoxygenase (LOX) isoenzymes under osmotic stress generated by 15 % PEG-6000. Our results showed that 25 or 50 μM SNP could significantly increase LR length and number whether or not the seedlings were under PEG stress. When 50 μM cPTIO, 50 μM SHAM or 50 μM NDGA was supplemented, the promotion effects of SNP were blocked. SNP could also induce the production of endogenous JAs in roots, and 25 μM SNP induced the maximum JA content. The effect of SNP on JA production could also be blocked by adding cPTIO, SHAM or NDGA. Furthermore, the activity of lipoxygenase (LOX) in roots was affected by SNP; the maximal activity of LOX also occurred in the roots treated by 25 μM SNP under PEG stress, or 50 μM SNP without PEG stress. LOX isoenzymes in roots were detected by electrophoresis; the results showed that 25 μM SNP could noticeably increase the activities of LOXII and LOXIII under PEG stress. Our results suggest that, under osmotic stress generated by PEG, the promotion effects of exogenous SNP on wheat LR and root hair development could be mediated by endogenous JAs through LOX activation.     

Effect of K-252a and abscisic acid on the efflux of citrate from soybean roots

The Al-induced release of organic acid has been suggested as an important mechanism for Al resistance in plants. In this study, the effect of K-252a and abscisic acid (ABA) on the efflux of citrate was investigated in soybean (Glycine max L.) roots. Al initiated citrate efflux from the root apices 30 min after the addition of Al. The Al-triggered efflux of citrate was sensitive to metabolic inhibitors and anion channel inhibitors. Pretreatment or treatment with K-252a, an inhibitor of protein kinase, severely inhibited the Al-induced efflux of citrate accompanying an increase in Al accumulation and intensified Al-induced root growth inhibition. Al-treatment increased the endogenous level of abscisic acid (ABA) in soybean roots in a dose- and time-dependent manner, while K-252a failed to inhibit the Al-induced increase in endogenous ABA. Exogenous application of ABA increased the activity of citrate synthase (EC 4.1.3.7) by 26.2%, and decreased Al accumulation by 32.3%, respectively. ABA-induced increases in citrate efflux and root elongation were suppressed by K-252a, while ABA could not reverse the K-252a effects. Taken together, these results suggest that ABA is probably involved in the early response, after which K-252a-sensitive protein kinases play a key step in regulating the activity of an anion channel, through which citrate is released from the apical cells of soybean roots.     

Water stress and indol-3yl-acetic acid content of maize roots

Water-stress conditions were applied to the apical 12 mm of intact or excised roots ofZea mays L. (cv. LG 11) using mannitol solutions (0 to 0.66 M) and changes in weight, water content, growth and IAA level of these roots were investigated. With increasing stress a decrease in growth, correlated with an increased IAA level, was observed. The largest increase in IAA (about 2.7-fold) was found in the apical 5 mm of the root and was obtained under a stress corresponding to an osmotic potential of −1.39 MPa in the solution. This stress led to an isotonic state in the cells after 1 h. When the duration of water stress (−1.09 MPa) was increased to 2 or 3 h, no further increase in the IAA content was observed in the root segments. This indicated that there was no correlation between a hypothetical passive penetration of mannitol in the cells and IAA content. Indol-3yl-acetic acid rose to the same level in excised as in intact roots. In both cases, IAA accumulation was apparently independent of the hydrolysis of the conjugated form. The caryopsis and shoot seem not to be necessary to induce the increase of the IAA level in the roots during water stress (−1.09 MPa). Therefore, there seems to be a high rate of IAA biosynthesis in excised maize roots under water-stress conditions. Exodiffusion of IAA was observed during an immersion in either buffer or stress (−1.09 MPa) solution. In both cases, this IAA efflux into the medium represented about 50% of the endogenous level. Considering the present results, IAA appears to play an important part in the regulation of maize root metabolism and growth under water deficiency.     

