[show abstract][hide abstract] ABSTRACT: * The mechanisms of enhanced root to shoot metal transport in heavy metal hyperaccumulators are incompletely understood. Here, we compared the distribution of nickel (Ni) over root segments and tissues in the hyperaccumulator Thlaspi caerulescens and the nonhyperaccumulator Thlaspi arvense, and investigated the role of free histidine in Ni xylem loading and Ni transport across the tonoplast. * Nickel accumulation in mature cortical root cells was apparent in T. arvense and in a high-Ni-accumulating T. caerulescens accession, but not in a low-accumulating T. caerulescens accession. * Compared with T. arvense, the concentration of free histidine in T. caerulescens was 10-fold enhanced in roots, but was only slightly higher in leaves, regardless of Ni exposure. Nickel uptake in MgATP-energized root- and shoot-derived tonoplast vesicles was almost completely blocked in T. caerulescens when Ni was supplied as a 1 : 1 Ni-histidine complex, but was uninhibited in T. arvense. Exogenous histidine supply enhanced Ni xylem loading in T. caerulescens but not in T. arvense. * The high rate of root to shoot translocation of Ni in T. caerulescens compared with T. arvense seems to depend on the combination of two distinct characters, that is, a greatly enhanced root histidine concentration and a strongly decreased ability to accumulate histidine-bound Ni in root cell vacuoles.
New Phytologist 05/2009; 183(1):106-16. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: The possible role of low molecular weight organic acids in the mechanism of increased zinc tolerance is investigated in Silene vulgaris (Moench) Garcke. In the leaves, the malate concentration is higher in zinc-tolerant plants than in zinc-sensitive ones, but no consistent differences are observed in the roots. Therefore, since the mechanism of increased zinc tolerance operates also in the roots themselves, a primary role for malate in the mechanism of increased zinc tolerance is doubtful. Exposure to 2inc does not affect the malate concentration in the plants. The citrate concentration in both the roots and leaves is higher in tolerant plants than in sensitive ones. A clear-cut effect of zinc on the concentration of citrate in the plants is not found. Citrate does not seem to play an important role in the sequestration of zinc in root cells, in view of its low concentration in the roots of tolerant plants. The possible role of citrate as a carrier for zinc in the cytosol is discussed. Oxalate, the most abundant organic acid, might play an important role in the sequestration of zinc, because of its high concentration in the plants. However, this cannot explain increased zinc tolerance, because the oxalate concentration is higher in the sensitive plants than in the tolerant ones. Neither succinate nor maleate play a prominent role in the mechanism of increased zinc tolerance. Malate and citrate are the only organic acids detected in the xylem fluid. The malate concentration in the xylem fluid is the same in sensitive and tolerant plants and it is not affected by the concentration of external zinc. The citrate concentration in the xylem fluid is about three times higher in the tolerant control plants than in the sensitive ones, but it decreases to the same level as in the sensitive plants after exposure to zinc. Differences in the citrate concentrations in the xylem fluid of sensitive and tolerant plants are not related to differences in the allocation of zinc.
New Phytologist 04/2006; 126(4):615 - 621. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: A few plant species, like Silene vulgaris, have evolved ecotypes, that are capable of growing on metal-enriched soils. These tolerant ecotypes exhibit metal-specific
resistance mechanisms, which are not completely understood to date. Resistance to copper, zinc and cadmium seems to be regulated
by different major genes and some hypostatic modifiers. Metallothionein (MT2b) expression is involved in the level of copper
tolerance, while an enhanced ATP-dependent copper efflux across the root cell plasma membrane may result in a decreased copper-accumulation
and contributes to the higher resistance of mine plants compared to the copper-sensitive plants. Given the lack of evidence
for the significance of similar strategies for zinc and cadmium (such as reduction of uptake and increased binding to intracellular
molecules) vacuolar compartmentation is generally believed to be the basic mechanism of tolerance of zinc and cadmium in higher
plant species. An enhanced tonoplast transport of zinc in roots plays a role in zinc tolerance, but the existence of a similar
tolerance mechanism for cadmium remains to be demonstrated. In our present research, the use of molecular biological techniques
to study the presence and expression of (transporter) genes (like ZAT, ZNT and other families) has been initiated. Introduction
A small number of higher plant species is capable of growing on soils heavily contaminated with zinc, copper, lead, cadmium,
nickel or arsenic (Ernst, 1974). The so-called metallophytes have evolved genetically based increased levels of resistance
to the metals involved (Schat and Verkleij, 1998). Resistance to copper, zinc and cadmium seems to be regulated by different
major genes and some hypostatic modifiers (in the case of copper resistance, Schat and Vooijs, 1996). Heavy metal resistance
may result from the ability to prevent uptake (avoidance) or from the ability to cope with high amounts of heavy metals in
the tissues (tolerance). If avoidance plays a role, it may be expected to result from changes in the permeability of the plasma
membrane. Evidence for an important role of the plasma membrane has been provided in the case of copper resistance (De Vos
et al., 1991). There are no indications that the plasma membrane plays a role in zinc or cadmium resistance. These resistances should
be based on increased capacities to sequester metals within the plant, which could be effected by synthesis of specific metal-binding
compounds, subcellular compartmentation (vacuole) or a combination of both. In this contribution we will give a short overview
of the state of the art with respect to the mechanisms of copper, zinc and cadmium resistance in Silene vulgaris, a very common metallophyte, which are under investigation in the authors’ laboratory.
