Publications (8)22.3 Total impact
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Article: Does singlet oxygen activate cell death in Arabidopsis cell suspension cultures?: analysis of the early transcriptional defense responses to high light stress.
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ABSTRACT: Can Arabidopsis cell suspension cultures (ACSC) provide a useful working model to investigate genetically-controlled defense responses with signaling cascades starting in chloroplasts? In order to provide a convincing answer, we analyzed the early transcriptional profile of Arabidopsis cells at high light (HL). The results showed that ACSC respond to HL in a manner that resembles the singlet oxygen ((1)O(2))-mediated defense responses described for the conditional fluorescent (flu) mutant of Arabidopsis thaliana. The flu mutant is characterized by the accumulation of free protochlorophyllide (Pchlide) in plastids when put into darkness and the subsequent production of (1)O(2) when the light is on. In ACSC, (1)O(2) is produced in chloroplasts at HL when excess excitation energy flows into photosystem II (PSII). Other reactive oxygen species are also produced in ACSC at HL, but to a lesser extent. When the HL stress ceases, ACSC recovers the initial rate of oxygen evolution and cell growth continues. We can conclude that chloroplasts of ACSC are both photosynthetically active and capable of initiating (1)O(2)-mediated signaling cascades that activate a broad range of genetically-controlled defense responses. The upregulation of transcripts associated with the biosynthesis and signaling pathways of OPDA (12-oxophytodienoic acid) and ethylene (ET) suggests that the activated defense responses at HL are governed by these two hormones. In contrast to the flu mutant, the (1)O(2)-mediated defense responses were independent of the upregulation of EDS1 (enhanced disease susceptibility) required for the accumulation of salicylic acid (SA) and genetically-controlled cell death. Interestingly, a high correlation in transcriptional expression was also observed between ACSC at HL, and the aba1 and max4 mutants of Arabidopsis, characterized by defects in the biosynthesis pathways of abscisic acid (ABA) and strigolactones, respectively.Plant signaling & behavior 12/2011; 6(12):1937-42. -
Article: Trolox, a water-soluble analogue of α-tocopherol, photoprotects the surface-exposed regions of the photosystem II reaction center in vitro. Is this physiologically relevant?
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ABSTRACT: Can Trolox, a water-soluble analogue of α-tocopherol and a scavenger of singlet oxygen ((1)O(2)), provide photoprotection, under high irradiance, to the isolated photosystem II (PSII) reaction center (RC)? To answer the question, we studied the endogenous production of (1)O(2) in preparations of the five-chlorophyll PSII RC (RC5) containing only one β-carotene molecule. The temporal profile of (1)O(2) emission at 1270 nm photogenerated by RC5 in D(2)O followed the expected biexponential behavior, with a rise time, unaffected by Trolox, of 13 ± 1 μs and decay times of 54 ± 2 μs (without Trolox) and 38 ± 2 μs (in the presence of 25 μM Trolox). The ratio between the total (k(t)) and chemical (k(r)) bimolecular rate constants for the scavenging of (1)O(2) by Trolox in aqueous buffer was calculated to be ~1.3, with a k(t) of (2.4 ± 0.2) × 10(8) M(-1) s(-1) and a k(r) of (1.8 ± 0.2) × 10(8) M(-1) s(-1), indicating that most of the (1)O(2) photosensitized by methylene blue chemically reacts with Trolox in the assay buffer. The photoinduced oxygen consumption in the oxygen electrode, when RC5 and Trolox were mixed, revealed that Trolox was a better (1)O(2) scavenger than histidine and furfuryl alcohol at low concentrations (i.e., <1 mM). After its incorporation into detergent micelles in unbuffered solutions, Trolox was able to photoprotect the surface-exposed regions of the D1-D2 heterodimer, but not the RC5 pigments, which were oxidized, together with the membrane region of the protein matrix of the PSII RC, by (1)O(2). These results are discussed and compared with those of studies dealing with the physiological role of tocopherol molecules as a (1)O(2) scavenger in thylakoid membranes of photosynthetic organisms.Biochemistry 08/2011; 50(39):8291-301. · 3.42 Impact Factor -
Article: Early transcriptional defense responses in Arabidopsis cell suspension culture under high-light conditions.
