Unravelling the response of biological systems towards abiotic stress using systems approaches (transcriptome-metabolome network/ signalling metabolites)

Plants are exposed to a suite of both biotic and abiotic stresses which furthermore often occur in combination. The capacity to cope with these challenges is a crucial factor in growth and thus finally biomass. Light and temperature represent key parameters influencing photosynthesis and, consequently, potential yield of plants. We have performed a comprehensive study to approach the response on the systems level.

A. thaliana plants were exposed to eight environmental conditions differing in light intensity and/or temperature. The eight conditions were chosen to allow the analysis of situations with only one (light intensity or temperature) or two (i.e. light and temperature) parameter(s) simultaneously being changed, thus enabling the analysis of interactions between the treatments. — *A. thaliana* plants were exposed to eight environmental conditions differing in light intensity and/or temperature. The eight conditions were chosen to allow the analysis of situations with only one (light intensity or temperature) or two (i.e. light and temperature) parameter(s) simultaneously being changed, thus enabling the analysis of interactions between the treatments.

© Lothar Willmitzer

*A. thaliana* plants were exposed to eight environmental conditions differing in light intensity and/or temperature. The eight conditions were chosen to allow the analysis of situations with only one (light intensity or temperature) or two (i.e. light and temperature) parameter(s) simultaneously being changed, thus enabling the analysis of interactions between the treatments.

© Lothar Willmitzer

To this end A. thaliana plants were exposed to eight environmental conditions differing in light intensity and/or temperature and the response followed over 22 time points each resulting in a total of 177 conditions analyzed. The eight conditions were chosen to allow the analysis of situations with only one (light intensity or temperature) or two (i.e. light and temperature) parameter(s) simultaneously being changed, thus enabling the analysis of interactions between the treatments. Time points were chosen to cover both a linear and a logarithmic scale for each of the conditions. In order to have an adequate systems description, the response was followed on two different readout levels, the transcript and the metabolite level. Data have been analyzed using a variety of statistical and graph based tools.

Hierarchical cluster analysis shows individual conditions (defined by temperature and light) diverge into distinct trajectories at condition dependent times following a different kinetics on the metabolome compared to the transcriptome level.

GO overrepresentation analysis identifies a common response for all changed conditions on the transcriptome level during the early response phase (5 – 60 minutes). Two conditions (darkness at 21 and 32°C) display patterns reminiscent of the classical stress response.

Metabolic networks reconstructed via metabolite-metabolite correlations reveal extensive environment-specific re-wiring. Detailed analysis identifies novel and conditional connections between amino acids and intermediates of the TCA cycle. Parallel analysis of transcriptional changes strongly support a model where under conditions of missing photosynthesis and normal/high temperatures protein degradation occurs rapidly with the products of protein degradation, i.e. amino acids getting further catabolised to serve as main energy supply.

Network of potential regulatory interactions based on the analysis of the kinetic response of A. thaliana to various environmental treatments (cf. Abb. 3). — Network of potential regulatory interactions based on the analysis of the kinetic response of *A. thaliana* to various environmental treatments (cf. Abb. 3).

© Lothar Willmitzer

Network of potential regulatory interactions based on the analysis of the kinetic response of *A. thaliana* to various environmental treatments (cf. Abb. 3).

© Lothar Willmitzer

In parallel to the study of A. thaliana towards environmental perturbations we performed a study of system level stress adjustments by analyzing detailed time-resolved E. coli response to five different perturbations (cold, heat, oxidative stress, lactose diauxie, and stationary phase) on both the transcriptome and the metabolome level. Analysis reveals that the metabolite response is more specific as compared with the general response observed on the transcript level and is reflected by much higher specificity during the early stress adaptation phase and when comparing the stationary phase response to other perturbations.

Despite these differences, the response on both levels still follows the same dynamics and general strategy of energy conservation as reflected by rapid decrease of central carbon metabolism intermediates coinciding with down regulation of genes related to cell growth. Application of co-clustering and canonical correlation analysis on combined metabolite and transcript data identifies a number of significant condition-dependent associations between metabolites and transcripts. The results confirm and extend existing models about co-regulation between gene expression and metabolites demonstrating the power of integrated systems oriented analysis.

References

Hannah, M.A., Caldana, C., Steinhauser, D., Balbo, I., Fernie, A.R. & Willmitzer, L. Combined Transcript and Metabolite Profiling of Arabidopsis Grown under Widely Variant Growth Conditions Facilitates the Identification of Novel Metabolite-Mediated Regulation of Gene Expression. Plant Physiology 152, 2120-2129 (2010).

Caldana, C., Degenkolbe, T., Cuadros-Inostroza, A., Klie, S., Sulpice, R., Leisse, A., Steinhauser, D., Fernie, A.R., Willmitzer, L. & Hannah, M.A. High-density kinetic analysis of the metabolomic and transcriptomic response of Arabidopsis to eight environmental conditions. Plant Journal 67, 869-884 (2011).

Hubberten, H.M., Klie, S., Caldana, C., Degenkolbe, T., Willmitzer, L. & Hoefgen, R. Additional role of O-acetylserine as a sulfur status-independent regulator during plant growth. Plant Journal 70, 666-677 (2012).

Jozefczuk, S., Klie, S., Catchpole, G., Szymanski, J., Cuadros-Inostroza, A., Steinhauser, D., Selbig, J. & Willmitzer, L. Metabolomic and transcriptomic stress response of Escherichia coli. Molecular Systems Biology 6, - (2010).