Use of comprehensive target analysis for determination of contaminants of emerging concern in a sediment core collected from Beppu Bay, Japan.

In recent years, concern about the release of anthropogenic organic micropollutants referred to as contaminants of emerging concern (CECs) has been growing. The objective of this study was to find potential CECs by means of an analytical screening method referred to as comprehensive target analysis with an automated identification and quantification system (CTA-AIQS), which uses gas and liquid chromatography combined with mass spectrometry (GC-MS and LC-QTOF-MS). We used CTA-AIQS to analyze samples from a sediment core collected in Beppu Bay, Japan. With this method, we detected 80 compounds in the samples and CTA-AIQA could work to useful tool to find CECs in environmental media. Among the detected chemicals, three PAHs (anthracene, chrysene, and fluoranthene) and tris(isopropylphenyl)phosphate (TIPPP) isomers were found to increase in concentration with decreasing sediment depth. We quantified TIPPP isomers in the samples by means of targeted analysis using LC-MS/MS for confirmation. The concentration profiles, combined with previous reports indicating persistent, bioaccumulative, and toxic properties, suggest that these chemicals can be categorized as potential CECs in marine environments.

Development and evaluation of a novel in-clinic automated hematology analyzer, ProCyte Dx, for canine erythrocyte indices, leukogram, platelet counts and reticulocyte counts.

A novel hematology analyzer for small animal medicine, ProCyte Dx, was developed from combination of the fluorescence laser flow cytometry and laminar flow impedance technologies, and its accuracy was evaluated by comparing with the conventional impedance-based hematology analyzer, pocH-100iV Diff, or microscopic manual cell counting methods with staining blood smears in the canine blood. Blood samples of 59 dogs were hematologically analyzed and compared by Pearson's correlation coefficients. Analyses between the two analyzers showed excellent correlation in RBC (r=0.998), HGB (r=0.999), HCT (r=0.998), MCV (r=0.994), MCH (r=0.974), MCHC (r=0.906), WBC (r=0.998) and PLT (r=0.993). Analyses between ProCyte Dx and microscopic manual counting results showed excellent correlation in neutrophils (r=0.920), lymphocytes (r=0.913) and reticulocyte percentages (r=0.924), good correlation in eosinophils (r=0.815) and reticulocyte numbers (r=0.850) and fair correlation in monocytes (r=0.770). The present study indicates that ProCyte Dx is acceptably accurate and can be a powerful tool for canine clinical medicine.

Overexpression of alternative oxidase gene confers aluminum tolerance by altering the respiratory capacity and the response to oxidative stress in tobacco cells.

Aluminum (Al) stress represses mitochondrial respiration and produces reactive oxygen species (ROS) in plants. Mitochondrial alternative oxidase (AOX) uncouples respiration from mitochondrial ATP production and may improve plant performance under Al stress by preventing excess accumulation of ROS. We tested respiratory changes and ROS production in isolated mitochondria and whole cell of tobacco (SL, ALT 301) under Al stress. Higher capacities of AOX pathways relative to cytochrome pathways were observed in both isolated mitochondria and whole cells of ALT301 under Al stress. AOX1 when studied showed higher AOX1 expression in ALT 301 than SL cells under stress. In order to study the function of tobacco AOX gene under Al stress, we produced transformed tobacco cell lines by introducing NtAOX1 expressed under the control of the cauliflower mosaic virus (CaMV) 35 S promoter in sensitive (SL) Nicotiana tabacum L. cell lines. The enhancement of endogenous AOX1 expression and AOX protein with or without Al stress was in the order of transformed tobacco cell lines > ALT301 > wild type (SL). A decreased respiratory inhibition and reduced ROS production with a better growth capability were the significant features that characterized AOX1 transformed cell lines under Al stress. These results demonstrated that AOX plays a critical role in Al stress tolerance with an enhanced respiratory capacity, reducing mitochondrial oxidative stress burden and improving the growth capability in tobacco cells.