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ABSTRACT: The early transcriptional defense responses and reactive oxygen species (ROS) production in Arabidopsis (Arabidopsis thaliana) cell suspension culture (ACSC), containing functional chloroplasts, were examined at high light (HL). The transcriptional analysis revealed that most of the ROS markers identified among the 449 transcripts with significant differential expression were transcripts specifically up-regulated by singlet oxygen ((1)O(2)). On the contrary, minimal correlation was established with transcripts specifically up-regulated by superoxide radical or hydrogen peroxide. The transcriptional analysis was supported by fluorescence microscopy experiments. The incubation of ACSC with the (1)O(2) sensor green reagent and 2',7'-dichlorofluorescein diacetate showed that the 30-min-HL-treated cultures emitted fluorescence that corresponded with the production of (1)O(2) but not of hydrogen peroxide. Furthermore, the in vivo photodamage of the D1 protein of photosystem II indicated that the photogeneration of (1)O(2) took place within the photosystem II reaction center. Functional enrichment analyses identified transcripts that are key components of the ROS signaling transduction pathway in plants as well as others encoding transcription factors that regulate both ROS scavenging and water deficit stress. A meta-analysis examining the transcriptional profiles of mutants and hormone treatments in Arabidopsis showed a high correlation between ACSC at HL and the fluorescent mutant family of Arabidopsis, a producer of (1)O(2) in plastids. Intriguingly, a high correlation was also observed with ABA deficient1 and more axillary growth4, two mutants with defects in the biosynthesis pathways of two key (apo)carotenoid-derived plant hormones (i.e. abscisic acid and strigolactones, respectively). ACSC has proven to be a valuable system for studying early transcriptional responses to HL stress.Plant physiology 04/2011; 156(3):1439-56. · 6.53 Impact Factor -
Article: Facile method for spectroscopic examination of radical ions of hydrophilic carotenoids.
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ABSTRACT: Hydrophilic carotenoids, unusual members of an intrinsically hydrophobic family, and their radical ions are important reactants. An all-optical method for generating singly charged radical ions of a hydrophilic carotenoid (Car) is described. It relies on photolyzing an aqueous mixture of Car and a photoionizable auxiliary solute (A), and making conditions conducive to the capture, by Car, of the hydrated electron (e(aq)(-)) or the positive hole in A(*)(+) or both. When A is Trolox (TOH), only e(aq)(-) can be captured, since TOH (*)(+) deprotonates too rapidly to be a hole donor; when A is Trolox methyl ether (TOMe), both Car(*)(-) and Car(*)(+) are formed, since TOMe (+) lives long enough to transfer its positive hole to Car; formation of Car(*)(-) is prevented under aerobic conditions.Physical Chemistry Chemical Physics 08/2009; 11(30):6401-5. · 3.57 Impact Factor -
Article: Peroxynitrite inhibits electron transport on the acceptor side of higher plant photosystem II.
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ABSTRACT: Peroxynitrite is a strong oxidant that has been proposed to form in chloroplasts. The interaction between peroxynitrite and photosystem II (PSII) has been investigated to determine whether this oxidant could be a hazard for PSII. Peroxynitrite is shown to inhibit oxygen evolution in PSII membranes in a dose-dependent manner. Analyses by PAM fluorimetry and EPR spectroscopy have demonstrated that the inhibition target of peroxynitrite is on the PSII acceptor side. In the presence of the herbicide DCMU, the chlorophyll (Chl) a fluorescence induction curve is inhibited by peroxynitrite, but the slow phase of the Chl a fluorescence decay does not change. EPR studies demonstrate that the Signal II(slow) and Signal II(fast) of peroxynitrite-treated Tris-washed PSII membranes are induced at room temperature, implying that the redox active tyrosines Y(Z) and Y(D) of PSII are not significantly nitrated. A featureless EPR signal with a g value of approximately 2.0043+/-0.0003 and a line width of 10+/-1G is induced under continuous illumination in the presence of peroxynitrite. This new EPR signal corresponds with the semireduced plastoquinone Q(A) in the absence of magnetic interaction with the non-heme Fe2+. We conclude that peroxynitrite impairs PSII electron transport in the Q(A)Fe2+ niche.Archives of Biochemistry and Biophysics 06/2008; 473(1):25-33. · 2.93 Impact Factor -
Article: Reaction center of photosystem II with no peripheral pigments in D2 allows secondary electron transfer in D1.
