Complexation of Hg with phytochelatins is important for plant Hg tolerance
14 Pag., 3 Tabl., 5 Fig. The definitive version is available at: www3.interscience.wiley.com ; Three-week-old alfalfa (Medicago sativa), barley (Hordeum vulgare) and maize (Zea mays) were exposed for 7 d to 30 µm of mercury (HgCl2) to characterize the Hg speciation in root, with no symptoms of being poisoned. The largest pool (99%) was associated with the particulate fraction, whereas the soluble fraction (SF) accounted for a minor proportion (<1%). Liquid chromatography coupled with electro-spray/time of flight mass spectrometry showed that Hg was bound to an array of phytochelatins (PCs) in root SF, which was particularly varied in alfalfa (eight ligands and five stoichiometries), a species that also accumulated homophytochelatins. Spatial localization of Hg in alfalfa roots by microprobe synchrotron X-ray fluorescence spectroscopy showed that most of the Hg co-localized with sulphur in the vascular cylinder. Extended X-ray Absorption Fine Structure (EXAFS) fingerprint fitting revealed that Hg was bound in vivo to organic-S compounds, i.e. biomolecules containing cysteine. Albeit a minor proportion of total Hg, Hg–PCs complexes in the SF might be important for tolerance to Hg, as was found with Arabidopsis thaliana mutants cad2-1 (with low glutathione content) and cad1-3 (unable to synthesize PCs) in comparison with wild type plants. Interestingly, high-performance liquid chromatography-electrospray ionization-time of flight analysis showed that none of these mutants accumulated Hg–biothiol complexes. ; This work was supported by Fundación Ramón Areces (http://www.fundacionareces.es), the Spanish Ministry of Science and Innovation (AGL2010-15151-PROBIOMET and AGL2007-61948), Comunidad de Madrid (EIADES S2009/AMB-1478), Junta Comunidades Castilla-La Mancha (FITOALMA, PBI07-0091-3644) and the Aragón Government (Group A03). The HPLC-ESI-TOFMS equipment was co-financed with EU FEDER funds. Portions of this research were carried out at the Stanford Synchrotron Radiation Lightsource, through the Structural Molecular Biology Program supported by the Department of Energy, Office of Biological and Environmental Research, and by the National Institute of Health, National Centre for Research Resources, Biomedical Technology Program. ; Peer reviewed