Characterization of the TaALMT1 protein as an Al3+-activated anion channel in transformed tobacco (Nicotiana tabacum L.) cells

TaALMT1 encodes a putative transport protein associated with Al(3+)-activated efflux of malate from wheat root apices. We expressed TaALMT1 in Nicotiana tabacum L. suspension cells and conducted a detailed functional analysis. Protoplasts were isolated for patch-clamping from cells expressing TaALMT1 and from control cells (empty vector transformed). With malate(2-) as the permeant anion in the protoplast, an inward current (anion efflux) that reversed at positive potentials was observed in protoplasts expressing TaALMT1 in the absence of Al(3+). This current was sensitive to the anion channel antagonist niflumate, but insensitive to Gd(3+). External AlCl(3) (50 microM), but not La(3+) and Gd(3+), increased the inward current in TaALMT1-transformed protoplasts. The inward current was highly selective to malate over nitrate and chloride (P(mal) > P(NO3) >or= P(Cl), P(mal)/P(Cl) >or=18, +/-Al(3+)), under conditions with higher anion concentration internally than externally. The anion currents displayed a voltage and time dependent deactivation at negative voltages. Voltage ramps revealed that inward rectification was caused by the imposed anion gradients. Single channels with conductances between 10 and 17 pS were associated with the deactivation of the current at negative voltages, agreeing with estimates from voltage ramps. This study of the electrophysiological function of the TaALMT1 protein in a plant heterologous expression system provides the first direct evidence that TaALMT1 functions as an Al(3+)-activated malate(2-) channel. We show that the Al(3+)-activated currents measured in TaALMT1-transformed tobacco cells are identical to the Al(3+)-activated currents observed in the root cells of wheat, indicating that TaALMT1 alone is likely to be responsible for those endogenous currents.     

A Comparison of Aluminum Resistance among Polygonum Species Originating on Strongly Acidic and Neutral Soils

Two kinds of Polygonum species (Polygonum aviculare L. and Polygonum lapathifolium L.) grown in tea garden soils at pH around 3.5 and one Polygonum bungeanum Turcz grown in neutral soils were collected to investigate the mechanisms involved in their high Al resistance. Hydroponic experiments showed that the root elongation was only inhibited by 15% in P. aviculare and 35% in P. lapathifolium after exposure to 50 μM Al for 24 h. Their Al resistance was respectively higher than and similar to that in an Al resistant buckwheat (Fagopyrum esculentum Moench) cultivar. In contrast, P. bungeanum was much more Al sensitive since the root elongation was inhibited by 80% under the same condition. The difference in Al resistance among Polygonum species was confirmed in a 10-d intermittent Al treatment experiment, the root biomass of the first two species were unaltered and decreased by 50% in the latter species. However, high Al accumulation was not found in the leaves, indicating these species were not Al accumulators. Oxalate efflux was detected in root exudates of both Al resistant species, efflux initiated within 30 min treatment of 50 μM Al. No organic acid anions were detected in the root exudates of the Al sensitive species. The anion channel inhibitor phenylglyoxal (PG) inhibited the oxalate efflux greatly. Inhibition of root elongation was greater in the presence of PG, confirming that oxalate efflux was associated with the Al resistance. However, since the efflux rate was much lower than their related species buckwheat, other mechanisms must be involved in Al resistance and these need to be studied further.     

Continuous secretion of organic acids is related to aluminium resistance during relatively long‐term exposure to aluminium stress

