Phytoextraction of metals in contaminated soils: transfer and sequestration of cadmium and zinc in the hyperaccumulating plant Arabidopsis halleri – PHYMET

We are studying A. halleri plants with contrasted phenotypes collected on metallicolous and non metallicolous sites. The localization of metals is determined using SEM/EDX and µSXRF. Metal speciation is investigated using X-ray Absotption techniques (XAS) and hyphenated analytical techniques based on chromatography and mass spectrometry.

Plants from 4 sites were collected and exposed to various Zn treatments. A. halleri from Auby (M) was investigated in parallel, and we could determine the Cd distribution in leaves and protoplasts. The Cd speciation was also obtained by EXAFS and XANES. The sequestration of metals in trichomes were studied by µFTIR. The analyticial set up for hyphenated analytical techniques is in progress.

The collection of xylem sap is scheduled for the automn 2012.
Plants from the four origines will be phenotyped (Zn accumulation) during the summer 2012, and the localization and speciation will be done on the most contrasted phenotypes.
The analytical set-up for hyphenated techniques is still in progress.

see the report

Hyperaccumulating plants are able to tolerate and accumulate high amounts of metals in their shoots at concentrations that are toxic to ‘normal’ plants. This fascinating property can be used for the remediation of contaminated sites (phytoextraction). For that, and more generally for a better understanding of metal homeostasis in hyperaccumulators, it is necessary to understand how metals are transferred and stored in the aerial parts. Arabidopsis halleri is a Zn hyperaccumulator and one of the four Cd hyperaccumulator identified up to now. It is also a close relative to the model plant Arabidopsis thaliana, whose genome has been totally sequenced, and thus genetic and molecular tools available for A. thaliana can be used for A. halleri. In that context, A. halleri appears as an emerging model to investigate hyperaccumulation, and recent studies showed the role of metal transporters in Zn hyperaccumulation and Cd tolerance. It is now important to determine if the tolerance and accumulation traits, governed by genes, are linked to the mechanisms of transfer and storage of metals in the plant, still unknown.
The aim of this project is to progress in the understanding of hyperaccumulation by determining the mechanisms of transfer and sequestration of Zn and Cd in A. halleri populations with contrasted properties of accumulation and tolerance (phenotypes), and to make the connection between these mechanisms and the tolerance and accumulation traits. It is also to determine if specific ligands unkown to date are excreted at the root level to enhance metal bioavailability. For that, we propose to study four A. halleri populations with contrasted phenotypes originated from four metallicolous and non metallicollous European sites. The mechanisms will be studied originally by combining microscopies and spectroscopic synchrotron-based techniques (X-ray fluorescence, and X- ray Absorption, XAS), and hyphenated analytical techniques coupling Inductively Coupled Plasma Mass Spectrometry used as an atomic detector in chromatography and electrophoresis, and supported by ElectroSpray Ionization Mass Spectrometry for molecular identification. These state-of-the art tools in the field of metal speciation are totally complementary in terms of sensitivity: limitations resulting from detection limit and poor sensibility to organic molecules in XAS can be circumvented by analytic techniques, and artifacts resulting from sample preparation in analytical techniques can be tested by physical techniques, which do not require any sample extraction. We particularly intend to clarify the role of organic acids in binding Zn and Cd in shoots, the mechanism of transfer in the xylem sap and the role of nicotianamine and phytochelatins, and the role of trichomes, epidermal hairs covering the surface of the leaves, and of root exudates. We will also focus our attention on the study at the cellular and sub-cellular level by investigating isolated compartments (protoplasts and vacuoles). Thanks to results obtained at the tissue and cellular level, we will propose a scheme of trafficking and storage of metals for the different phenotypes. This information will be compared to results obtained in parallel studies on the gene expression induced under metal stress. The project consortium profits from the complementarity of its members having experience in the field of genetic adaptation of plants to metallic conditions, sample preparation, physical techniques, and analytical techniques, both applied to plant samples. With this project, we intend to promote this scarce interdisciplinary approach for future works.