Evaluation of P5CS and ProDH activity in Paulownia tomentosa (Steud.) as an indicator of oxidative changes induced by drought stress.

The aim of the study was to investigate changes in proline metabolism in seedlings of tree species during drought stress. One month old Paulownia tomentosa seedlings were exposed to moisture conditions at various levels (irrigation at 100, 75, 50 and 25% of field capacity), and then the material (leaves and roots) was collected three times at 10-day intervals. The activity of enzymes involved in proline metabolism was closely related to drought severity; however, proline content was not directly impacted. The activity of pyrroline-5-carboxylate synthetase (P5CS), which catalyzes proline biosynthesis, increased in response to hydrogen peroxide accumulation, which was correlated with soil moisture. In contrast, the activity of proline dehydrogenase (ProDH), which catalyzes proline catabolism, decreased. Compared to proline, the activity of these enzymes may be a more reliable biochemical marker of stress-induced oxidative changes. The content of proline is dependent on numerous additional factors, i.e., its degradation is an important alternative energy source. Moreover, we noted tissue-specific differences in this species, in which roots appeared to be proline biosynthesis sites and leaves appeared to be proline catabolism sites. Further research is needed to examine a broader view of proline metabolism as a cycle regulated by multiple mechanisms and differences between species.

High-throughput method for Oxygen Consumption Rate measurement (OCR) in plant mitochondria.

BACKGROUND: Conventional methods to measure oxygen consumption, such as Clark-type electrodes, have limitations such as requiring a large amount of starting material. Moreover, commercially available kits for high-throughput methods are usually optimized for animal cells and mitochondria. Here, we present a novel method to measure the oxygen consumption rate using a high-throughput assay in isolated mitochondria of European beech seeds. To perform the measurements, we adapted the Agilent Seahorse XF Cell Mito Stress Test Kit protocol for measurements on plant mitochondria. RESULTS: The optimized protocol for OCR measurement of mitochondria isolated from beech seeds allowed the observation of storage period-dependent gradual decreases in non-phosphorylating respiration, phosphorylating respiration and maximal FCCP-stimulated respiration. The longer the seeds were stored, the greater the impairment of respiratory function. CONCLUSIONS: Thanks to this method it is possible to minimize the amount of plant material and conduct research to obtain information on the respiratory condition and activity of plant mitochondria, including the efficiency of oxidative phosphorylation and the maximum oxidative capacity of the respiratory chain. We demonstrated that the improved protocol is suitable for study of plant material.

Proline concentrations in seedlings of woody plants change with drought stress duration and are mediated by seed characteristics: a meta-analysis.

Proline accumulation represents one of mechanisms used by plants to prevent the adverse consequences of water stress. The effects of increased proline levels in response to drought differ among species. Trees are exposed to the long-term effects of climate change. The reproductive success of species in a specific environment depends on the functional trait of tree seeds. We conducted a meta-analysis to evaluate the effects of drought stress on the proline concentrations in seedling leaf tissues of woody plant species and their relationships to drought duration, seed mass, seed category and coniferous/deciduous classification. Drought duration exhibited a nonlinear effect on proline accumulations. The drought effect on proline accumulations is greater for deciduous than for coniferous species and is higher for orthodox seed species than for recalcitrant. The seedlings of large-seeded species showed greater effect sizes than those of small-seeded species. Our results suggest that there is an optimum level at which proline accumulations under the influence of drought are the highest. A link between seed functional traits, as well as the coniferous/deciduous classification, and proline concentrations in tree seedlings during water stress were determined for the first time. Proline may help to identify high-quality seeds of trees used for reforestation.