Co-segregation analysis of cadmium and zinc accumulation in Thlaspi caerulescens interecotypic crosses.

• Cadmium (Cd) hyperaccumulation in Thlaspi caerulescens varies among ecotypes. Here we investigated segregation of Cd and zinc (Zn) accumulation in F2 crosses between high (Ganges) and low (Prayon) Cd-accumulating ecotypes. • Accumulation was measured in plants grown in compost treated with 5 and 100 mg kg-1 Cd and Zn, respectively, and in hydroponics with 50 m Zn and 10 or 50 m Cd. Another hydroponic experiment examined the relationship between Cd tolerance and accumulation. • Parental phenotype distributions for shoot metal concentrations were distinct for Cd, but not consistent for Zn. Shoot Cd and Zn in F2 s varied continuously, with significant transgression for Zn in all treatments. Shoot Cd correlated strongly with shoot manganese (Mn), and to a lesser degree with shoot Zn. Shoot Cd concentrations in the Cd nontolerant F2 s were lower than, or similar to, those in the Cd-tolerant F2 s. • We conclude that Cd and Zn accumulation is governed by multiple genes, and that Cd tolerance and accumulation are independent traits in T. caerulescens. Two uptake systems with distinctive affinities for Cd, Zn and Mn are proposed.

Derivatization of phytochelatins from Silene vulgaris, induced upon exposure to arsenate and cadmium: comparison of derivatization with Ellman's reagent and monobromobimane.

Phytochelatins (PCs) are a family of thiol-rich peptides, with the general structure (gamma-Glu-Cys)(n)()-Gly, with n = 2-11, induced in plants upon exposure to excessive amounts of heavy metals and some metalloids, such as arsenic. Two types of PC analyses are currently used, i.e., acid extraction and separation on HPLC with either precolumn derivatization (pH 8.2) with monobromobimane (mBBr) or postcolumn derivatization (pH 7.8) with Ellman's reagent [5, 5'-dithiobis(2-nitrobenzoic acid), DTNB]. Although both methods were satisfactory for analysis of Cd-induced PCs, formation of (RS)(3)-As complexes during extraction of As-induced PCs rendered the DTNB method useless. This paper shows that precolumn derivatization with mBBr, during which the (RS)(3)-As complexes are disrupted, provides a qualitative and quantitative analysis of both Cd- and As-induced PCs. In addition, derivatization efficiencies of both methods for the oligomers with n = 2-4 (PC(2)(-)(4)) are compared. Derivatization efficiency decreased from 71.8% and 81.4% for mBBr and DTNB derivatization, respectively, for PC(2) to 27.4% and 50.2% for PC(4). This decrease is most likely due to steric hindrance. Correction of measured thiol concentration is therefore advised for better quantification of PC concentrations in plant material.

Copper tolerance in Silene cucubalus : Subcellular distribution of copper and its effects on chloroplasts and plastocyanin synthesis.

The uptake, translocation and subcellular distribution of copper as well as its effect on chloroplasts and plastocyanin synthesis were studied in a copper-sensitive and a copper-tolerant population of Silene cucubalus (L.) Wib. As a function of time, the copper concentration in roots of tolerant plants increased more slowly than that in roots of sensitive ones. Translocation to the shoot occurred more rapidly in tolerant plants than in sensitive ones. Although it was accumulated in leaf cells, copper was not accumulated in the chloroplasts of either sensitive or tolerant plants. Chlorophyll content was not affected by copper in tolerant plants, whereas sensitive plants became chlorotic. Plastocyanin synthesis was not enhanced as a result of high copper concentrations and no difference in plastocyanin content between tolerant and sensitive plants was detected. Measurements of copper in purified cell walls revealed that storage of the metal in cell-wall material does not play an important role in tolerance mechanism. Uptake characteristics, distribution and cytoplasmic detoxification of copper are discussed.

Bioindication of a surplus of heavy metals in terrestrial ecosystems.

A survey of the methods of boindication of heavy metals in terrestrial ecosystems and their effectiveness for predicting the consequences of environmental stress on organisms is presented. Two main inputs of heavy metals for terrestrial ecosystems have been considered: airborne and soil-borne.Airborne metals can be monitored due to physical adsorption on plant surfaces or due to chemical exchange processes in cell walls. Active biomonitoring widely uses both aspects, however, without predictive values.Meaningful bioindication of soilborne heavy metals can only be achieved by passive monitoring. Due to the different functions of heavy metals in organisms-micronutrients and trace elements-the knowledge of natural background values is important, considering the qualitative aspects of metals in the soil. In exceptional situations morphological and anatomical changes of plant organs will facilitate bioindication; in every case chemical analysis of the concentration of heavy metals is an essential part of the monitoring program.A long-term exposure of organisms to heavy metals will influence the genetic structure of populations. Therefore measurement of heavy metal tolerance of plants has to be a standard procedure in monitoring programs.

Glutathione Depletion Due to Copper-Induced Phytochelatin Synthesis Causes Oxidative Stress in Silene cucubalus.

The relation between loss of glutathione due to metal-induced phytochelatin synthesis and oxidative stress was studied in the roots of copper-sensitive and tolerant Silene cucubalus (L.) Wib., resistant to 1 and 40 micromolar Cu, respectively. The amount of nonprotein sulfhydryl compounds other than glutathione was taken as a measure of phytochelatins. At a supply of 20 micromolar Cu, which is toxic for sensitive plants only, phytochelatin synthesis and loss of total glutathione were observed only in sensitive plants within 6 h of exposure. When the plants were exposed to a range of copper concentrations for 3 d, a marked production of phytochelatins in sensitive plants was already observed at 0.5 micromolar Cu, whereas the production in tolerant plants was negligible at 40 micromolar or lower. The highest production in tolerant plants was only 40% of that in sensitive plants. In both varieties, the synthesis of phytochelatins was coupled to a loss of glutathione. Copper at toxic concentrations caused oxidative stress, as was evidenced by both the accumulation of lipid peroxidation products and a shift in the glutathione redox couple to a more oxidized state. Depletion of glutathione by pretreatment with buthionine sulfoximine significantly increased the oxidative damage by copper. At a comparably low glutathione level, cadmium had no effect on either lipid peroxidation or the glutathione redox couple in buthionine sulfoximine-treated plants. These results indicate that copper may specifically cause oxidative stress by depletion of the antioxidant glutathione due to phytochelatin synthesis. We conclude that copper tolerance in S. cucubalus does not depend on the production of phytochelatins but is related to the plant's ability to prevent glutathione depletion resulting from copper-induced phytochelatin production, e.g. by restricting its copper uptake.

Phytochelatins are involved in differential arsenate tolerance in Holcus lanatus.

Arsenate tolerance is conferred by suppression of the high-affinity phosphate/arsenate uptake system, which greatly reduces arsenate influx in a number of higher plant species. Despite this suppressed uptake, arsenate-tolerant plants can still accumulate high levels of As over their lifetime, suggesting that constitutive detoxification mechanisms may be required. Phytochelatins are thiol-rich peptides, whose production is induced by a range of metals and metalloids including arsenate. This study provides evidence for the role of phytochelatins in the detoxification of arsenate in arsenate-tolerant Holcus lanatus. Elevated levels of phytochelatin were measured in plants with a range of tolerance to arsenate at equivalent levels of arsenate stress, measured as inhibition of root growth. The results suggest that arsenate tolerance in H. lanatus requires both adaptive suppression of the high-affinity phosphate uptake system and constitutive phytochelatin production.