Secretion of organic acid has been suggested to be one of the mechanisms for Al resistance in short‐term experiments. In the present study, relatively long‐term response of roots to Al stress was investigated in terms of organic acid secretion. Eight plant cultivars belonging to 5 species that exhibited differential sensitivity to Al were used. Ten days of intermittent exposure to Al (one day in 0.5 m M CaCl₂ containing 50 µ M AlCl₃ at pH 4.5, alternating with one day in nutrient solution without Al) inhibited root growth by 65% in an Al‐sensitive cultivar of wheat ( Triticum aestivum L. Scout 66) and by 25‐50% in two cultivars of oilseed rape ( Brassica napus L. 94008 and H166), two cultivars of oat ( Avena sativa L. Tochiyutaka and Heoats), and an Al‐tolerant cultivar of wheat (Atlas 66). However, root growth was hardly affected by the same treatment in buckwheat ( Fagopyrum esculentum Moench Jianxi) and radish ( Raphanus sativus L. Guangxi). Organic acids were monitored during the first 6 h of each day of Al treatment, and both the kind and amount of organic acids secreted were found to differ among different species. Roots of buckwheat secreted oxalic acid, those of wheat exuded malic acid, while those of rapeseed, oats, and radish secreted both citric and malic acids. Three different patterns in response to relatively long‐term treatment of Al were found in terms of total amount of organic acids secreted: (1) the amount secreted was very low during the treatment (wheat cv. Scout 66, oat), (2) the amount gradually decreased with duration of treatment (wheat cv. Atlas 66, oilseed rape), and (3) the amount maintained at a high level during the whole period of Al treatment (buckwheat and radish). Combined with the results of growth inhibition, it is suggested that the continuous secretion of organic acids at a high level is related to high Al resistance.     

Aluminum Tolerance in Wheat (Triticum aestivum L.) (II. Aluminum-Stimulated Excretion of Malic Acid from Root Apices)

We investigated the role of organic acids in conferring Al tolerance in near-isogenic wheat (Triticum aestivum L.) lines differing in Al tolerance at the Al tolerance locus (Alt1). Addition of Al to nutrient solutions stimulated excretion of malic and succinic acids from roots of wheat seedlings, and Al-tolerant genotypes excreted 5- to 10-fold more malic acid than Al-sensitive genotypes. Malic acid excretion was detectable after 15 min of exposure to 200 [mu]M Al, and the amount excreted increased linearly over 24 h. The amount of malic acid excreted was dependent on the external Al concentration, and excretion was stimulated by as little as 10 [mu]M Al. Malic acid added to nutrient solutions was able to protect Al-sensitive seedlings from normally phytotoxic Al concentrations. Root apices (terminal 3-5 mm of root) were the primary source of the malic acid excreted. Root apices of Al-tolerant and Al-sensitive seedlings contained similar amounts of malic acid before and after a 2-h exposure to 200 [mu]M Al. During this treatment, Al-tolerant seedlings excreted about four times the total amount of malic acid initially present within root apices, indicating that continual synthesis of malic acid was occurring. Malic acid excretion was specifically stimulated by Al, and neither La, Fe, nor the absence of Pi was able to elicit this response. There was a consistent correlation of Al tolerance with high rates of malic acid excretion stimulated by Al in a population of seedlings segregating for Al tolerance. These data are consistent with the hypothesis that the Alt1 locus in wheat encodes an Al tolerance mechanism based on Al-stimulated excretion of malic acid.     

Malate-permeable channels and cation channels activated by aluminum in the apical cells of wheat roots

Aluminum (Al(3+))-dependent efflux of malate from root apices is a mechanism for Al(3+) tolerance in wheat (Triticum aestivum). The malate anions protect the sensitive root tips by chelating the toxic Al(3+) cations in the rhizosphere to form non-toxic complexes. Activation of malate-permeable channels in the plasma membrane could be critical in regulating this malate efflux. We examined this by investigating Al(3+)-activated channels in protoplasts from root apices of near-isogenic wheat differing in Al(3+) tolerance at a single locus. Using whole-cell patch clamp we found that Al(3+) stimulated an electrical current carried by anion efflux across the plasma membrane in the Al(3+)-tolerant (ET8) and Al(3+)-sensitive (ES8) genotypes. This current occurred more frequently, had a greater current density, and remained active for longer in ET8 protoplasts than for ES8 protoplasts. The Al(3+)-activated current exhibited higher permeability to malate(2-) than to Cl(-) (P(mal)/P(Cl) > or = 2.6) and was inhibited by anion channel antagonists, niflumate and diphenylamine-2-carboxylic acid. In ET8, but not ES8, protoplasts an outward-rectifying K(+) current was activated in the presence of Al(3+) when cAMP was included in the pipette solution. These findings provide evidence that the difference in Al(3+)-induced malate efflux between Al(3+)-tolerant and Al(3+)-sensitive genotypes lies in the differing capacity for Al(3+) to activate malate permeable channels and cation channels for sustained malate release.     

