Effects of Interaction Between Cadmium and Plumbum on Phytochelatins and Glutathione Production in Wheat （Triticum aestivum L.）

Phytochelatins (PCs) may function as a potential biomarker for metal toxicity. However, less attention has been paid to the effects of metal interactions on the production of PCs and glutathione (GSH), the most prominent cellular thiol. In the present study, the effects of interactions between cadmium (Cd) and plumbum (Pb) on the production of PCs and GSH were monitored over a period of 14 d in wheat (Triticum aestivum L.) tissues. The results showed that combination of Cd and Pb led to synergistic growth inhibition in wheat. Exposure to Cd or Pb increased levels of PCs in a concentration-, tissue-, and time-dependent manner. Cadmium was more effective that Pb in increasing PCs production. Compared with the effects of Cd or Pb alone on the production of PCs, the combination of Cd and Pb acted synergistically, resulting in an enhanced production of PCs. Cadmium also stimulated GSH production in a concentration-, tissue-, and time-dependent manner. However, Pb had no obvious effects on GSH levels. The combination of Pb and Cd antagonized GSH production over the course of the growth period. The results of the present study suggest that metal interactions should be considered in the application of PCs and GSH as potential biomarkers for the evaluation of metal toxicity.

The importance of glutathione and phytochelatins on the selenite and arsenate detoxification in Arabidopsis thaliana

We investigated the role of glutathione（GSH） and phytochelatins（PCs） on the detoxification of selenite using Arabidopsis thaliana. The wild-type（WT） of Arabidopsis thaliana and its mutants（glutathione deficient Cad 2–1 and phytochelatins deficient Cad 1–3） were separately exposed to varying concentrations of selenite and arsenate and jointly to both toxicants to determine their sensitivities. The results of the study revealed that, the mutants were about 20-fold more sensitive to arsenate than the WT, an indication that the GSH and PCs affect arsenate detoxification. On the contrary, the WT and both mutants showed a similar level of sensitivity to selenite, an indication that the GSH and PCs do not significantly affect selenite detoxification. However, the WT is about 8 times more sensitive to selenite than to arsenate, and the mutants were more resistant to selenite than arsenate by a factor of 2. This could not be explained by the accumulation of both elements in roots and shoots in exposure experiments. The co-exposure of the WT indicates a synergistic effect with regards to toxicity since selenite did not induce PCs but arsenic and selenium compete in their PC binding as revealed by speciation analysis of the root extracts using HPLC–ICP–MS/ESI–MS. In the absence of PCs an antagonistic effect has been detected which might suggest indirectly that the formation of Se glutathione complex prevent the formation of detrimental selenopeptides. This study, therefore, revealed that PC and GSH have only a subordinate role in the detoxification of selenite.

Root cell walls and phytochelatins in low-cadmium cultivar of Brassica parachinensis

‘Lvbao-701’ is a cultivar of Chinese flowering cabbage(Brassica parachinensis) that exhibits low cadmium(Cd) accumulation and high Cd tolerance.In this study, this cultivar was compared with a high-Cd accumulating cultivar, ‘Chixin-4’, to characterize the mechanisms influencing Cd accumulation in B. parachinensis. Root cell walls were isolated by dissolving the cytoplasm with an organic solvent, and root Cd and phytochelatin(PC) contents were analyzed. In addition, a PC synthase gene fragment was cloned and expressed under Cd stress conditions. The proportions of Cd bound to root cell walls were higher in the ‘Lvbao-701’ plants(68.32%–76.80%) than in the ‘Chixin-4’ plants(35.36%–54.18%) after exposure to Cd stress. The proportions of Cd bound to root cell walls measured using cell walls isolated with a non-destructive method were higher than those obtained using a conventional method that required grinding and centrifugation. Exposure to Cd stress induced the PC production and resulted in higher PC contents in the ‘Lvbao-701’ plants than in the ‘Chixin-4’ plants. Cloning and expression analysis of a B. parachinensis PC synthase cDNA fragment indicated that PC synthase gene expression was induced by Cd and occurred mainly in the roots of both ‘Lvbao-701’ and ‘Chixin-4’ plants. However, the PC synthase gene expression level was higher in the‘Lvbao-701’ roots than in the ‘Chixin-4’ roots. Therefore, a higher abundance of Cd in the root cell walls of ‘Lvbao-701’ and up-regulated PC production in the roots are probably the main reasons why ‘Lvbao-701’ exhibits lower Cd translocation to the shoots and higher tolerance to Cd than ‘Chixin-4’.

