No abstract

Nuclear DNA and other molecules in living systems are continuously exposed to endogenously generated oxygen species. Such species range from the unreactive superoxide radical (O2*-)the precursor of hydrogen peroxide (H2O2)to the highly reactive hydroxyl radical (*OH). Exogenous chemical and physical agents, such as ionizing radiation and the UVA component of solar light, can also oxidatively damage both the bases and the 2-deoxyribose moieties of cellular DNA. Over the last two decades, researchers have made major progress in understanding the oxidation degradation pathways of DNA that are most likely to occur from either oxidative metabolism or exposure to various exogenous agents. In the first part of this Account, we describe the mechanistic features of one-electron oxidation reactions of the guanine base in isolated DNA and related model compounds. These reactions illustrate the complexity of the various degradation pathways involved. Then, we briefly survey the analytical methods that can detect low amounts of oxidized bases and nucleosides in cells as they are formed. Recent data on the formation of oxidized guanine residues in cellular DNA following exposure to UVA light, ionizing radiation, and high-intensity UV pulses are also provided. We discuss these chemical reactions in the context of *OH radical, singlet oxygen, and two-quantum photoionization processes.

No abstract

No abstract

Carotenoids are considered to be the first line of defense of plants against singlet oxygen ( 1 O 2 ) toxicity because of their capacity to quench 1 O 2 as well as triplet chlorophylls through a physical mechanism involving transfer of excitation energy followed by thermal deactivation. Here, we show that leaf carotenoids are also able to quench 1 O 2 by a chemical mechanism involving their oxidation. In vitro oxidation of b-carotene, lutein, and zeaxanthin by 1 O 2 generated various aldehydes and endoperoxides. A search for those molecules in Arabidopsis (Arabidopsis thaliana) leaves revealed the presence of 1 O 2 -specific endoperoxides in low-light-grown plants, indicating chronic oxidation of carotenoids by 1 O 2 . b-Carotene endoperoxide, but not xanthophyll endoperoxide, rapidly accumulated during high-light stress, and this accumulation was correlated with the extent of photosystem (PS) II photoinhibition and the expression of various 1 O 2 marker genes. The selective accumulation of b-carotene endoperoxide points at the PSII reaction centers, rather than the PSII chlorophyll antennae, as a major site of 1 O 2 accumulation in plants under high-light stress. b-Carotene endoperoxide was found to have a relatively fast turnover, decaying in the dark with a half time of about 6 h. This carotenoid metabolite provides an early index of 1 O 2 production in leaves, the occurrence of which precedes the accumulation of fatty acid oxidation products.

The survey focuses on recent aspects of photochemical reactions to cellular DNA that are implicated through the predominant formation of mostly bipyrimidine photoproducts in deleterious effects of human exposure to sunlight. Recent developments in analytical methods have allowed accurate and quantitative measurements of the main DNA photoproducts in cells and human skin. Highly mutagenic CC and CT bipyrimidine photoproducts, including cyclobutane pyrimidine dimers and pyrimidine (6-4) pyrimidone photoproducts (6-4PPs) are generated in low yields with respect to TT and TC photoproducts. Another striking finding deals with the formation of Dewar valence isomers, the third class of bipyrimidine photoproducts that is accounted for by UVA-mediated isomerization of initially UVB generated 6-4PPs. Cyclobutadithymine (T< >T) has been unambiguously shown to be involved in the genotoxicity of UVA radiation. Thus, T< >T is formed in UVA-irradiated cellular DNA according to a direct excitation mechanism with a higher efficiency than oxidatively generated DNA damage that arises mostly through the Type II photosensitization mechanism. C<>C and C< >T are repaired at rates intermediate between those of T< >T and 6-4TT. Evidence has been also provided for the occurrence of photosensitized reactions mediated by exogenous agents that act either in an independent way or through photodynamic effects.

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.

A method involving high-performance liquid chromatography (HPLC) separation associated with tandem mass spectrometry (MS/MS) detection in the multiple-reaction monitoring mode was set up for the assessment of radiation-induced degradation products of DNA bases. This sensitive and specific assay is aimed at assessing six oxidized 2'-deoxyribonucleosides and two modified purine bases within both isolated and cellular DNA. For this purpose, stable isotopically labeled internal standards were prepared and used for isotope dilution mass spectrometry measurements. The latter method was validated through a comparison with two other assays, including HPLC associated with electrochemical detection and gas chromatography coupled to mass spectrometry. Using the specific and sensitive HPLC-MS/MS approach, 5,6-dihydroxy-5,6-dihydrothymidine, 5-hydroxy-2'-deoxyuridine, 5-(hydroxymethyl)-2'-deoxyuridine, 5-formyl-2'-deoxyuridine, 8-oxo-7,8-dihydro-2'-deoxyadenosine, 8-oxo-7,8-dihydro-2'-deoxyguanosine, 4, 6-diamino-5-formamidopyrimidine, and 2, 6-diamino-4-hydroxy-5-formamidopyrimidine were quantified within both isolated and cellular DNA upon exposure to gamma-radiation.