chlorophyll-a and diphenyleneiodonium

Diatoms are especially important microorganisms because they constitute the larger group of microalgae. To survive the constant variations of the light environment, diatoms have developed mechanisms aiming at the dissipation of excess energy, such as the xanthophyll cycle and the non-photochemical chlorophyll (Chl) fluorescence quenching. This contribution is dedicated to the relaxation of the latter process when the adverse conditions cease. An original nonlinear regression analysis of the relaxation of non-photochemical Chl fluorescence quenching, qN, in diatoms is presented. It was used to obtain experimental evidence for the existence of three time-resolved components in the diatom Phaeodactylum tricornutum: qNf, qNi and qNs. qNf (s time-scale) and qNs (h time-scale) are exponential in shape. By contrast, qNi (min time-scale) is of sigmoidal nature and is dominant among the three components. The application of metabolic inhibitors (dithiothreitol, ammonium chloride, cadmium and diphenyleneiodonium chloride) allowed the identification of the mechanisms on which each component mostly relies. qNi is linked to the relaxation of the ΔpH gradient and the reversal of the xanthophyll cycle. qNs quantifies the stage of photoinhibition caused by the high light exposure, qNf seems to reflect fast conformational changes within thylakoid membranes in the vicinity of the photosystem II complexes.

Topics: Ammonium Chloride; Cadmium; Chlorophyll; Diatoms; Dithiothreitol; Fluorescence; Kinetics; Light; Onium Compounds; Photosystem II Protein Complex; Regression Analysis; Thylakoids; Time Factors; Xanthophylls

Non-photochemical (dark) increases in chlorophyll a fluorescence yield associated with non-photochemical reduction of redox carriers (Fnpr) have been attributed to the reduction of plastoquinone (PQ) related to cyclic electron flow (CEF) around photosystem I. In vivo, this rise in fluorescence is associated with activity of the chloroplast plastoquinone reductase (plastid. plastoquinone oxidoreductase) complex. In contrast, this signal measured in isolated thylakoids has been attributed to the activity of the protein gradient regulation-5 (PGR5)/PGR5-like (PGRL1)-associated CEF pathway. Here, we report a systematic experimentation on the origin of Fnpr in isolated thylakoids. Addition of NADPH and ferredoxin to isolated spinach thylakoids resulted in the reduction of the PQ pool, but neither its kinetics nor its inhibitor sensitivities matched those of Fnpr. Notably, Fnpr was more rapid than PQ reduction, and completely insensitive to inhibitors of the PSII QB site and oxygen evolving complex as well as inhibitors of the cytochrome b6f complex. We thus conclude that Fnpr in isolated thylakoids is not a result of redox equilibrium with bulk PQ. Redox titrations and fluorescence emission spectra imply that Fnpr is dependent on the reduction of a low potential redox component (Em about − 340 mV) within photosystem II (PSII), and is likely related to earlier observations of low potential variants of QA within a subpopulation of PSII that is directly reducible by ferredoxin. The implications of these results for our understanding of CEF and other photosynthetic processes are discussed.

Topics: Antimycin A; Chlorophyll; Chloroplasts; Cytochrome b6f Complex; Diuron; Electron Transport; Enzyme Inhibitors; Ferredoxin-NADP Reductase; Ferredoxins; Fluorescence; Kinetics; NAD; Onium Compounds; Oxidation-Reduction; Photosynthesis; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Plant Proteins; Plastoquinone; Quinone Reductases; Spectrometry, Fluorescence; Spinacia oleracea; Thylakoids

Ethephon, an ethylene releasing compound, promoted leaf senescence, H2O2 elevation, and senescence-associated gene expression in sweet potato. It also affected the glutathione and ascorbate levels, which in turn perturbed H2O2 homeostasis. The decrease of reduced glutathione and the accumulation of dehydroascorbate correlated with leaf senescence and H2O2 elevation at 72h in ethephon-treated leaves. Exogenous application of reduced glutathione caused quicker and significant increase of its intracellular level and resulted in the attenuation of leaf senescence and H2O2 elevation. A small H2O2 peak produced within the first 4h after ethephon application was also eliminated by reduced glutathione. Diphenyleneiodonium (DPI), an NADPH oxidase inhibitor, delayed leaf senescence and H2O2 elevation at 72h, and its influence was effective only within the first 4h after ethephon treatment. Ethephon-induced senescence-associated gene expression was repressed by DPI and reduced glutathione at 72h in pretreated leaves. Leaves treated with l-buthionine sulfoximine, an endogenous glutathione synthetase inhibitor, did enhance senescence-associated gene expression, and the activation was strongly repressed by reduced glutathione. In conclusion, ethephon-mediated leaf senescence, H2O2 elevation and senescence-associated gene expression are all alleviated by reduced glutathione and NADPH oxidase inhibitor DPI. The speed and the amount of intracellular reduced glutathione accumulation influence its effectiveness of protection against ethephon-mediated effects. Reactive oxygen species generated from NADPH oxidase likely serves as an oxidative stress signal and participates in ethephon signaling. The possible roles of NADPH oxidase and reduced glutathione in the regulation of oxidative stress signal in ethephon are discussed.