Aluminum toxicity recovery processes in root apices. Possible association with oxidative stress.

Al inhibits root apex elongation with concomitant morphological injuries such as ruptures punctuated by the regions stained with Evans blue. The recovery can be investigated by transfer of Al-injured roots to a solution lacking Al. In the Al-injured root apex, superoxide anion, H(2)O(2), Al, and lignin accumulate. During the recovery process, the central cylinder elongates leaving the region stained with Evans blue without marked disappearance. The obvious function of the region is not clear but may trigger the elongation of central cylinder during the recovery process. Thus the function of the region stained with Evans blue might be derived from the programmed cell-like idea. Oxidative stress concerns events induced under Al toxicity and the recovery process. The superoxide anion is primarily formed by plasma membrane-associated NADPH oxidase and is dismuted to H(2)O(2) and O(2) by superoxide dismutase. H(2)O(2) provides the electrons for the polymerization of phenolics to lignin, which causes the stiffening of the cell wall. The distortion of the cell wall caused by lignin may induce the breaking and tearing of cells, which results in the formation of ruptures at the rhizodermis and outer cortex layers. The production of superoxide anion, H(2)O(2), and lignin was reduced during the recovery process and thereby the elongation of the central cylinder may be induced.

DNA display selection of peptide ligands for a full-length human G protein-coupled receptor on CHO-K1 cells.

The G protein-coupled receptors (GPCRs), which form the largest group of transmembrane proteins involved in signal transduction, are major targets of currently available drugs. Thus, the search for cognate and surrogate peptide ligands for GPCRs is of both basic and therapeutic interest. Here we describe the application of an in vitro DNA display technology to screening libraries of peptide ligands for full-length GPCRs expressed on whole cells. We used human angiotensin II (Ang II) type-1 receptor (hAT1R) as a model GPCR. Under improved selection conditions using hAT1R-expressing Chinese hamster ovary (CHO)-K1 cells as bait, we confirmed that Ang II gene could be enriched more than 10,000-fold after four rounds of selection. Further, we successfully selected diverse Ang II-like peptides from randomized peptide libraries. The results provide more precise information on the sequence-function relationships of hAT1R ligands than can be obtained by conventional alanine-scanning mutagenesis. Completely in vitro DNA display can overcome the limitations of current display technologies and is expected to prove widely useful for screening diverse libraries of mutant peptide and protein ligands for receptors that can be expressed functionally on the surface of CHO-K1 cells.

Changes in rupture formation and zonary region stained with Evans blue during the recovery process from aluminum toxicity in the pea root apex.

We investigated how the pea (Pisum sativum cv. Harunoka) root, upon return to an Al-free condition, recovers from injury caused by exposure to Al. Elongation and re-elongation of the root during the recovery process from Al injury occurred only in the apical 5-mm region of the pea root. With the model system of the pea root for recovery from Al injury, images of the root characterized by zonal staining with Evans blue showed the existence of two regions in the root apex consisting of rupture and zonary stained regions. Ruptures enlarged by increase in their depth but without widening of the intervals among zonary stained regions in the roots treated with Al continuously. On the other hand, intervals of the zonary stained regions were widened due to re-elongation of the root and were narrow in the rupture region in the recovery root.

Aluminium reduces sugar uptake in tobacco cell cultures: a potential cause of inhibited elongation but not of toxicity.

