We have quantitatively measured nitric oxide production in the leaves of Arabidopsis thaliana and Vicia faba by adapting ferrous dithiocarbamate spin tapping methods previously used in animal systems. Hydrophobic diethyldithiocarbamate complexes were used to measure NO interacting with membranes, and hydrophilic N-methyl-d-glucamine dithiocarbamate was used to measure NO released into the external solution. Both complexes were able to trap levels of NO, readily detectable by EPR spectroscopy. Basal rates of NO production (in the order of 1 nmol g– 1 h–1) agreed with previous studies. However, use of methodologies that corrected for the removal of free NO by endogenously produced superoxide resulted in a significant increase in trapped NO (up to 18 nmol g– 1 h–1). Basal NO production in leaves is therefore much higher than previously thought, but this is masked by significant superoxide production. The effects of nitrite (increased rate) and nitrate (decreased rate) are consistent with a role for nitrate reductase as the source of this basal NO production. However, rates under physiologically achievable nitrite concentrations never approach that reported following pathogen induction of plant nitric-oxide synthase. In Hibiscus rosa sinensis, the addition of exogenous nitrite generated sufficient NO such that EPR could be used to detect its production using endogenous spin traps (forming paramagnetic dinitrosyl iron complexes). Indeed the levels of this nitrosylated iron pool are sufficiently high that they may represent a method of maintaining bioavailable iron levels under conditions of iron starvation, thus explaining the previously observed role of NO in preventing chlorosis under these conditions.
Endogenous Superoxide Production and the Nitrite/Nitrate Ratio Control the Concentration of Bioavailable Free Nitric Oxide in Leaves*
A. Vanin,D. Svistunenko,V. Mikoyan,V. Serezhenkov,M. Fryer,N. Baker,C. Cooper
Published 2004 in Journal of Biological Chemistry
ABSTRACT
PUBLICATION RECORD
- Publication year
2004
- Venue
Journal of Biological Chemistry
- Publication date
2004-06-04
- Fields of study
Biology, Medicine, Chemistry, Environmental Science
- Identifiers
- External record
- Source metadata
Semantic Scholar, PubMed
CITATION MAP
EXTRACTION MAP
CLAIMS
CONCEPTS
- bioavailable iron
The pool of iron accessible for biological use, discussed here in relation to iron starvation and dinitrosyl iron complexes.
Aliases: available iron
- diethyldithiocarbamate complexes
Hydrophobic dithiocarbamate complexes used as spin traps to measure nitric oxide associated with membranes.
Aliases: DETC complexes
- dinitrosyl iron complexes
Paramagnetic iron-nitrosyl complexes formed from endogenous spin traps in Hibiscus rosa sinensis.
Aliases: DNICs
- endogenous superoxide production
Superoxide generated by the plant tissue itself that can react with free nitric oxide during the measurements.
Aliases: superoxide production
- epr spectroscopy
An electron paramagnetic resonance method used to detect the trapped nitric oxide signals in this assay.
Aliases: electron paramagnetic resonance spectroscopy, EPR
- hibiscus rosa sinensis
A plant species examined for nitric oxide generation from exogenous nitrite and for endogenous dinitrosyl iron complex formation.
- nitrate reductase
A plant enzyme proposed here as the source of basal nitric oxide production from nitrite.
Aliases: NR
- nitric oxide production
The generation of nitric oxide in plant leaves measured in this work as a rate of release or trapping.
Aliases: NO production, NO
- nitrite/nitrate ratio
The balance between nitrite and nitrate conditions used to probe how nitrogen oxides affect nitric oxide formation in leaves.
Aliases: nitrite, nitrate
- n-methyl-d-glucamine dithiocarbamate
A hydrophilic dithiocarbamate spin trap used to measure nitric oxide released into the external solution.
Aliases: MGD
REFERENCES
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