Effects of nanomolar heavy metal concentrations on plants - comparison of biochemical&biophysical mechanisms of deficiency and sublethal toxicity under environmentally relevant conditions

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In this project we are investigating mechanisms of heavy metal toxicity and deficiency. We started this in the water plant Ceratophyllum demersum as a model for shoots, and now also transfer the knowledge obtained from this model to crop plants. While many potential mechanisms of metal toxicity induced damage have been proposed, most of them were investigated only under rather artificial laboratory conditions. Therefore, we are evaluating the relevance of different proposed mechanisms of metal-induced inhibition under environmentally relevant conditions and are comparing these results with those obtained under conditions that were used in identifying the putative toxicity mechanisms. We have chosen copper, chromium and cadmium as model heavy metals, plus arsenic as a toxic metalloid, because they most frequently reach toxic concentrations in contaminated environments. We apply environmentally relevant conditions in terms of concentrations, biomass/water volume ratios, light and temperature cycles and incubation times. In the past 39 months of the project, the focus was at the beginning on the analysis of the toxicity of lake Ammelshain samples and the determination of reasons for their toxicity by systematic re-constitution of field conditions in the laboratory. Afterwards we systematically investigated mechanisms of deficiency of copper and chromium, as well as toxicity of arsenic, cadmium and copper. The deficiency & toxicity analysis was started with in vivo measurements of photosynthesis biophysics, formation of reactive oxygen species and oxygen exchange, combined with pigment, starch and metal analyses, and is currently being continued by analysis of cellular and subcellular metal distribution and speciation, as well as metalloproteomics. In the coming years, we want to transfer the knowledge obtained with the model plant to crop species (mainly soybean and barley), with special emphasis on cellular and subcellular metal distribution (via µXRF) and speciation (via µXANES), as well as metalloproteomics and metabolomics. Thus we want to investigate further how in plants metal deficiency and onset of metal toxicity lead to a change in metal distribution between different tissues, different cellular compartments and different metal-binding proteins, rather than a general concentration change in many tissues. Further, we want to clone the genes of novel metal-binding proteins to further characterise them and reveal their function by modifying their expression in the crop species and Arabidopsis thaliana. This particularly applies the proteins that bind Cr under optimal and Cr-deficient conditions, where we already have a preliminary identification of one of these proteins.