Aluminium is well known to inhibit plant elongation, but the role in this inhibition played by water relations remains unclear. To investigate this, tobacco (Nicotiana tabacum L.) suspension-cultured cells (line SL) was used, treating them with aluminium (50 microM) in a medium containing calcium, sucrose, and MES (pH 5.0). Over an 18 h treatment period, aluminium inhibited the increase in fresh weight almost completely and decreased cellular osmolality and internal soluble sugar content substantially; however, aluminium did not affect the concentrations of major inorganic ions. In aluminium-treated cultures, fresh weight, soluble sugar content, and osmolality decreased over the first 6 h and remained constant thereafter, contrasting with their continued increases in the untreated cultures. The rate of sucrose uptake, measured by radio-tracer, was reduced by approximately 60% within 3 h of treatment. Aluminium also inhibited glucose uptake. In an aluminium-tolerant cell line (ALT301) isogenic to SL, all of the above-mentioned changes in water relations occurred and tolerance emerged only after 6 h and appeared to involve the suppression of reactive oxygen species. Further separating the effects of aluminium on elongation and cell survival, sucrose starvation for 18 h inhibited elongation and caused similar changes in cellular osmolality but stimulated the production of neither reactive oxygen species nor callose and did not cause cell death. We propose that the inhibition of sucrose uptake is a mechanism whereby aluminium inhibits elongation, but does not account for the induction of cell death.

Aluminum stress signaling in plants.

Aluminum (Al) toxicity is a major constraint for crop production in acidic soil worldwide. When the soil pH is lower than 5, Al(3+) is released to the soil and enters into root tip cell ceases root development of plant. In acid soil with high mineral content, Al is the major cause of phytotoxicity. The target of Al toxicity is the root tip, in which Al exposure causes inhibition of cell elongation and cell division, leading to root stunting accompanied by reduced water and nutrient uptake. A variety of genes have been identified that are induced or repressed upon Al exposure. At tissue level, the distal part of the transition zone is the most sensitive to Al. At cellular and molecular level, many cell components are implicated in the Al toxicity including DNA in nucleus, numerous cytoplastic compounds, mitochondria, the plasma membrane and the cell wall. Although it is difficult to distinguish the primary targets from the secondary effects so far, understanding of the target sites of the Al toxicity is helpful for elucidating the mechanisms by which Al exerts its deleterious effects on root growth. To develop high tolerance against Al stress is the major goal of plant sciences. This review examines our current understanding of the Al signaling with the physiological, genetic and molecular approaches to improve the crop performance under the Al toxicity. New discoveries will open up new avenues of molecular/physiological inquiry that should greatly advance our understanding of Al tolerance mechanisms. Additionally, these breakthroughs will provide new molecular resources for improving the crop Al tolerance via molecular-assisted breeding and biotechnology.

Mitochondrial alterations related to programmed cell death in tobacco cells under aluminium stress.

The present investigation was undertaken to verify whether mitochondria play a significant role in aluminium (Al) toxicity, using the mitochondria isolated from tobacco cells (Nicotiana tabacum, non-chlorophyllic cell line SL) under Al stress. An inhibition of respiration was observed in terms of state-III, state-IV, succinate-dependent, alternative oxidase (AOX)-pathway capacity and cytochrome (CYT)-pathway capacity, respectively, in the mitochondria isolated from tobacco cells subjected to Al stress for 18 h. In accordance with the respiratory inhibition, the mitochondrial ATP content showed a significant decrease under Al treatment. An enhancement of reactive oxygen species (ROS) production under state-III respiration was observed in the mitochondria isolated from Al-treated cells, which would create an oxidative stress situation. The opening of mitochondrial permeability transition pore (MPTP) was seen more extensively in mitochondria isolated from Al-treated cells than in those isolated from control cells. This was Ca(2+) dependent and well modulated by dithioerythritol (DTE) and Pi, but insensitive to cyclosporine A (CsA). The collapse of inner mitochondrial membrane potential (DeltaPsi(m)) was also observed with a release of cytochrome c from mitochondria. A great decrease in the ATP content was also seen under Al stress. Transmission electron microscopy analysis of Al-treated cells also corroborated our biochemical data with distortion in membrane architecture in mitochondria. TUNEL-positive nuclei in Al-treated cells strongly indicated the occurrence of nuclear fragmentation. From the above study, it was concluded that Al toxicity affects severely the mitochondrial respiratory functions and alters the redox status studied in vitro and also the internal structure, which seems to cause finally cell death in tobacco cells.

