Reactive oxygen species act as signaling molecules but can also directly provoke cellular damage by rapidly oxidizing cellular components, including lipids. We developed a high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry-based quantitative method that allowed us to discriminate between free radical (type I)-and singlet oxygen ( 1 O 2 ; type II)-mediated lipid peroxidation (LPO) signatures by using hydroxy fatty acids as specific reporters. Using this method, we observed that in nonphotosynthesizing Arabidopsis (Arabidopsis thaliana) tissues, nonenzymatic LPO was almost exclusively catalyzed by free radicals both under normal and oxidative stress conditions. However, in leaf tissues under optimal growth conditions, 1 O 2 was responsible for more than 80% of the nonenzymatic LPO. In Arabidopsis mutants favoring 1 O 2 production, photooxidative stress led to a dramatic increase of 1 O 2 (type II) LPO that preceded cell death. Furthermore, under all conditions and in mutants that favor the production of superoxide and hydrogen peroxide (two sources for type I LPO reactions), plant cell death was nevertheless always preceded by an increase in 1 O 2 -dependent (type II) LPO. Thus, besides triggering a genetic cell death program, as demonstrated previously with the Arabidopsis fluorescent mutant, 1 O 2 plays a major destructive role during the execution of reactive oxygen species-induced cell death in leaf tissues.Plant leaves capture sun-derived light energy to drive CO 2 fixation during photosynthesis. During this process, leaves need to cope with photooxidative stress when the balance between energy absorption and consumption is disturbed. Excess excitation energy in the photosystems (PSI and PSII) leads to the inhibition of photosynthesis via the production of various reactive oxygen species (ROS) at different spatial levels of the cell (Apel and Hirt, 2004;Asada, 2006;Van Breusegem and Dat, 2006). Both exposure to high light intensities and decreased CO 2 availability direct linear electron transfer toward the reduction of molecular oxygen, generating superoxide radicals (O 2 2. ) at PSI (the Mehler reaction). Superoxide dismutation generates hydrogen peroxide (H 2 O 2 ), which is detoxified in the chloroplast by ascorbate peroxidases. As such, this so-called water-water cycle participates in the dissipation of excess energy (Asada, 2006). Decreased CO 2 availability affects the first step in CO 2 fixation by shifting the carboxylation of Rubisco by the Rubisco carboxylase-oxygenase enzyme toward oxygenation, a process called photorespiration. This leads, through the action of glycolate oxidase, to peroxisomal H 2 O 2 production that is counteracted by catalases. Finally, when the intersystem electron carriers are overreduced, triplet excited P680 in the PSII reaction center as well as triplet chlorophylls in the light-harvesting antennae are produced, with the production of singlet oxygen ( 1 O 2 ) as a consequence (Krieger-Liszkay, 2005). In photosynthetic membranes, 1 O 2 ...

Singlet oxygen ( 1 O 2 ) is a reactive oxygen species that can function as a stress signal in plant leaves leading to programmed cell death. In microalgae, 1 O 2 -induced transcriptomic changes result in acclimation to 1 O 2 . Here, using a chlorophyll b-less Arabidopsis thaliana mutant (chlorina1 [ch1]), we show that this phenomenon can also occur in vascular plants. The ch1 mutant is highly photosensitive due to a selective increase in the release of 1 O 2 by photosystem II. Under photooxidative stress conditions, the gene expression profile of ch1 mutant leaves very much resembled the gene responses to 1 O 2 reported in the Arabidopsis mutant flu. Preexposure of ch1 plants to moderately elevated light intensities eliminated photooxidative damage without suppressing 1 O 2 formation, indicating acclimation to 1 O 2 . Substantial differences in gene expression were observed between acclimation and high-light stress: A number of transcription factors were selectively induced by acclimation, and contrasting effects were observed for the jasmonate pathway. Jasmonate biosynthesis was strongly induced in ch1 mutant plants under high-light stress and was noticeably repressed under acclimation conditions, suggesting the involvement of this hormone in 1 O 2 -induced cell death. This was confirmed by the decreased tolerance to photooxidative damage of jasmonatetreated ch1 plants and by the increased tolerance of the jasmonate-deficient mutant delayed-dehiscence2.

The xanthophyll cycle has a major role in protecting plants from photooxidative stress, although the mechanism of its action is unclear. Here, we have investigated Arabidopsis plants overexpressing a gene encoding ␤-carotene hydroxylase, containing nearly three times the amount of xanthophyll cycle carotenoids present in the wild-type. In high light at low temperature wild-type plants exhibited symptoms of severe oxidative stress: lipid peroxidation, chlorophyll bleaching, and photoinhibition. In transformed plants, which accumulate over twice as much zeaxanthin as the wild-type, these symptoms were significantly ameliorated. The capacity of non-photochemical quenching is not significantly different in transformed plants compared with wild-type and therefore an enhancement of this process cannot be the cause of the stress tolerant phenotype. Rather, it is concluded that it results from the antioxidant effect of zeaxanthin. 80 -90% of violaxanthin and zeaxanthin in wildtype and transformed plants was localized to an oligomeric LHCII fraction prepared from thylakoid membranes. The binding of these pigments in intact membranes was confirmed by resonance Raman spectroscopy. Based on the structural model of LHCII, we suggest that the protein/lipid interface is the active site for the antioxidant activity of zeaxanthin, which mediates stress tolerance by the protection of bound lipids.