Aluminium resistance requires resistance to acid stress: a case study with spinach that exudes oxalate rapidly when exposed to Al stress

Spinach is a vegetable with a high oxalate concentration in its tissues. Oxalate efflux from spinach (Spinacia oleracea L. cv. Quanneng) roots was rapidly stimulated (within 30 min) by aluminium (Al) treatment. The efflux was constant within 6 h, but increased with increasing Al concentration. The efflux was confined to the root tip (0-5 mm), which showed a 5-fold greater efflux than the root zone distal to the tip (5-10 mm). Oxalate efflux could not be triggered by treatment with the trivalent cation lanthanum or by phosphorus deficiency, indicating that the efflux was specific to the Al treatment. All this evidence suggested that spinach possesses Al-resistance mechanisms. However, spinach was found to be as sensitive to Al toxicity as the Al-sensitive wheat line ES8, which had no Al-dependent organic acids efflux. The Al accumulated in the apical 5 mm of the roots of spinach which was also similar to that in the Al-sensitive wheat after 24 h treatment with 50 microM AlCl(3), indicating a non-exclusion mechanism. In addition, root elongation in spinach was significantly inhibited at pH 4.5, compared with that at pH 6.5. Based on this evidence, it is concluded that the sensitivity to acid stress in spinach could mask the potential role for oxalate to protect the plant roots from Al toxicity.     

三峡库区消落带土壤水分变化对落羽杉（Taxodium distichum）幼苗根部次生代谢物质含量及根生物量的影响

模拟三峡库区消落带土壤淹水变化特征设置了常规生长水分条件(CK组,土壤含水量为田间持水量的60%～63%)、轻度干旱水分胁迫(T1组,土壤含水量为田间持水量的47%～50%)、土壤水饱和(T2组,土壤表面一直处于潮湿状态)以及水淹(T3组,苗木根部淹水超过土壤表面1cm)4个不同处理组,研究落羽杉当年实生幼苗根部次生代谢物质含量与生物量的变化(均以干重计)。研究表明,不同水分处理对落羽杉幼苗主根、侧根和根部苹果酸、莽草酸含量以及生物量的影响程度有所差异,其中以T3组受到的影响最为明显。主根苹果酸、莽草酸含量在整个试验期的总体平均值,T3组显著低于CK组,分别达28.0%和16.4%;相反侧根苹果酸、莽草酸总体平均含量T3组则极其显著地高于CK组分别达105.7%和152.6%,根部平均值T3组显著地高于CK组分别达32.7%和26.2%。与之形成鲜明对比,主根、侧根和根部苹果酸、莽草酸含量在整个试验期的总体平均值,T1、T2与CK组相互之间分别均无显著差异。各处理组之间主根生物量没有显著差异;与CK和T1组相比,T3组侧根生物量分别降低38.3%和40.8%,根部生物量分别降低31.9%和31.1%,但与T2组相比均未达到显著差异。主根苹果酸含量与根系各部分莽草酸含量和生物量之间均无显著相关性,相反侧根、根部的苹果酸与莽草酸含量以及二者与生物量相互间均表现出显著或极显著相关性。在三峡库区消落带土壤含水量变化条件下,落羽杉幼苗将充分利用侧根增强代谢适应调节能力,通过产生大量苹果酸和莽草酸、减少根部生物量积累适应根部水淹环境,通过维持与CK组同样水平的代谢和生长而适应轻度干旱与饱和水环境。     

Mechanisms Underlying the Regulation of Root Formation in <Emphasis Type="Italic">Malus hupehensis</Emphasis> Stem Cuttings by Using Exogenous Hormones