Aluminum toxicity and Ca depletion may enhance cell death of tobacco cells via similar syndrome.

The main objective of this work is to find out whether aluminum (Al) toxicity and Ca depletion cause cell death of tobacco cells via similar sequence of events. Tobacco cell suspension culture exhibited maximum fresh weight in the presence of a wide range of Ca concentrations between 0.1-1.0 mM whereas higher concentrations (>1.0-5.0 mM) gradually lowered cell fresh weight. However, this decrease in fresh weight does not imply a negative impact on cell viability since cell growth recommenced in fresh MS medium with rates mostly higher than those of low Ca. In addition, high Ca seems to be crucial for survival of Al-treated cells. On the other side, tobacco cells exhibited extreme sensitivity to complete deprivation of Ca. Without Ca, cells could not survive for 18 h and substantially lost their growth capability. Evans blue uptake proved membrane damage of Ca-depleted same as Al-treated cells; relative to maintained membrane intactness of calcium-supplemented (control) ones. Percentage of membrane damage and the growth capability (survival) of tobacco cells exhibited a clear negative correlation.Alterations in growth (fresh weight per aliquot) could not be ascribed neither to cell number nor to decreased dry matter allocation (dry weight/fresh weight percentage) but was mainly due to decreased cellular water content. In this context, Ca-depleted cells lost about half their original water content while 100 microM Al-treated ones retained most of it (ca 87%). This represented the single difference between the two treatments (discussed in the text). Nevertheless, such high water content of the Al-treated cells seems physiologically useless since it did not result in improved viability. Similarities, however, included negligible levels of growth capability, maximum levels of membrane damage, and comparable amounts of NO(3) (-) efflux. As well, both types of treatments led to a sharp decline in osmotic potential that is, in turn, needed for water influx. The above-mentioned sequence of events, induced by Al application looks, to a great extent, similar to Ca depletion syndrome leading finally to cell death of tobacco cells.

Overexpression of an auxilin-like gene (F9E10.5) can suppress Al uptake in roots of Arabidopsis.

Plants resistant to aluminium (Al) stress were isolated from Arabidopsis thaliana enhancer-tagged mutant lines. Compared with the parental Col-7 control line, one of the resistant candidates, #355-2, showed a higher expression of the F9E10.5 gene (At1g75100) on chromosome 1, a lower Al content in whole roots, and a shorter root hair length (approximately 30%). Both Al influx and associated oxidative stress occurred in root hairs, as well as in root tips of Col-7; however, they were seen only in root tips of #355-2. Transgenic plants overexpressing the F9E10.5 gene showed a slightly higher Al resistance than their parental control line (Ler). The F9E10.5 gene encodes an auxilin-like protein related to the clathrin-uncoating process in endocytosis. Microscopic observation indicated that both Al ion influx and endocytosis activity were lower in root hair cells of the #355-2 line than in those of Col-7. These results suggested that overexpression of this auxilin-like protein inhibits endocytosis in root hair cells by a disturbance of the transport system as in animal cells shown previously. It was also suggested that a part of the Al influx occurred via endocytosis in root hair cells in Arabidopsis. The Al resistance in the #355-2 line may therefore be due to a lower Al uptake via endocytosis in the root hair region.

The Membrane Topology of ALMT1, an Aluminum-Activated Malate Transport Protein in Wheat (Triticum aestivum).

The wheat ALMT1 gene encodes an aluminum (Al)-activated malate transport protein which confers Al-resistance. We investigated the membrane topology of this plasma-membrane localized protein with immunocytochemical techniques. Several green fluorescent protein (GFP)-fused and histidine (His)-tagged chimeras of ALMT1 were prepared based on a computer-predicted secondary structure and transiently expressed in cultured mammalian cells. Antibodies raised to polypeptide epitopes of ALMT1 were used in conjunction with the antibody to the His-tags to determine the topology of ALMT1. This study shows that the ALMT1 protein contains six transmembrane domains with the amino and carboxyl termini located on the extracellular side of the plasma membrane.