Selenium Modulates Growth and Thiol Metabolism in Wheat (<i>Triticum aestivum</i>L.) during Arsenic Stress

Arsenic affects plants by disrupting their growth and metabolism while selenium, an essential micronutrient has beneficial role in stress tolerance. Owing to the antioxidative capacity of selenium, it can counteract detrimental effects of arsenic induced stress in plants. The interactive influence of arsenate and selenate on the growth, arsenic and selenium accumulation, activities of non-enzymatic and enzymatic antioxidants, levels of ascorbate, α-tocopherol, total glutathione and activities of glutathione regulatory enzymes along with characterization and quantification of phytochelatins in growing wheat (Triticum aestivum L.) seedlings were investigated. Rate of arsenic accumulation was increased in arsenate treated seedlings while in seedlings treated jointly with arsenate and selenate, arsenic accumulation decreased. Arsenate stress resulted decrease in ascorbate and total glutathione contents, activities of the glutathione metabolism enzymes while significantly increased the levels of α-tocopherol and phytochelatins (PC2 and PC4), along with activities of ascorbate peroxidase and ascorbic acid oxidase in the test seedlings. The effects were more pronounced in roots than in shoots. Joint application of arsenate along with selenate was able to overcome the adverse effects caused by arsenic alone to variable extents by exhibiting significant alterations of all parameters tested, imparting better growth and thiol metabolism in the test seedlings. Our results conclude that application of selenium fertilizer in arsenic contaminated soil might be favourable to improve growth and defense ability in wheat against arsenic toxicity.

Comparison of physiological responses to oxidative and heavy metal stress in seedlings of rice paddy, Oryza sativa L.

Physiological responses on the bases of activities of antioxidant enzymes: peroxidase, catalase, superoxide dismutase and glutathione reductase as well as estimation of total protein, lipid peroxidation and thiols in the form of protein, non\|protein, glutathione and phytochelatin measured in growing seedlings of paddy, Oryza sativa L., from day 2 to 8 were compared following treatment of seeds for 5h with oxidative agents, paraquat 5 × 10 -5 , 10 -4 , 10 -3 mol/L, H 2O 2 10 -3 , 5×10 -3 , 10 -2 mol/L, and CdCl 2 10 -5 , 10 -4 , 5×10 -3 mol/L. A significant induction of all antioxidant enzymes along with an increase in the levels of protein, lipid peroxidation and glutathione was noted in response to oxidative stress, CdCl 2 induced significant peroxidase and catalase activities but not superoxide dismutase. In a marked contrast from oxidative stress, CdCl 2 decreased glutathione reductase activity as well as glutathione levels but increased phytochelatin level. The different physiological responses thus underlined the crucial involvement of glutathione and phytochelatin in the oxidative and heavy metal\|linduced adaptive responses respectively.

Arsenic Stress Responses and Tolerance in Rice: Physiological, Cellular and Molecular Approaches

Arsenic （As）, a potentially toxic metalloid released in the soil environment as a result of natural as well as anthropogenic processes, is subsequently taken up by crop plants. In rice grains, As has been reported in Asia, North America and Europe, suggesting a future threat to food security and crop production. As3＋ by dint of its availability, mobility and phytotoxicity, is the most harmful species of As for the rice crop. Specific transporters mediate the transport of different species of As from roots to the aboveground parts of the plant body. Accumulation of As leads to toxic reactions in plants, affecting its growth and productivity. Increase in As uptake leads to oxidative stress and production of antioxidants to counteract this stress. Cultivars tolerant to As stress are efficient in antioxidant metabolism compared to sensitive ones. Iron and selenium are found to have ameliorating effect on the oxidative stress caused by As. Microbes, even many indigenous ones, in the plant rhizosphere are also capable of utilizing As in their metabolism, both independently and in association. Some of these microbes impart tolerance to As-stress in plants grown in As contaminated sites.