Malus hupehensis is one of the most important Malus ornamental and rootstock species in the south China Yellow River Basin. In the present study, we treated the stem cuttings of M. hupehensis with one of three exogenous hormones, indole acetic acid (IAA), naphthalene acetic acid (NAA), or a compound plant growth regulator (GGR) to investigate the mechanisms underlying root formation in stem cuttings and to optimize stem cutting propagation techniques. The results showed that immersing the stem cuttings in 100 mg/L of IAA for 2 h before planting was most effective, which reduced the time to root formation by 21 days and increased rooting percentage by 129.4 %, compared to that in the control group. In addition, the levels of endogenous substances (endogenous hormones, soluble proteins, and carbohydrates) dynamically changed, with the time to peak value or time to valley value of each parameter synchronized well with the initiation of adventitious roots. The synchronized change suggested that root formation was coordinated with physiological metabolism. However, exogenous hormone treatment significantly accelerated the catabolism of the root inhibiting hormone, abscisic acid. On the other hand, exogenous hormone treatment significantly enhanced the accumulation of root promoting hormones [IAA, gibberellic acid (GA₃), and zeatin riboside (ZR)] and soluble proteins. Moreover, exogenous hormone treatments accelerated the consumption of starch and soluble sugars. Overall, the results indicated that exogenous hormone treatment (IAA) accelerated the synthesis of endogenous hormones (IAA, GA₃, and ZR), therefore, sped up the metabolism of carbohydrates and soluble proteins, and consequently quickened the root formation process.     

Citrate transporters play a critical role in aluminium-stimulated citrate efflux in rice bean (Vigna umbellata) roots

BACKGROUND AND AIMS: Aluminium (Al) stimulates the efflux of citrate from apices of rice bean (Vigna umbellata) roots. This response is delayed at least 3 h when roots are exposed to 50 microm Al, indicating that some inducible processes leading to citrate efflux are involved. The physiological bases responsible for the delayed response were examined here. METHODS: The effects of several antagonists of anion channels and citrate carriers, and of the protein synthesis inhibitor, cycloheximide (CHM) on Al-stimulated citrate efflux and/or citrate content were examined by high-pressure liquid chromatography (HPLC) or an enzymatic method. KEY RESULTS: Both anion channel inhibitors and citrate carrier inhibitors can inhibit Al-stimulated citrate efflux, with anthracene-9-carboxylic acid (A-9-C, an anion channel inhibitor) and phenylisothiocyanate (PI, a citrate carrier inhibitor) the most effective inhibitors. A 6 h pulse of 50 microm Al induced a significant increase of citrate content in root apices and release of citrate. However, the increase in citrate content preceded the efflux. Furthermore, the release of citrate stimulated by the pulse treatment was inhibited by both A-9-C and PI, indicating the importance of the citrate carrier on the mitochondrial membrane and the anion channel on the plasma membrane for the Al-stimulated citrate efflux. CHM (20 microm) also significantly inhibited Al-stimulated citrate efflux, confirming that de novo protein synthesis is required for Al-stimulated citrate efflux. CONCLUSIONS: These results indicate that the activation of genes possibly encoding citrate transporters plays a critical role in Al-stimulated citrate efflux.     

Levels of endogenous abscisic acid and indole-3-acetic acid influence shoot organogenesis in callus cultures of rice subjected to osmotic stress

Osmotic stress and endogenous hormone levels may have a role in shoot organogenesis, but a systematic study has not yet to investigate the links. We evaluated the changes of the endogenous indole-3-acetic acid (IAA) and abscisic acid (ABA) levels in rice (Oryza sativa L. cv. Tainan 5) callus during shoot organogenesis induced by exogenous plant growth regulator treatments or under osmotic stress. Non-regenerable callus showed low levels of endogenous ABA and IAA, with no fluctuation in level during the period evaluated. The addition of 100 μM ABA or 2 mM anthranilic acid (IAA precursor) into Murashige and Skoog basal induction medium containing 10 μM 2,4-D enhanced the regeneration frequency slightly, to 5 and 35%, respectively, and their total cellular ABA or IAA levels were increased significantly, correspondingly to the treatments. However, the regeneration frequency was greatly increased to 80% after treatment with 0.6 M sorbitol or 100 μM ABA and 2 mM anthranilic acid combined. Both treatments produced high levels of total cellular ABA and IAA at the callus stage, which was quickly decreased on the first day after transfer to regeneration medium. Thus, osmotic stress-induced simultaneous accumulation of endogenous ABA and IAA is involved in shoot regeneration in rice callus.     