[show abstract][hide abstract] ABSTRACT: We studied copper uptake in inside-out plasma membrane vesicles derived from roots of copper-sensitive, moderately copper-tolerant and highly copper-tolerant populations of Silene vulgaris (Amsterdam, Marsberg and Imsbach, respectively). Plasma membrane vesicles were isolated using the two-phase partitioning method and copper efflux was measured using direct filtration experiments. Vesicles derived from Imsbach plants accumulated two and three times more copper than those derived from Marsberg and Amsterdam plants, respectively. This accumulation was ATP-dependent. Also, 9-amino-6-chloro-2-methoxyacridine fluorescence quenching rates upon copper addition decreased in the order Imsbach>Marsberg>Amsterdam. Our results support the hypothesis that efflux of copper across the root plasma membrane plays a role in the copper tolerance mechanism in S. vulgaris.
Physiologia Plantarum 11/2001; 113(2):225-232. · 3.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: 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.
Journal of Agricultural and Food Chemistry 10/2000; 48(9):4014-9. · 2.91 Impact Factor
[show abstract][hide abstract] ABSTRACT: Terrestrial metal-toxic soils are found all over the world. They originate either from geological anomalies (e.g., ore outcrops,
serpentines), or from human activities (e.g., mining, smelting, agricultural use of metal containing fertilizers, pesticides,
sludges, etc.). Even extremely metal-toxic soils eventually become vegetated, though often sparsely and without trees. These
vegetations are usually poor in species, particularly in parts of the world that have been glaciated during the Pleistocene
(Brooks et al., 1985), most probably mainly due to a strong selective effect of metal-toxicity as such (Schat and Verkleij, 1998).
[show abstract][hide abstract] ABSTRACT: The results presented in this paper describe the short- and long-term toxicity of arsenate in Silene vulgaris. Short-term toxicity, measured as inhibition of root elongation, depended on phosphate nutrition, arsenate being much less toxic at high phosphate supply. At low phosphate levels more arsenic was taken up by the plants. Under chronic exposure, toxicity (measured as inhibition of biomass production) did not increase with time. In addition, the accumulation of phytochelatins (PCs) as a function of toxicity and duration of exposure was studied. Short-term PC accumulation (over a 3 d period) was positively correlated with exposure. Isolation of peptide complexes from prolongedly exposed plants showed that PC2, PC3 and PC4 were present, although the latter not until at least 3 d exposure. Arsenic co-eluted mainly with PC2 and PC3. Fractions containing PC4 were devoid of As, probably due to dissociation of the complexes during extraction or elution. The breakdown of PCs after arresting As exposure was very slow. This could explain the continuous accumulation of PCs throughout longer periods of As exposure.
New Phytologist 10/1999; 144(2):223 - 232. · 6.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: It was demonstrated recently that isolated tonoplast vesicles derived from plants of a Zn-tolerant ecotype of Silene vulgaris accumulate more Zn than vesicles derived from a Zn-sensitive ecotype. We have now characterized the tonoplast-transport system that causes this uptake difference and demonstrated its genetic correlation to Zn tolerance using plant crosses. We conclude that the tonoplast Zn uptake system of the tolerant ecotype differs greatly in its characteristics from that of the sensitive one, with the most prominent differences being its insensitivity to protonophores and ortho-vanadate and its stimulation by Mg-GTP. These differences in characteristics are most likely due to the fact that Zn can be taken up by two or more parallel pathways, which are not present in the same proportions in both ecotypes. In both ecotypes, Zn is actively transported across the tonoplast (temperature coefficient > 1.6), most likely as a free ion, since citrate does not accumulate in vesicles. Most importantly, the uptake difference found using the ecotypes was also found between homozygous Zn-tolerant and Zn-sensitive F3 plants, proving the genetic correlation between increased tonoplast Zn transport and naturally selected Zn tolerance in S. vulgaris.
[show abstract][hide abstract] ABSTRACT: Compartmentalization of zinc in the vacuole has been proposed to explain enhanced zinc tolerance in higher plants. In order to determine whether the vacuolar membrane is involved in differential zinc tolerance, tonoplast vesicles were isolated from roots of zinc tolerant and sensitive Silene vulgaris. In vitro transport studies with K+/nigericin energized tonoplast vesicles showed that zinc (supplied as zinc citrate) was transported across the tonoplast only in the presence of MgATP. Zinc transport was about 2.5 times higher in tonoplast vesicles derived from zinc tolerant Silene vulgaris compared with those derived from sensitive Silene vulgaris at high zinc concentration (0.8 mmol/L). At low zinc concentrations (0.1–0.3 mmol/L) there was no significant difference in zinc transport between both types of plants. The difference in zinc transport at high external zinc concentrations could be due to an additional transport system with lower zinc affinity that may operate at higher zinc concentrations.
Overall, these data strongly suggest that the tonoplast plays an important role in naturally selected zinc tolerance.
Journal of Plant Physiology. 08/1998; 153(s 1–2):188–191.
[show abstract][hide abstract] ABSTRACT: In response to external Cd supply Cd-tolerant Silene vulgaris plants produce three times less phytochelatins than Cd-sensitive ones. Phytochelatin synthase activity in roots of both ecotypes was measured in order to determine whether a lower enzyme activity could be responsible for the lower phytochelatin production. The phytochelatin synthase activity in tolerant plants, grown in the absence or in the presence of Cd, tended to be slightly lower than that in sensitive ones. However, the difference was too small to account for the differential phytochelatin contents observed. Observations on phytochelatin synthesis in excised roots and on the recovery of the level of glutathione, the precursor of phytochelatins, after arresting the Cd exposure, suggested that tolerant plants did not exhibit a decreased capacity to synthesize glutathione. Furthermore, the relative decrease in the phytochelatin concentration after arresting the Cd-exposure was identical for both populations, indicating that the difference in phytochelatin concentrations was not the result of a differential rate of phytochelatin degradation.