Arsenic speciation for the phytoremediation by the Chinese brake fern,Pteris vittata

Arsenic(As)speciation for the phytoremediation by the Chinese brake fern was studied.In particular,the mechanism of how plants induce compounds containing thiol(SH)and proteins by As exposure in terms of the relationship between As and phosphate uptaken into plant cells was examined.Pteris vittata callus could effciently reduce As(V)to As(III)by the rapid introduction of reductase and synthesize thiols leading to phytochelatins production.Furthermore,Pteris vittata could control phosphate concentration in t...

Physiological and biochemical analysis of mechanisms underlying cadmium tolerance and accumulation in turnip

The capacity of plants to accumulate cadmium(Cd) is significant for phytoremediation of Cd-polluted soils. Turnips cultivated in China include species featuring high Cd accumulation and some of these plants act as Cd hyperaccumulator landraces. These plants can accumulate over 100 mg Cd kg^(-1) dry weight in leaves without injury. Hence, studies that explore mechanisms underlying Cd detoxification and transport in turnip plants are essential. In the present study, we compared physiological and biochemical changes in turnip leaves treated with two Cd concentrations to controls. We discovered that Cd stress significantly increased the enzymatic activities or compound contents in the antioxidant system, including members of the glutathione-ascorbic acid cycle, whereas oxidation of reactive oxygen species(ROS) remained stable. Cd treatments also increased the contents of phytochelatins as well as a number of amino acids. Based on these results, we conclude that turnips initiate a series of response processes to manage Cd treatment. First, the antioxidant system maintaining ROS homeostasis and osmotic adjustment is excited to maintain stability of cell osmotic potential. Cd is chelated into its stable form to reduce its toxicity. Cd is possibly transported to vacuoles or non-protoplasts for isolation. Amino acid synthesis may directly and indirectly play an important role in these processes. This study partly revealed physiological and biochemical mechanisms underlying turnip response to Cd stress and provides information on artificially increasing or decreasing Cd accumulation in turnips and other plants.

Homologous cloning, characterization and expression of a new halophyte phytochelatin synthase gene in Suaeda salsa

The halophyte Suaeda salsa can grow in heavy metal-polluted areas along intertidal zones having high salinity.Since phytochelatins can effectively chelate heavy metals,it was hypothesized that S.salsa possessed a phytochelatin synthase(PCS) gene.In the present study,the cDNA of PCS was obtained from S.salsa(designated as SsPCS) using homologous cloning and the rapid amplification of cDNA ends(RACE).A sequence analysis revealed that SsPCS consisted of 1 916 bp nucleotides,encoding a polypeptide of 492 amino acids with one phytochelatin domain and one phytochelatin C domain.A similarity analysis suggested that SsPCS shared up to a 58.6%identity with other PCS proteins and clustered with PCS proteins from eudicots.There was a new kind of metal ion sensor motif in its C-terminal domain.The SsPCS transcript was more highly expressed in elongated and fibered roots and stems(P<0.05) than in leaves.Lead and mercury exposure significantly enhanced the mRNA expression of SsPCS(P<0.05).To the best of our knowledge,SsPCS is the second PCS gene cloned from a halophyte,and it might contain a different metal sensing capability than the first PCS from Thellungiella halophila.This study provided a new view of halophyte PCS genes in heavy metal tolerance.

