Drought affects carbon partitioning into volatile organic compound biosynthesis in Scots pine needles.

The effect of drought on the interplay of processes controlling carbon partitioning into plant primary and secondary metabolisms, such as respiratory CO2 release and volatile organic compound (VOC) biosynthesis, is not fully understood. To elucidate the effect of drought on the fate of cellular C sources into VOCs vs CO2 , we conducted tracer experiments with 13 CO2 and position-specific 13 C-labelled pyruvate, a key metabolite between primary and secondary metabolisms, in Scots pine seedlings. We determined the stable carbon isotope composition of leaf exchanged CO2 and VOC. Drought reduced the emission of the sesquiterpenes α-farnesene and ß-farnesene but did not affect 13 C-incorporation from 13 C-pyruvate. The labelling patterns suggest that farnesene biosynthesis partially depends on isopentenyl diphosphate crosstalk between chloroplasts and cytosol, and that drought inhibits this process. Contrary to sesquiterpenes, drought did not affect emission of isoprene, monoterpenes and some oxygenated compounds. During the day, pyruvate was used in the TCA cycle to a minor degree but was mainly consumed in pathways of secondary metabolism. Drought partly inhibited such pathways, while allocation into the TCA cycle increased. Drought caused a re-direction of pyruvate consuming pathways, which contributed to maintenance of isoprene and monoterpene production despite strongly inhibited photosynthesis. This underlines the importance of these volatiles for stress tolerance.

Physiological responses of date palm (Phoenix dactylifera) seedlings to seawater and flooding.

In their natural environment along coast lines, date palms are exposed to seawater inundation and, hence, combined stress by salinity and flooding. To elucidate the consequences of this combined stress on foliar gas exchange and metabolite abundances in leaves and roots, date palm seedlings were exposed to flooding with seawater and its major constituents under controlled conditions. Seawater flooding significantly reduced CO2 assimilation, transpiration and stomatal conductance, but did not affect isoprene emission. A similar effect was observed upon NaCl exposure. By contrast, flooding with distilled water or MgSO4 did not affect CO2 /H2 O gas exchange or stomatal conductance significantly, indicating that neither flooding itself, nor seawater sulfate, contributed greatly to stomatal closure. Seawater exposure increased Na and Cl contents in leaves and roots, but did not affect sulfate contents significantly. Metabolite analyses revealed reduced abundances of foliar compatible solutes, such as sugars and sugar alcohols, whereas nitrogen compounds accumulated in roots. Reduced transpiration upon seawater exposure may contribute to controlling the movement of toxic ions to leaves and, therefore, can be seen as a mechanism to cope with salinity. The present results indicate that date palm seedlings are tolerant towards seawater exposure to some extent, and highly tolerant to flooding.

Heat Waves Change Plant Carbon Allocation Among Primary and Secondary Metabolism Altering CO2 Assimilation, Respiration, and VOC Emissions.

Processes controlling plant carbon allocation among primary and secondary metabolism, i.e., carbon assimilation, respiration, and VOC synthesis are still poorly constrained, particularly regarding their response to stress. To investigate these processes, we simulated a 10-day 38°C heat wave, analysing real-time carbon allocation into primary and secondary metabolism in the Mediterranean shrub Halimium halimifolium L. We traced position-specific 13C-labeled pyruvate into daytime VOC and CO2 emissions and during light-dark transition. Net CO2 assimilation strongly declined under heat, due to three-fold higher respiration rates. Interestingly, day respiration also increased two-fold. Decarboxylation of the C1-atom of pyruvate was the main process driving daytime CO2 release, whereas the C2-moiety was not decarboxylated in the TCA cycle. Heat induced high emissions of methanol, methyl acetate, acetaldehyde as well as mono- and sesquiterpenes, particularly during the first two days. After 10-days of heat a substantial proportion of 13C-labeled pyruvate was allocated into de novo synthesis of VOCs. Thus, during extreme heat waves high respiratory losses and reduced assimilation can shift plants into a negative carbon balance. Still, plants enhanced their investment into de novo VOC synthesis despite associated metabolic CO2 losses. We conclude that heat stress re-directed the proportional flux of key metabolites into pathways of VOC biosynthesis most likely at the expense of reactions of plant primary metabolism, which might highlight their importance for stress protection.

Heat stress increases the use of cytosolic pyruvate for isoprene biosynthesis.