The BnALMT1 Protein That is an Aluminum-Activated Malate Transporter is Localized in the Plasma Membrane.

We have previously reported that Al-induces citrate and malate efflux from P-sufficient and P-deficient plants of rape (Brassica napus L.) and that P-deficiency alone could not induce this response. Further investigation showed that the transcript of two genes designated BnALMT1 and BnALMT2 is accumulated in roots by Al-treatment. Transgenic tobacco cells (Nicotiana tabacum) and Xenopus laevis oocytes expressing the BnALMT1 and BnALMT2 proteins released more malate than control cells in the presence of Al, indicating that the BnALMT genes encode an Al-activated malate transporter. The transgenic tobacco cells exposed to toxic level of Al grew better than control cells indicating that the genes can enhance Al-resistance of plant cells. In this study we showed the subcellular localization of BnALMT1 fused to the green fluoresce protein (GFP). The BnALMT1:: GFP construct was transiently expressed in protoplasts prepared from Arabidopsis leaves using the polyethylene glycol (PEG) method. The result showed that the BnALMT1 protein is localized in the plasma membrane. This provides further evidence that the BnALMT proteins facilitate the transport of malate across the plasma membrane (PM).

The BnALMT1 and BnALMT2 genes from rape encode aluminum-activated malate transporters that enhance the aluminum resistance of plant cells.

The release of organic anions from roots can protect plants from aluminum (Al) toxicity and help them overcome phosphorus (P) deficiency. Our previous findings showed that Al treatment induced malate and citrate efflux from rape (Brassica napus) roots, and that P deficiency did not induce the efflux. Since this response is similar to the malate efflux from wheat (Triticum aestivum) that is controlled by the TaALMT1 gene, we investigated whether homologs of TaALMT1 are present in rape and whether they are involved in the release of organic anions. We isolated two TaALMT1 homologs from rape designated BnALMT1 and BnALMT2 (B. napus Al-activated malate transporter). The expression of these genes was induced in roots, but not shoots, by Al treatment but P deficiency had no effect. Several other cations (lanthanum, ytterbium, and erbium) also increased BnALMT1 and BnALMT2 expression in the roots. The function of the BnALMT1 and BnALMT2 proteins was investigated by heterologous expression in cultured tobacco (Nicotiana tabacum) cells and in Xenopus laevis oocytes. Both transfection systems showed an enhanced capacity for malate efflux but not citrate efflux, when exposed to Al. Smaller malate fluxes were also activated by ytterbium and erbium treatment. Transgenic tobacco cells grew significantly better than control cells following an 18 h treatment with Al, indicating that the expression of BnALMT1 and BnALMT2 increased the resistance of these plant cells to Al stress. This report demonstrates that homologs of the TaALMT1 gene from wheat perform similar functions in other species.

Sequence upstream of the wheat (Triticum aestivum L.) ALMT1 gene and its relationship to aluminum resistance.

Aluminum (Al) resistance in wheat relies on the Al-activated malate efflux from root apices, which appears to be controlled by an Al-activated anion transporter encoded by the ALMT1 gene on chromosome 4DL. Genomic regions upstream and downstream of ALMT1 in 69 wheat lines were characterized to identify patterns that might influence ALMT1 expression. The first 1,000 bp downstream of ALMT1 was conserved among the lines examined apart from the presence of a transposon-like sequence which did not correlate with Al resistance. In contrast, the first 1,000 bp upstream of the ALMT1 coding region was more variable and six different patterns could be discerned (types I-VI). Type I had the simplest structure, while the others had blocks of sequence that were duplicated or triplicated in different arrangements. A pattern emerged among the lines of non-Japanese origin such that the number of repeats in this upstream region was positively correlated with the levels of ALMT1 expression and Al resistance. In contrast, many of the Japanese lines exhibited a large variation in ALMT1 expression and Al resistance despite possessing the same type of upstream region. Although ALMT1 expression was also poorly correlated with Al-activated malate efflux in the Japanese lines, a strong correlation between malate efflux and Al resistance suggested that malate efflux was still the primary mechanism for Al resistance, and that additional genes are involved in the post-transcriptional regulation of ALMT1 function.

