Nitrogen often limits primary productivity in the terrestrial ecosystems and is also the element whose biogeochemical cycling has been most altered since the industrial revolution. In forests of the temperate and boreal climate zones, most nitrogen is assimilated in trees through ectomycorrhizal (ECM) fungi. In vitro studies suggest a large heterogeneity in the use of different nitrogen sources by the diverse range of mycobionts. Only a few researchers have investigated this functional diversity in the presence of an intact plant sink under controlled environmental conditions. In soil, several N sources are present in the same solution and it is not clear which sources particular fungi prefer. We also do not know whether some of these sources can be assimilated simultaneously and how uptake is regulated. It is remarkeble that mineral N uptake processes are still studied in non-mycorrhizal trees. The objective of this study are: 1. to onalyse the impact of functionally distinctive ECM fungi on the assimilation of various N sources in tree seedlings. The uptake capacity and kinetics of intact plants will be determined for different N sources, that will be applied separately or in combinations. Simultaneously, these experiments will be used to analyse the D15N patterns in plants and fungi at a low natural abundance of 15N. Complete N isotope budgets will be made and the effect of different N sources on the D15N patterns will be investigated. 2. to disentangle the metabolic pathways of N assimilation and transfer to the host plant we will perform a number of classical traces experiments with nitrogen sources labelled with 15N. In those cases, where amino acid are applied, additional information will be gained by using position specific isotope labels (amido-N or amino-N labelled glutamine) and by double labelling of amino acids with 13C and 15N. In this way, we will be able to distinguish different mechanisms that may be involved in the N transfer from fungus to plant. For the modelling of the N fluxes and N fractionation in the ecosystem, it is of great importance to determine the N pool(s) that participate in the transfer from fungus to plant. 3. We also will investigate the expression and regulation of plant and fungal genes that are involved in the N assimilation in mycorrhizal plants. Therefore, a new molecular technique (SAGE) will be used in order to analyse gene expression of microorganisms in the plant rhizosphere. The effect of various N sources on the expression of genes such as glutamine synthetase/glutamate synthase, nitrate reductase and other will be studied.
created:2011-12-14 14:18:59 UTC, source:web