Cold stress and freezing tolerance negatively affect the fitness of Arabidopsis thaliana accessions under field and controlled conditions.

MAIN CONCLUSION: Higher acclimated freezing tolerance improved winter survival, but reduced reproductive fitness of Arabidopsis thaliana accessions under field and controlled conditions. Low temperature is one of the most important abiotic factors influencing plant fitness and geographical distribution. In addition, cold stress is known to influence crop yield and is therefore of great economic importance. Increased freezing tolerance can be acquired by the process of cold acclimation, but this may be associated with a fitness cost. To assess the influence of cold stress on the fitness of plants, long-term field trials over 5 years were performed with six natural accessions of Arabidopsis thaliana ranging from very tolerant to very sensitive to freezing. Fitness parameters, as seed yield and 1000 seed mass, were measured and correlation analyses with temperature and freezing tolerance data performed. The results were compared with fitness parameters from controlled chamber experiments over 3 years with application of cold priming and triggering conditions. Winter survival and seed yield per plant were positively correlated with temperature in field experiments. In addition, winter survival and 1000 seed mass were correlated with the cold-acclimated freezing tolerance of the selected Arabidopsis accessions. The results provide strong evidence for a trade-off between higher freezing tolerance and reproductive fitness in A. thaliana, which might have ecological impacts in the context of global warming.

Response of Arabidopsis primary metabolism and circadian clock to low night temperature in a natural light environment.

Plants are exposed to varying irradiance and temperature within a day and from day to day. We previously investigated metabolism in a temperature-controlled greenhouse at the spring equinox on both a cloudy and a sunny day [daily light integral (DLI) of 7 mol m-2 d-1 and 12 mol m-2 d-1]. Diel metabolite profiles were largely captured in sinusoidal simulations at similar DLIs in controlled-environment chambers, except that amino acids were lower in natural light regimes. We now extend the DLI12 study by investigating metabolism in a natural light regime with variable temperature including cool nights. Starch was not completely turned over, anthocyanins and proline accumulated, and protein content rose. Instead of decreasing, amino acid content rose. Connectivity in central metabolism, which decreased in variable light, was not further weakened by variable temperature. We propose that diel metabolism operates better when light and temperature are co-varying. We also compared transcript abundance of 10 circadian clock genes in this temperature-variable regime with the temperature-controlled natural and sinusoidal light regimes. Despite temperature compensation, peak timing and abundance for dawn- and day-phased genes and GIGANTEA were slightly modified in the variable temperature treatment. This may delay dawn clock activity until the temperature rises enough to support rapid metabolism and photosynthesis.

Enzyme activity profiles during fruit development in tomato cultivars and Solanum pennellii.

Enzymes interact to generate metabolic networks. The activities of more than 22 enzymes from central metabolism were profiled during the development of fruit of the modern tomato cultivar Solanum lycopersicum 'M82' and its wild relative Solanum pennellii (LA0716). In S. pennellii, the mature fruit remains green and contains lower sugar and higher organic acid levels. These genotypes are the parents of a widely used near introgression line population. Enzymes were also profiled in a second cultivar, S. lycopersicum 'Moneymaker', for which data sets for the developmental changes of metabolites and transcripts are available. Whereas most enzyme activities declined during fruit development in the modern S. lycopersicum cultivars, they remained high or even increased in S. pennellii, especially enzymes required for organic acid synthesis. The enzyme profiles were sufficiently characteristic to allow stages of development and cultivars and the wild species to be distinguished by principal component analysis and clustering. Many enzymes showed coordinated changes during fruit development of a given genotype. Comparison of the correlation matrices revealed a large overlap between the two modern cultivars and considerable overlap with S. pennellii, indicating that despite the very different development responses, some basic modules are retained. Comparison of enzyme activity, metabolite profiles, and transcript profiles in S. lycopersicum 'Moneymaker' revealed remarkably little connectivity between the developmental changes of transcripts and enzymes and even less between enzymes and metabolites. We discuss the concept that the metabolite profile is an emergent property that is generated by complex network interactions.

Adjustment of growth and central metabolism to a mild but sustained nitrogen-limitation in Arabidopsis.

We have established a simple soil-based experimental system that allows a small and sustained restriction of growth of Arabidopsis by low nitrogen (N). Plants were grown in a large volume of a peat-vermiculite mix that contained very low levels of inorganic N. As a control, inorganic N was added in solid form to the peat-vermiculite mix, or plants were grown in conventional nutrient-rich solids. The low N growth regime led to a sustained 20% decrease of the relative growth rate over a period of 2 weeks, resulting in a two- to threefold decrease in biomass in 35- to 40-day-old plants. Plants in the low N regime contained lower levels of nitrate, lower nitrate reductase activity, lower levels of malate, fumarate and other organic acids and slightly higher levels of starch, as expected from published studies of N-limited plants. However, their rosette protein content was unaltered, and total and many individual amino acid levels increased compared with N-replete plants. This metabolic phenotype reveals that Arabidopsis responds adaptively to low N by decreasing the rate of growth, while maintaining the overall protein content, and maintaining or even increasing the levels of many amino acids.

GC-EI-TOF-MS analysis of in vivo carbon-partitioning into soluble metabolite pools of higher plants by monitoring isotope dilution after 13CO2 labelling.

The established GC-EI-TOF-MS method for the profiling of soluble polar metabolites from plant tissue was employed for the kinetic metabolic phenotyping of higher plants. Approximately 100 typical GC-EI-MS mass fragments of trimethylsilylated and methoxyaminated metabolite derivatives were structurally interpreted for mass isotopomer analysis, thus enabling the kinetic study of identified metabolites as well as the so-called functional group monitoring of yet non-identified metabolites. The monitoring of isotope dilution after (13)CO(2) labelling was optimized using Arabidopsis thaliana Col-0 or Oryza sativa IR57111 plants, which were maximally labelled with (13)C. Carbon isotope dilution was evaluated for short (2h) and long-term (3 days) kinetic measurements of metabolite pools in root and shoots. Both approaches were shown to enable the characterization of metabolite specific partitioning processes and kinetics. Simplifying data reduction schemes comprising calculation of (13)C-enrichment from mass isotopomer distributions and of initial (13)C-dilution rates were employed. Metabolites exhibited a highly diverse range of metabolite and organ specific half-life of (13)C-label in their respective pools ((13)C-half-life). This observation implied the setting of metabolite specific periods for optimal kinetic monitoring. A current experimental design for the kinetic metabolic phenotyping of higher plants is proposed.

Starch content and yield increase as a result of altering adenylate pools in transgenic plants.

Starch represents the most important carbohydrate used for food and feed purposes. With the aim of increasing starch content, we decided to modulate the adenylate pool by changing the activity of the plastidial adenylate kinase in transgenic potato plants. As a result, we observed a substantial increase in the level of adenylates and, most importantly, an increase in the level of starch to 60% above that found in wild-type plants. In addition, concentrations of several amino acids were increased by a factor of 2-4. These results are particularly striking because this genetic manipulation also results in an increased tuber yield. The modulation of the plastidial adenylate kinase activity in transgenic plants therefore represents a potentially very useful strategy for increasing formation of major storage compounds in heterotrophic tissues of higher plants.