AtALMT1, which encodes a malate transporter, is identified as one of several genes critical for aluminum tolerance in Arabidopsis.

Aluminum (Al) tolerance in Arabidopsis is a genetically complex trait, yet it is mediated by a single physiological mechanism based on Al-activated root malate efflux. We investigated a possible molecular determinant for Al tolerance involving a homolog of the wheat Al-activated malate transporter, ALMT1. This gene, named AtALMT1 (At1g08430), was the best candidate from the 14-member AtALMT family to be involved with Al tolerance based on expression patterns and genomic location. Physiological analysis of a transferred DNA knockout mutant for AtALMT1 as well as electrophysiological examination of the protein expressed in Xenopus oocytes showed that AtALMT1 is critical for Arabidopsis Al tolerance and encodes the Al-activated root malate efflux transporter associated with tolerance. However, gene expression and sequence analysis of AtALMT1 alleles from tolerant Columbia (Col), sensitive Landsberg erecta (Ler), and other ecotypes that varied in Al tolerance suggested that variation observed at AtALMT1 is not correlated with the differences observed in Al tolerance among these ecotypes. Genetic complementation experiments indicated that the Ler allele of AtALMT1 is equally effective as the Col allele in conferring Al tolerance and Al-activated malate release. Finally, fine-scale mapping of a quantitative trait locus (QTL) for Al tolerance on chromosome 1 indicated that AtALMT1 is located proximal to this QTL. These results indicate that AtALMT1 is an essential factor for Al tolerance in Arabidopsis but does not represent the major Al tolerance QTL also found on chromosome 1.

Root plasma membrane H+-ATPase is involved in the adaptation of soybean to phosphorus starvation.

The plasma membrane H+-ATPase plays an important role in the plant response to nutrient and environmental stresses. However, the involvement of plant root plasma membrane H+-ATPase in adaptation to phosphate (P) starvation is not yet fully elucidated. In this study, experiments were performed with soybean roots in low-P nutrient solution (10 microM). Treatment with fusicoccin, an activator of the plasma membrane H+-ATPase, increased P uptake by 35%, while vanadate, an inhibitor of plasma membrane H+-ATPase, severely suppressed it. These results suggested that P uptake might be regulated via the modulation of the activity of plasma membrane H+-ATPase under P starvation. The relationship between P uptake and the activity of plasma membrane H+-ATPase was examined further by using plasma membrane H+-ATPase transgenic Arabidopsis thaliana under low-P conditions. Transgenic plants absorbed more P compared with wild-type Arabidopsis. Results from real-time RT-PCR, western-blotting and immunolocalization analysis indicated that the increase in activity of the plasma membrane H+-ATPase by P starvation was caused by its transcriptional and translational regulation. A higher expression was observed at the translational level than at the transcriptional level. P starvation could induce a transient increase of endogenous indole-3-acetic acid (IAA) in soybean roots. The exogenous application of IAA stimulated the activity of plasma membrane H+-ATPase and P uptake, while naphthylphthalamic acid (NPA), an IAA transport inhibitor, blocked IAA effects. Taken together, these results suggested an involvement of root plasma membrane H+-ATPase in the adaptation of soybean to P starvation. IAA might be involved in signal transduction of P starvation by activating the plasma membrane H+-ATPase in soybean roots.

Cytotoxic thio-malate is transported by both an aluminum-responsive malate efflux pathway in wheat and the MAE1 malate permease in Schizosaccharomyces pombe.

