Impact of root- and leaf-associated bacteria on metabolic priming of Arabidopsis for subsequent bacteria colonisation

Principal Investigator: Dr. habil. Joachim Kopka

PROJECT IS COMPLETED IN PHASE I.

The aim of this project is to understand the impact of Arabidopsis thaliana-associated bacteria on metabolic priming of plant primary carbohydrate metabolism that results in an altered stress response towards pathogenic bacteria. Specifically, we will investigate how a first bacterial contact primes metabolism such that the metabolic response is altered upon subsequent bacterial infection. We will apply beneficial rhizobacteria, and avirulent and virulent pathogenic leaf-associated bacteria of the genus Pseudomonas. So far, studies on priming of plant responses to pathogen infection did not yet investigate the impact of metabolic changes in the course of priming against pathogenic attack in detail; however, there is growing evidence that primary metabolism may play a major role during priming of defence against pathogens. To determine priming effects on the primary carbohydrate and energy metabolism, we will measure levels of sugars, sugar phosphates, amino acids and organic acids of naïve, primed and triggered plants. Since variation of metabolic activity is not necessarily reflected by changes of metabolite concentrations, we will also conduct metabolic flux experiments using ¹³C-substrate labelling. This approach allows monitoring the distribution of labelled carbon throughout the plant and to simultaneously determine the carbon-flux through selected metabolic pathways, thus providing detailed insight into changes in metabolic activity in the course of a first priming stimulus and a subsequent triggering bacterial stimulus. The introduction of a lag phase between the two subsequent stimuli will show how the metabolic pattern changes through time following the initial priming stimulus. Furthermore, we will evaluate activities of enzymes that are likely involved in regulating the observed changes in carbohydrate metabolism and that, therefore, are potential targets for metabolic priming. Preliminary experiments already revealed first evidence that invertase activity is affected upon priming, but other enzymes will be evaluated as well, depending on the results from the metabolic flux analyses. With plants altered in the activity of different invertases, we will investigate their role in the priming response. To consider the putative impact on metabolic regulation of bacteria-induced changes in phytohormone levels we will determine changes in plant hormone levels with available hormone reporter lines that contain a luciferase reporter system. Finally, we will determine how priming of the primary carbohydrate metabolism by the bacterial priming/triggering setup correlates with plant fitness parameters such as growth, development, reproductive success, and defence capacity.

References

• Cartieaux, F., Contesto, C., Gallou, A., Desbrosses, G., Kopka, J., Taconnat, L., Renou, J. P. and Touraine, B. 2008. Simultaneous interaction of Arabidopsis thaliana with Bradyrhizobium sp. strain ORS278 and Pseudomonas syringae pv. tomato DC3000 leads to complex transcriptome changes. Mol. Plant-Microbe Interact. 21: 244-259.
• 24*)    Hummel, J., Strehmel, N., Selbig, J., Walther, D. and Kopka, J. 2010. Decision tree supported substructure prediction of metabolites from GC-MS profiles. Metabolomics 6:322-333.
• Luedemann, A., Strassburg, K., Erban, A. and Kopka, J. 2008. TagFinder for the quantitative analysis of gas chromatography - mass spectrometry (GC-MS) based metabolite profiling experiments. Bioinformatics 24:732 -737.
• Strehmel, N., Hummel, J., Erban, A., Strassburg, K. and Kopka, J. 2008. Estimation of retention index thresholds for compound matching using routine gas chromatography-mass spectrometry based metabolite profiling experiments. J. Chromatogr. 871:182-190.
• van Dongen, J.T., Froehlich, A., Ramirez-Aguilar, S.J., Schauer, N., Fernie, A.R., Erban, A., Kopka, J., Clark, J., Langer, A. and Geigenberger, P. 2009. Transcript and metabolite profiling of the adaptive response to mild decreases in oxygen concentration in the roots of Arabidopsis plants. Ann. Bot. 103:269-280.
• Siahpoosha, M.R., Sanchez, D.H., Schlereth, A., Scofield, G.N., Furbank, R.T., van Dongen, J.T. and Kopka, J. 2011. Modification of OsSUT1 gene expression modulates the salt response of rice Oryza sativa cv. Taipei309. Plant Sci. DOI: 10.1016/j.plantsci.2011.01.001.
• Lisec, J., Schauer, N., Kopka,J., Willmitzer, L. and Fernie, A. R. 2006. Gas chromatography mass spectrometry-based metabolite profiling in plants. Nature Prot. 1: 387-396.