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See detailFunctional analysis of hydrogen photoproduction in respiratory-deficient mutants of Chlamydomonas reinhardtii
Lecler, Renaud ULg; Godaux, Damien ULg; Vigeolas, Hélène ULg et al

in International Journal of Hydrogen Energy (2011), 36

In this paper, mitochondrial mutants of Chlamydomonas reinhardtii defective for respiratory complex I (NADH:ubiquinone oxidoreductase), complex III (ubiquinol cytochrome c oxidoreductase) and both ... [more ▼]

In this paper, mitochondrial mutants of Chlamydomonas reinhardtii defective for respiratory complex I (NADH:ubiquinone oxidoreductase), complex III (ubiquinol cytochrome c oxidoreductase) and both complexes I and III were analyzed for H2 photoproduction. Several parameters were followed during the S-deficiency stage and the anaerobic stage leading to H2 photoproduction. At the early aerobic S-deficiency stage, starch and neutral lipids accumulated in all strains but their amount was significantly decreased in mutants compared to wild type. During the H2 photoproduction process, whereas starch content strongly decreased in all strains, neutral lipid amount remained nearly unchanged, suggesting that starch degraded by glycolysis is the preferential substrate for energy production during anaerobiosis. The mutants displayed a decrease in H2 photoproduction correlating to the number of active mitochondrial proton-pumping sites lost in the strains. Our results thus highlight the critical role of oxidative phosphorylation during the first (aerobic) stage of S-starvation when carbon resources are accumulated. [less ▲]

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See detailLa réponse photosynthétique d'une algue verte à la carence en soufre
de Marchin, Thomas ULg; Ghysels, Bart ULg; Franck, Fabrice ULg

Book published by Editions universitaires europeennes (2010)

Chlamydomonas reinhardtii possède la capacité de produire de l'hydrogène à la lumière en absence d'oxygène. Cette condition peut être obtenue en cultivant les algues dans un milieu carencé en soufre. La ... [more ▼]

Chlamydomonas reinhardtii possède la capacité de produire de l'hydrogène à la lumière en absence d'oxygène. Cette condition peut être obtenue en cultivant les algues dans un milieu carencé en soufre. La carence en soufre entraîne une forte diminution de l'activité du photosystème II tout en maintenant une respiration élevée, ce qui provoque un passage de cultures fermées en anoxie et induit la production d'hydrogène. Dans cette étude, nous avons caractérisé la réponse photosynthétique à la carence en soufre chez la souche sauvage et la souche déficiente en oxydase alternative mictochondriale (AOX) dans des milieux contenant de l'ammonium ou du nitrate comme source d'azote. L'AOX, inductible par le nitrate, fait partie de la chaîne de transport d'électrons mitochondriale et catalyse l'oxydation de l'ubiquinol en transférant directement ses électrons à l'oxygène. Ainsi l'AOX entre en compétition avec le complexe III et est impliquée dans une voie de dissipation du pouvoir réducteur en excès. [less ▲]

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See detailAlternative photosynthetic electron pathways in different clades of Symbiodinium: the Mehler reaction
Roberty, Stéphane ULg; Poulicek, Mathieu ULg; Franck, Fabrice ULg

Poster (2010, December)

The high productivity of coral reef ecosystems is largely attributed to the mutualistic symbiosis between reef-building corals and their intracellular dinoflagellate in the genus Symbiodinium commonly ... [more ▼]

The high productivity of coral reef ecosystems is largely attributed to the mutualistic symbiosis between reef-building corals and their intracellular dinoflagellate in the genus Symbiodinium commonly referred to as zooxanthellae. These photosynthetic algae translocate a majority of their photosynthetically fixed carbon to the host and contribute greatly to their metabolic needs (Muscatine, 1990) and the calcification process (Gattuso, 1999). In the natural environment the holobiont have to cope with significant daily variations in light intensities that sometimes exceed Symbiodinium photosynthetic capacity. Fortunately, photosynthetic organisms possess regulatory features that help to ensure that high light intensities can be endured without the accumulation of photodamage. Thus, the regulation of photosynthesis can be viewed as a dynamic balance between photosynthetic efficiency (photochemical quenching) and photoprotection processes (i.e. non-photochemical quenching). Among them, the role of O2 as an alternative electron acceptor within the chloroplast could play a critical role (Ort & Baker, 2002). Under particular environmental conditions when sinks for photosynthetic electrons are scarce, the direct reduction of oxygen by the PSI could sustain significant levels of photosynthetic electron flux by initiating the ΔpH formation and of NPQ, regulating the ratio of ATP/NADPH to match the requirements of carbon reduction. However, this process leads to the formation of reactive oxygen species that are rapidly detoxified by superoxide dismutase and ascorbate peroxidase. An additional electron flux associated with this oxygen pathway is directed to the reduction of monodehydroascorbate (MDA), which is generated as a result of peroxide reduction by ascorbate (Asada, 2000). The present study aimed to highlight the existence of alternative photosynthetic electron pathways and more especially the Mehler ascorbate peroxidase pathway in different clades of Symbiodinium, cultivated at low and high light intensities. -Muscatine L (1990) The role of symbiotic algae in carbon and energy flux in reef corals. In: Dubinsky Z(ed) Ecosystems of the world: coral reefs. Elsevier, Amsterdam, pp 1-9. -Gattuso JP, Allemand D and M Frankignoulle (1999) Photosynthesis and calcification at cellular, organismal and community levels in coral reefs: A review on interactions and control by carbonate chemistry. American Zoologist 39(1): 160-183. -Ort, D. R. and N. R. Baker (2002). A photoprotective role for O2 as an alternative electron sink in photosynthesis? Current Opinion in Plant Biology 5(3): 193-198. -Asada, K. (2000) The water-water cycle as alternative photon and electron sinks. Philosophical Transactions of the Royal Society B-Biological Sciences 355(1402): 1419–1431. [less ▲]