How Plants Cope with Cadmium: Staking All on Metabolism and Gene Expression

Environmental pollution is one of the major problems for human health. Toxic heavy metals are normally present as soil constituents or can also be spread out in the environment by human activity and agricultural techniques. Soil contamination by heavy metals as cadmium, highlights two main aspects： on one side they interfere with the life cycle of plants and therefore reduce crop yields, and on the other hand, once adsorbed and accumulated into the plant tissues, they enter the food chain poisoning animals and humans. Considering this point of view, understanding the mechanism by which plants handle heavy metal exposure, in particular cadmium stress, is a primary goal of plant-biotechnology research or plant breeders whose aim is to create plants that are able to recover high amounts of heavy metals, which can be used for phytoremediation, or identify crop varieties that do not accumulate toxic metal in grains or fruits. In this review we focus on the main symptoms of cadmium toxicity both on root apparatus and shoots. We elucidate the mechanisms that plants activate to prevent absorption or to detoxify toxic metal ions, such as synthesis of phytochelatins, metallothioneins and enzymes involved in stress response. Finally we consider new plant-biotechnology applications that can be applied for phytoremediation.

Molecular Mechanisms and Genetic Basis of Heavy Metal Tolerance/ Hyperaccumulation in Plants

Phytoremediation has gained increased attention as a cost-effective method for the remediation of heavy metal-contaminated sites. Because some plants possess a range of potential mechanisms that may be involved in the detoxification of heavy metals, they manage to survive under metal stresses. High tolerance to heavy metal toxicity could rely either on reduced uptake or increased plant internal sequestration, which is manifested by an interaction between a genotype and its environment. The growing application of molecular genetic technologies has led to increased understanding of mechanisms of heavy metal tolerance/accumulation in plants and, subsequently, many transgenic plants with increased heavy metal resistance, as well as increased uptake of heavy metals, have been developed for the purpose of phytoremediation. In the present review, our major objective is to concisely evaluate the progress made so far in understanding the molecular/cellular mechanisms and genetic basis that control the uptake and detoxification of metals by plants.

Physiological responses and detoxific mechanisms to Pb,Zn,Cu and Cd in young seedlings of Paulownia fortunei

Paulownia fortunei has been successfully used in the phytoremediation of many Pb/Zn mine tailings. However, seed germination and young seedlings of P. fortunei rarely occurred in these mine tailings. The physiological responses and detoxific mechanisms of P. fortunei young seedling to Pb, Zn, Cu and Cd stress were investigated. The germinated rate, shoot length, chlorophyll and carotenoid contents in leaves of young seedlings had a great reduction under Zn and Cu treatments, but had little decrease under Pb and Cd treatments. The production rate of O2＊-, H2O2 and malondialdehyde （MDA） contents significantly increased in response to added Zn and Cu indicating great oxidative stress for young seedlings, but they had no significant change to added Pb and Cd. Young seedlings had effective detoxific mechanism to Pb and Cd, as antioxidant enzymes activities, phytochelatins （PCs-SH） and proline contents increased with increasing rates of added Pb and Cd. However, young seedlings had un-effective detoxific mechanisms to Zn and Cu stress. Results revealed the heavy metals （such as Ca） that present at low concentrations in mine tailings may be major constraint for the survival of young seedlings.

Effects of EDTA and plant growth-promoting rhizobacteria on plant growth and heavy metal uptake of hyperaccumulator Sedum alfredii Hance

Phytoremediation is a cost-effective and environment-friendly strategy for decontaminating heavy-metal-contaminated soil.However, the practical use of phytoremediation is constrained by the low biomass of plants and low bioavailability of heavy metals in soil.A pot experiment was conducted to investigate the effects of the metal chelator ethylenediaminetetraacetic acid(EDTA) and EDTA in combination with plant growth-promoting rhizobacteria(Burkholderia sp.D54 or Burkholderia sp.D416) on the growth and metal uptake of the hyperaccumulator Sedum alfredii Hance.According to the results, EDTA application decreased shoot and root biomass by 50% and 43%, respectively.The soil respiration and Cd,Pb, Zn uptake were depressed, while the photosynthetic rate, glutathione and phytochelatin(PC) contents were increased by EDTA application.Interestingly, Burkholderia sp.D54 and Burkholderia sp.D416 inoculation significantly relieved the inhibitory effects of EDTA on plant growth and soil respiration.Compared with the control, EDTA + D416 treatment increased the Cd concentration in shoots and decreased the Pb concentration in shoots and roots, but did not change the Zn concentration in S.alfredii plants.Furthermore,EDTA, EDTA + D54 and EDTA + D416 application increased the cysteine and PC contents in S.alfredii(p < 0.05);among all tested PCs, the most abundant species was PC2, and compared with the control, the PC2 content was increased by 371.0%, 1158.6% and 815.6%,respectively.These results will provide some insights into the practical use of EDTA and PGPR in the phytoremediation of heavy-metal-contaminated soil by S.alfredii.