The increasing occurrence of heatwaves has intensified temperature stress on terrestrial vegetation. Here, we investigate how two contrasting isoprene-emitting tropical species, Ficus benjamina and Pachira aquatica, cope with heat stress and assess the role of internal plant carbon sources for isoprene biosynthesis in relation to thermotolerance. To our knowledge, this is the first study to report isoprene emissions from P. aquatica. We exposed plants to two levels of heat stress and determined the temperature response curves for isoprene and photosynthesis. To assess the use of internal C sources in isoprene biosynthesis, plants were fed with 13C position-labelled pyruvate. F. benjamina was more heat tolerant with higher constitutive isoprene emissions and stronger acclimation to higher temperatures than P. aquatica, which showed higher induced isoprene emissions at elevated temperatures. Under heat stress, both isoprene emissions and the proportion of cytosolic pyruvate allocated into isoprene synthesis increased. This represents a mechanism that P. aquatica, and to a lesser extent F. benjamina, has adopted as an immediate response to sudden increase in heat stress. However, in the long run under prolonged heat, the species with constitutive emissions (F. benjamina) was better adapted, indicating that plants that invest more carbon into protective emissions of biogenic volatile organic compounds tend to suffer less from heat stress.

Real-time carbon allocation into biogenic volatile organic compounds (BVOCs) and respiratory carbon dioxide (CO2) traced by PTR-TOF-MS, 13CO2 laser spectroscopy and 13C-pyruvate labelling.

Our understanding of biogenic volatile organic compound (BVOC) emissions improved substantially during the last years. Nevertheless, there are still large uncertainties of processes controlling plant carbon investment into BVOCs, of some biosynthetic pathways and their linkage to CO2 decarboxylation at central metabolic branching points. To shed more light on carbon partitioning during BVOC biosynthesis, we used an innovative approach combining δ13CO2 laser spectroscopy, high-sensitivity proton-transfer-reaction time-of-flight mass spectrometry and a multiple branch enclosure system in combination with position-specific 13C-metabolite labelling. Feeding experiments with position-specific 13C-labelled pyruvate, a central metabolite of BVOC synthesis, enabled online detection of carbon partitioning into 13C-BVOCs and respiratory 13CO2. Measurements of trace gas emissions of the Mediterranean shrub Halimium halimifolium revealed a broad range of emitted BVOCs. In general, [2-13C]-PYR was rapidly incorporated into emitted acetic acid, methyl acetate, toluene, cresol, trimethylbenzene, ethylphenol, monoterpenes and sesquiterpenes, indicating de novo BVOC biosynthesis of these compounds. In contrast, [1-13C]-pyruvate labelling substantially increased 13CO2 emissions in the light indicating C1-decarboxylation. Similar labelling patterns of methyl acetate and acetic acid suggested tightly connected biosynthetic pathways and, furthermore, there were hints of possible biosynthesis of benzenoids via the MEP-pathway. Overall, substantial CO2 emission from metabolic branching points during de novo BVOC biosynthesis indicated that decarboxylation of [1-13C]-pyruvate, as a non-mitochondrial source of CO2, seems to contribute considerably to daytime CO2 release from leaves. Our approach, combining synchronised BVOC and CO2 measurements in combination with position-specific labelling opens the door for real-time analysis tracing metabolic pathways and carbon turnover under different environmental conditions, which may enhance our understanding of regulatory mechanisms in plant carbon metabolism and BVOC biosynthesis.

The carnivorous Venus flytrap uses prey-derived amino acid carbon to fuel respiration.

The present study was performed to elucidate the fate of carbon (C) and nitrogen (N) derived from protein of prey caught by carnivorous Dionaea muscipula. For this, traps were fed 13 C/15 N-glutamine (Gln). The release of 13 CO2 was continuously monitored by isotope ratio infrared spectrometry. After 46 h, the allocation of C and N label into different organs was determined and tissues were subjected to metabolome, proteome and transcriptome analyses. Nitrogen of Gln fed was already separated from its C skeleton in the decomposing fluid secreted by the traps. Most of the Gln-C and Gln-N recovered inside plants were localized in fed traps. Among nonfed organs, traps were a stronger sink for Gln-C compared to Gln-N, and roots were a stronger sink for Gln-N compared to Gln-C. A significant amount of the Gln-C was respired as indicated by 13 C-CO2 emission, enhanced levels of metabolites of respiratory Gln degradation and increased abundance of proteins of respiratory processes. Transcription analyses revealed constitutive expression of enzymes involved in Gln metabolism in traps. It appears that prey not only provides building blocks of cellular constituents of carnivorous Dionaea muscipula, but also is used for energy generation by respiratory amino acid degradation.