Abstract
BackgroundReactive oxygen species (ROS) such as hydrogen peroxide and superoxide anions significantly accumulate during biotic and abiotic stress and cause oxidative damage and eventually cell death. There is accumulating evidence that ROS are also involved in regulating beneficial plant–microbe interactions, signal transduction and plant growth and development. Due to the relevance of ROS throughout the life cycle and for interaction with the multifactorial environment, the physiological phenotyping of the mechanisms controlling ROS homeostasis is of general importance.ResultsIn this study, we have developed a robust and resource-efficient experimental platform that allows the determination of the activities of the nine key ROS scavenging enzymes from a single extraction that integrates posttranscriptional and posttranslational regulations. The assays were optimized and adapted for a semi-high throughput 96-well assay format. In a case study, we have analyzed tobacco leaves challenged by pathogen infection, drought and salt stress. The three stress factors resulted in distinct activity signatures with differential temporal dynamics.ConclusionsThis experimental platform proved to be suitable to determine the antioxidant enzyme activity signature in different tissues of monocotyledonous and dicotyledonous model and crop plants. The universal enzymatic extraction procedure combined with the 96-well assay format demonstrated to be a simple, fast and semi-high throughput experimental platform for the precise and robust fingerprinting of nine key antioxidant enzymatic activities in plants.
Highlights
Reactive oxygen species (ROS) such as hydrogen peroxide and superoxide anions significantly accumulate during biotic and abiotic stress and cause oxidative damage and eventually cell death
As the plant extract containing the activity of interest is the limiting factor in each assay, the slope of the curves reveals the enzymatic activity in the samples
This is of great advantage in comparison to the cuvette assays, which in most cases measure endpoint activities and lack the inbuilt quality control
Summary
Reactive oxygen species (ROS) such as hydrogen peroxide and superoxide anions significantly accumulate during biotic and abiotic stress and cause oxidative damage and eventually cell death. There is accumulating evidence that ROS are involved in regulating beneficial plant–microbe interactions, signal transduction and plant growth and development. Reactive oxygen species (ROS) produced in plant cells have taken important roles in the biology of plants, Fimognari et al Plant Methods (2020) 16:42 environmental conditions is key for ensuring high reproductive fitness in general and high yield of crop plants in particular. ROS are not always acting as toxic molecules, but they have been shown to play important roles in the regulation of metabolism and development. Several studies showed that ROS can alter a number of metabolic pathways during organ development and response to stresses; the precise control of ROS in terms of abundance, cellular location and time is of paramount importance. Plant morphology is affected by the metabolism of ROS, where pollen formation and root hair growth are primary examples [12, 13]
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