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ABSTRACT: A pigment-deficient reaction center of photosystem II (PSII)-with all the core pigments (two molecules of chlorophyll a and one of pheophytin a in each D protein) but with only one molecule each of peripheral chlorophyll a (Chlz) and beta-carotene (Car)-has been investigated by pump-probe spectroscopy. The data imply that Car and Chlz are both bound to D1. The absence of Car and Chlz in D2 allows the unprecedented observation of secondary electron transfer in D1 of PSII reaction centers at room temperature. The absorption band of the Car cation in D1 (Car(D1)(+*)) peaks around 910 nm (as against 990 nm for Car(D2)(+*)), and its positive hole is shared by ChlzD1, whereas Car(D2)(+*) can disappear by capturing an electron from ChlzD2.Biochemistry 01/2008; 46(51):15027-32. · 3.42 Impact Factor -
Article: Reaction center of photosystem II with no peripheral pigments in D2 allows secondary electron transfer in D1
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ABSTRACT: El articulo está disponible en: http://pubs.acs.org/cgi-bin/article.cgi/bichaw/2007/46/i51/pdf/bi701440z.pdf. La editorial no permite la inclusión del articulo en un repositorio institucional, pero está disponible on-line para quien tenga acceso a la suscripción. A pigment-deficient reaction center of photosystem II (PSII)-with all the core pigments (two molecules of chlorophyll a and one of pheophytin a in each D protein) but with only one molecule each of peripheral chlorophyll a (Chlz) and beta-carotene (Car)-has been investigated by pump-probe spectroscopy. The data imply that Car and Chlz are both bound to D1. The absence of Car and Chlz in D2 allows the unprecedented observation of secondary electron transfer in D1 of PSII reaction centers at room temperature. The absorption band of the Car cation in D1 (Car(D1)(+·)) peaks around 910 nm (as against 990 nm for Car(D2)(+·)), and its positive hole is shared by ChlzD1, whereas Car(D2)(+·) can disappear by capturing an electron from ChlzD2. This work was funded by the Research Council of Norway, the Spanish Ministry of Science and Education (Grant No. BFU2004-04914-C02-02), the Czech Ministry of Education, Youth and Sports (Grant Nos. MSM6007665808 and AV0Z50510513), and the Spanish and Czech bilateral collaboration program (Grant No. CZ2004001). Peer reviewed -
Article: Femtosecond laser disruption of filamentous cyanobacteria unveils dissimilar cellular stability between heterocysts and vegetative cells.
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ABSTRACT: Filamentous cyanobacteria develop heterocysts in response to deprivation for combined nitrogen under aerobic conditions. The most prominent structural change in heterocysts is the biosynthesis of an envelope that restricts gas permeability, providing an appropriate micro-oxic environment for N2 fixation inside. The additional thickness of the differentiated cells, when compared to vegetative cells, makes filamentous cyanobacteria an attractive biological system to investigate cellular response against femtosecond laser processing. By irradiating the cyanobacterial filaments with 120 fs, 795 nm, 1 kHz pulses focused through a 100x microscope objective with a numerical aperture of 0.85, we have determined that the pulse energy threshold for an apparent disruption of the cell wall of vegetative cells is 13 +/- 4 nJ per pulse. A further increase in the pulse energy to 43 +/- 13 nJ causes the complete removal of vegetative cells. In contrast, the pulse energy threshold has to be augmented about three-fold for heterocyst envelope disruption or two-fold for complete removal of heterocysts. We propose that the singular cross-linked structure of the glycolipid multilayer of the envelope, required to restrict gas permeability, accounts for the remarked difference in the ablation energy threshold between vegetative cells and heterocysts.Photochemistry and Photobiology 84(6):1576-82. · 2.41 Impact Factor
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Institutions
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2011
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Spanish National Research Council
Madrid, Madrid, Spain
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2008–2009
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Norwegian University of Science and Technology (NTNU)
- Department of Physics
Trondheim, Sor-Trondelag Fylke, Norway
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