Topics: Ascorbic Acid; Buthionine Sulfoximine; Cellular Senescence; Chlorophyll; Ethylenes; Gene Expression Regulation, Plant; Glutathione; Hydrogen Peroxide; Ipomoea batatas; NADPH Oxidases; Onium Compounds; Organophosphorus Compounds; Plant Leaves

Here, the link between UV-B stimulus and the abscisic acid (ABA)-induced nitricoxide (NO) synthesis pathway was studied in leaves of maize (Zea mays).The ABA concentration increased by 100% in UV-B irradiated leaves. Leaves of viviparous 14 (vp14), a mutant defective in ABA synthesis, were more sensitive to UV-B-induced damage than those of the wild type (wt). ABA supplementation attenuated UV-B-induced damage in both the wt and vp14. The hydrogen peroxide(H2O2) concentration increased in the irradiated wt, but changed only slightly in vp14. This increase was prevented by diphenylene iodonium (DPI), an inhibitor of NADPH oxidase (pNOX).NO was detected using the fluorophore 4,5-diamino-fluorescein diacetate(DAF-2DA). DAF-2DA fluorescence increased twofold in UV-B-irradiated wt leaves but not in vp14 leaves. H2O2 and NO production was restored in vp14 plants supplied with 100 μM ABA. Catalase, DPI and the NO synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME) partially blocked UV-B-induced NO accumulation, suggesting that H2O2 as well as NOS-like activity is required for a full plant response to UV-B. NO protects against UV-B-induced cell damage.Our results suggest that UV-B perception triggers an increase in ABA concentration,which activates pNOX and H2O2 generation, and that an NOS-like-dependent mechanism increases NO production to maintain cell homeostasis and attenuate UV-B-derived cell damage.

Topics: Abscisic Acid; Adaptation, Physiological; Chlorophyll; Helianthus; Hydrogen Peroxide; NADPH Oxidases; Nitric Oxide; Onium Compounds; Plant Leaves; Stress, Physiological; Ultraviolet Rays; Zea mays

Expression of the Chlamydomonas reinhardtii gsa gene encoding the chlorophyll biosynthetic enzyme glutamate 1-semialdehyde aminotransferase was previously shown to be induced by blue light. Possible blue light photoreceptors include flavins and carotenoids. Light induction of gsa was investigated in carotenoid-deficient mutant C. reinhardtii cells. Strain CC-2682 cells are sensitive to light, produce only small amounts of chlorophyll, and do not exhibit phototaxis. Solvent extracts show the absence of carotenoids and carotenoid precursors beyond phytoene in dark-grown mutant cells. Although apparently devoid of carotenoids, the cells did show light induction of gsa. The gsa transcript level was very low in dark-grown cells but increased significantly after 2 h of exposure to dim (1.5 x 10(-5) mol m(-2) s(-1)) green (480-585 nm) light. This light regime was previously determined not to injure these photosensitive cells and to fully induce gsa in wild-type cells. Exposure to this light did not cause the mutant cells to produce measurable carotenoids or to become phototactic. Growth of the mutant cells in the presence of exogenous beta-carotene or all-trans retinol restored phototaxis but did not affect the degree of gsa induction by light. The induction of gsa by light in the absence of carotenoids, and the fact that incorporation of physiologically usable carotenoids (as indicated by the restoration of phototaxis) did not affect the degree of light induction, indicate that the photoreceptor for light induction of gsa in C. reinhardtii is not a carotenoid. The flavin antagonist diphenyleneiodonium blocked light induction of gsa in both wild-type and mutant cells under conditions where respiration was not inhibited. These results suggest that the photoreceptor or a signal transduction effector for light induction of the C. reinhardtii gsa gene is a flavoprotein.

Topics: Animals; Carotenoids; Chlamydomonas reinhardtii; Chlorophyll; Enzyme Inhibitors; Gene Expression Regulation, Plant; Intramolecular Transferases; Light; Onium Compounds