Aluminum (Al) tolerance in wheat (Triticum aestivum L.) is mainly achieved by malate efflux, which is regulated by the expression of the recently identified gene, presumably encoding an Al-activated malate efflux transporter (ALMT1). However, the transport mechanism is not fully understood, partly as a result of the rapid turnover of its substrate. We developed a tool to study malate transport in wheat by screening biological compounds using the well-characterized Schizosaccharomyces pombe malate transporter (SpMAE1). Expression of SpMAE1 in both S. pombe and Saccharomyces cerevisiae, which has no SpMAE1 homologue, caused hypersensitivity to thio-malic acid. This hypersensitivity was prominent at pH 3.5, but not pH 4.5, and was accompanied by an increase in thiol content, indicating that SpMAE1 mediates the uptake of thio-malic acid at a specific low pH. In wheat, root apices were able to accumulate thio-malic acid without growth reduction at pH values above 4.2. Pretreatment of root apices with thio-malic acid followed by Al treatment induced thio-malate efflux. Al-induced thio-malate efflux was much higher in Al-resistant cultivars/genotypes than in Al-sensitive ones, and was accompanied by a decrease in thiol-content. Thio-malate efflux in the Al-resistant cultivar was slightly activated by lanthanum or ytterbium ion. Thio-malic acid did not alleviate the Al-induced inhibition of root elongation in wheat. Taken together, our results suggest that thio-malate acts as an analogue for malate in malate transport systems in wheat and yeast, and that it may be a useful tool for the analysis of malate transport involved in Al-tolerance and of other organic ion transport processes.

The role of the plasma membrane in the response of plant roots to aluminum toxicity.

Al(3+), the predominant form of solubilized aluminum at pH values below 5.0, has been shown to exert a profound inhibitory effect on root elongation. Al is known to accumulate at the root apex. The plasma membrane represents the first potential target for Al toxicity, due to its pronounced binding to phospholipids. Al appears to alter both the structure and functions of the plasma membrane, and a great deal of research has been conducted concerning the interactions between Al and the plasma membrane. In this review, recent findings regarding the interactions between Al and the plasma membrane are described, specifically findings involving Al-induced alterations in the structure and function of the plasma membrane.

Functions of two genes in aluminium (Al) stress resistance: repression of oxidative damage by the AtBCB gene and promotion of efflux of Al ions by the NtGDI1gene.

The functions of two genes whose expression provides tolerance to aluminium (Al) stress were investigated using plants and Saccharomyces cerevisiae (yeast): the Arabidopsis thaliana blue copper binding gene (AtBCB) and Nicotiana tabacum guanosine diphosphate (GDP) dissociation inhibitor gene (NtGDI1). To determine the localization of these proteins, each gene was fused to the green fluorescent protein (GFP) gene and introduced into onion epidermal cells. AtBCB was localized to cell membrane region and NtGDI1 to cytoplasm. Transgenic lines over-expressing the AtBCB gene showed constitutive lignin production in whole roots. By contrast, wild-type Arabidopsis (Ler) produced a negligible level of lignin and enhanced lignin production in the root-tip region by Al stress. Compared with Ler, the AtBCB-expressing lines showed a lower deposition of malon dialdehyde after Al stress. Microscopic observation of the Al-treated roots indicated that the deposition of lipid peroxides was clearly low in the area where lignin accumulated. It was proposed that lipid peroxidation caused by Al stress was diminished by the formation of lignin. Expression of the NtGDI1 gene in yeast complemented the temperature-sensitive phenotype of a sec19 mutant at 37 degrees C. This gene also complemented an Al-sensitive phenotype shown by the sec19 mutant at the permissive temperature of 32 degrees C. These results suggested that the yeast Sec19 vesicle transport system has a function in providing basal Al resistance in yeast by the export of Al ions. It was also proposed that over-expression of the NtGDI1 protein activates an Al efflux system that protects Arabidopsis against Al toxicity.