Enhanced Cadmium Accumulation in Transgenic Tobacco Expressing the Phytochelatin Synthase Gene of Cynodon dactylon L

Bermudagrass （Cynodon dactylon L. cv. Goldensun） is highly resistant to and accumulates large amounts of cadmium （Cd）. A phytochelatin synthase （PCS） cDNA （CdPCS1） was isolated from this grass by rapid amplification of cDNA ends. The putative CdPCS1 protein shared a high homology with PCS from other plants, with 79% homology at the N-terminal and 47% homology at the C-terminah However, 16 Cys residues were found at the C-terminal of CdPCS1, and among these residues, three positions were different from other PCS proteins. Semiquantitative reverse transcription-polymerase chain reaction analysis showed that Cd stress induced CdPCS1 expression in both roots and leaves in Bermudagrass. We verified that CdPCS1 plays an important role in Cd tolerance in yeast cells by expressing the gene in ABDE1, a Cdsensitive mutant. CdPCS1 was then introduced into tobacco plants. The phytochelatin level in some transgenic tobacco lines increased 3.88-fold more than in wild type plants and Cd accumulation in these transgenic plants was enhanced 3.21-fold accordingly. The results suggested that CdPCS1 could be used as a gene element for phytoremediation in the future.

RNA Interference-mediated Silencing of Phytochelatin Synthase Gene Reduce Cadmium Accumulation in Rice Seeds

Phytochelatins （PCs） play an important role in heavy metal resistance and accumulation. To reduce the accumulation of cadmium （Cd） in rice seeds, the expression of phytochelatin synthase （PCS） gene OsPCS1 was suppressed by RNA interference （RNAi）. A hairpin construct of a PCS fragment was designed in the pRNAi-OsPCS1 under the control of ZMM1, a seed-specific promoter from maize. The construct was introduced into rice （japonica） through Agrobacterium tumefaciens. The RNAi rice plantlets were selected and cultivated in pots exposured to 10 mg/kg Cd. The transcriptional level of OsPCS1 declined in seeds of some RNAi rice compared to the wild type. As a result Cd accumulation was reduced by about half in the seeds of RNAi rice. As expected, no apparent difference of growth appeared between RNAi and wild-type plants. The results suggest that this new approach can be used to control heavy metal accumulation in crops.

The Purification and Rapid Identification of Heavy Metal-binding Peptides of Water Hyacinth

This paper studies the rapid identification of heavy metal-binding peptides (phytochelatin) by taking Water Hyacinth as a model plant. Plants were cultured in water containing 2 μg/ml Cd2+ for 13 days. The Sephadex G-50 chromatography of root extract under low salt concentration (0. 01 mol/L PBS) gave a Cd-binding peak with MW of 10 ,000 determined by SEC HPLC. After oxidation with performic acid, its SEC HPLC molecular weight decreased to below 1300 and the reverse phase HPLC showed one peptide peak, whose amino acid composition is the same as that of the sample never undertaking oxidation, and (Glu/Gln):Cys:Gly=2:2:1. According to the general structure of phytochelatin (γ-Glu-Cys)n-Gly, n is 2 in this case. The protocol including the sequential steps of Sephadex G-50 chromatography→performic acid oxidation→reverse phase HPLC→amino acid analysis is a rapid and effective method to identify the existence of phytochelatin and determine its values of n.