Citrate transporters play an important role in regulating aluminum-induced citrate secretion in Glycine max

To further understand the process of Al-induced citrate secretion from soybean roots, the effect of protein synthesis inhibitor, anion channel blockers, and citrate carrier inhibitors on Al-induced citrate exudation was investigated in Al-resistant soybean cultivar PI 416937. Citrate exudation from roots increased with the increase of Al concentration from 10 to 50 μM and initiated after 4 h of Al exposure. Protein synthesis inhibitor, cycloheximide (CHM; 25 μM) completely inhibited Al-induced citrate secretion during 12-h exposure, suggesting that novel protein synthesis was necessary in Al-induced citrate efflux. Also both anion channel blocker anthracene-9-carboxylic acid (A-9-C) and citrate carrier inhibitor mersalyl acid (Mersalyl) significantly reduced citrate secretion, suggesting that both anion channels in plasma membrane and citrate carriers in mitochondria membrane were the rate limiting factors of Al dependent citrate release. However, Al-induced citrate secretion was insensitive to anion channel blockers phenylglyoxal (PG), 4,4′-diisothiocyanostibene-2,2′-disulfonat (DIDS) and citrate carrier inhibitor pyridoxal 5′-P (PP).     

Differential effects of aluminium on osmotic potential and sugar accumulation in the root cells of Al-resistant and Al-sensitive wheat

The changes in osmotic potential and the concentration of osmotic solutes in the cell sap of the root tips exposed to Al were examined in two cultivars of wheat ( Triticum aestivum ) differing in Al resistance. Root elongation was less influenced by an 8-h exposure to 20 μ M or 50 μ M Al in Al-resistant cv. Atlas 66 than in Al-sensitive cv. Scout 66. After Al treatment the osmotic potential of the root cells was decreased in Atlas 66 but increased in Scout 66 indicating that the Al treatment osmotically stimulated the driving force for water uptake in Atlas 66 but suppressed it in Scout 66. Al increased the concentration of soluble sugars, the major osmotic solute in the root cells in Atlas 66, but decreased it in Scout 66. Al at both low (5 μ M ) and high (50 μ M ) concentrations, also increased the concentration of soluble sugars in the Al-resistant genotype ET8 but a high Al concentration decreased it in Al-sensitive genotype ES8. Enzymatic analyses and thin-layer chromatography revealed that soluble sugars in the root cells of both Atlas 66 and Scout 66 mainly consisted of monosaccharides such as glucose, fructose and a small amount of sucrose. These results suggest that the accumulation of soluble sugars in Al-resistant wheat Atlas 66 keeps the osmotic potential in the root cells low and thus, enables the root cells to take up water and to elongate against the pressure produced by cell wall rigidification under Al stress.     

Interaction of boron and aluminum on the physiological characteristics of rape (<Emphasis Type="Italic">Brassica napus</Emphasis> L.) seedlings

It has been reported that aluminum (Al) toxicity is a major limiting factor for plant growth and production on acidic soils. Boron (B) is indispensable micronutrient for normal growth of higher plants, and its addition could alleviate Al toxicity. The rape seedlings were grown under three B (0.25, 25 and 500 μM) and two Al concentrations [0 (?Al) and 100 μM (+Al) as AlCl₃·6H₂O]. The results indicated that Al stress severely hampered root elongation and root activity at 0.25 μM B while the normal (25 μM) and excess (500 μM) B improved the biomass of rape seedlings under Al exposure. Additionally, normal and excess B treatment reduced accumulation of Al in the roots and leaves under Al toxicity, which was also confirmed by hematoxylin with light staining. This indicates that both normal and excess B could alleviate Al toxicity. Furthermore, it also decreased the contents of malondialdehyde and soluble protein under Al toxicity. Likewise, superoxide dismutase activity (SOD) improved by 97.82 and 131.96% in the roots, and 168 and 119.88% in the leaves at 25 and 500 µM B, respectively, while the peroxidase and catalase activities dropped as a result of Al stress. The study results demonstrated that appropriate B application is necessary to avoid the harmful consequences of Al toxicity in rape seedlings.     

Effects of indole-3-acetic acid on arsenic uptake and antioxidative enzymes in Pteris cretica var. nervosa and Pteris ensiformis

A hydroponic experiment was conducted to investigate the effects of indole-3-acetic acid (IAA) on arsenic (As) uptake and antioxidative enzymes in fronds of Pteris cretica var. nervosa (As hyperaccumulator) and Pteris ensiformis (non-hyperaccumulator). Plants were exposed to 2 mg L^?1 As(III), As(V) or dimethylarsinic acid (DMA) and IAA concentrations for 14 d. The biomass and total As in the plants significantly increased at 30 mg L^?1 IAA. Superoxide dismutase (SOD) activities significantly increased with IAA addition. Catalase (CAT) activities showed a significant increase in P. ensiformis exposed to three As species at 30 or 50 mg L^?1 IAA but varied in P. cretica var. nervosa. Peroxidase (POD) activities were unchanged in P. ensiformis except for a significant decrease at 50 mg L^?1 IAA under As(III) treatment. However, a significant increase was observed in P. cretica var. nervosa at 10 mg L^?1 IAA under As(III) or DMA treatment and at 50 mg L^?1 IAA under As(V) treatment. Under DMA stress, malondialdehyde contents in fronds of P. cretica var. nervosa showed a significant decrease at 10 mg L^?1 IAA but remained unchanged in P. ensiformis. Therefore, IAA enhanced As uptake and frond POD activity in P. cretica var. nervosa under As stress.     

Functional characterization of plasma membrane-localized organic acid transporter (CsALMT1) involved in aluminum tolerance in <Emphasis Type="Italic">Camelina sativa</Emphasis> L.

Aluminum (Al) toxicity is a major limiting factor that inhibits root elongation and decreases crop production in acidic soils. The symptoms of inhibited root growth include a reduced uptake of nutrients because the roots become stubby and brittle. The release of organic anions from roots can protect a plant from Al toxicity. The mechanism relies on the efflux of organic anions, such as malate or citrate, which protect roots by chelating the Al³⁺. In this study, homologs of TaALMT1, a Camelina gene that encodes an aluminum-activated malate transporter, were investigated. The expression of this gene was induced by Al in the root, but not in the shoots. Using green fluorescent protein (GFP) fusion constructs and Western-blot analysis, we observed that CsALMT1 was localized in the plasma membrane. Also, to determine the degree to which Al tolerance was affected by malate secretion in Camelina root, we generated CsALMT1 overexpressing plants. CsALMT1 overexpressing transgenic plants showed a higher root elongation rate than the wild-type plant. Damaged cell staining analysis by hematoxylin under 25 µM Al treatment for 2, 4, and 6 h showed a pattern of less damage in CsALMT1 transgenic plants than in wild-type plant, especially in the root elongation zone. Furthermore, the rate of increase of secretion of organic acid in overexpressed plants after Al treatment was higher than that in the wild-type plant. In addition, in the Al-specific dye morin staining on root protoplast under 50 µM Al treatment, less Al accumulation was observed in the CsALMT1 transgenic plants than in the wild-type plant. The Al contents in the roots of the transgenic plants were at a lower level than those in the wild-type plant. These results show that the overexpression of CsALMT1 improves Al tolerance by increasing the release of malate from the root to the soil and, thereby, detoxifies the Al³⁺.