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See detail(Functionnal) analysis of hydrogen production in Chlamydomonas reinhardtii mitochondrial mutants
Lecler, Renaud ULg; Godaux, Damien ULg; Hamilton, Christopher ULg et al

Poster (2010, June 27)

Mitochondrial Chlamydomonas mutants for respiratory complexes present a decreased dark respiration and apparent yield of photosynthetic linear electron flow. They accumulate reducing power such as NAD(P)H ... [more ▼]

Mitochondrial Chlamydomonas mutants for respiratory complexes present a decreased dark respiration and apparent yield of photosynthetic linear electron flow. They accumulate reducing power such as NAD(P)H and show lower levels of ATP. Under restrictive conditions, like sulfur depletion and anoxia, Chlamydomonas is able to produce hydrogen towards the activation of a chloroplatic O2-sensitive Fe-hydrogenase which catalyses the reduction of electrons to H2. In this study we used an adapted Melis protocol to analyse hydrogen evolution of mitochondrial mutants. For this aim a simple-flask system was built with gaz collecting tubes. A parallel flask was used for GC analyses. [less ▲]

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See detailEffect of pH on glucose and starch fermentation in batch and sequenced-batch mode with a recently isolated strain of hydrogen-producing Clostridium butyricum CWBI1009
Masset, Julien ULg; Hiligsmann, Serge ULg; Hamilton, Christopher ULg et al

in International Journal of Hydrogen Energy (2010), 35(8), 3371-3378

This paper reports investigations carried out to determine the optimum culture conditions for the production of hydrogen with a recently isolated strain Clostridium butyricum CWBI1009. The production ... [more ▼]

This paper reports investigations carried out to determine the optimum culture conditions for the production of hydrogen with a recently isolated strain Clostridium butyricum CWBI1009. The production rates and yields were investigated at 30 °C in a 2.3 l bioreactor operated in batch and sequenced-batch mode using glucose and starch as substrates. In order to study the precise effect of a stable pH on hydrogen production, and the metabolite pathway involved, cultures were conducted with pH controlled at different levels ranging from 4.7 to 7.3 (maximum range of 0.15 pH unit around the pH level). For glucose the maximum yield (1.7 mol H2 mol-1 glucose) was measured when the pH was maintained at 5.2. The acetate and butyrate yields were 0.35 mol acetate mol-1 glucose and 0.6 mol butyrate mol-1 glucose. For starch a maximum yield of 2.0 mol H2 mol-1 hexose, and a maximum production rate of 15 mol H2 mol-1 hexose h-1 were obtained at pH 5.6 when the acetate and butyrate yields were 0.47 mol acetate mol-1 hexose and 0.67 mol butyrate mol-1 hexose. [less ▲]

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See detailLa production photosynthétique d'oxygène
Franck, Fabrice ULg

Conference given outside the academic context (2010)

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See detailProteomic and functional characterization of a Chlamydomonas reinhardtii mutant lacking the mitochondrial alternative oxidase 1
Mathy, Grégory ULg; Cardol, Pierre ULg; Dinant, Monique et al

in Journal of Proteome Research (2010), 9

In the present work we have isolated by RNA interference and characterized at the functional and the proteomic levels a Chlamydomonas reinhardtii strain devoid of the mitochondrial alternative oxidase ... [more ▼]

In the present work we have isolated by RNA interference and characterized at the functional and the proteomic levels a Chlamydomonas reinhardtii strain devoid of the mitochondrial alternative oxidase (AOX). The AOX-deficient strain displays a doubling of the cell volume and biomass without any alteration of the generation time, a significantly higher ROS production, no change in total respiration rate, and a slight decrease of the photosynthesis efficiency. In order to identify the molecular adaptation underlying these phenotypical effects, we carried out a comparative proteomic study at the level of the mitochondrial and cellular soluble proteomes. Our results indicate a strong up-regulation of the ROS scavenging systems and important modifications of proteins involved in the primary metabolism, namely an increase of enzymes involved in anabolic pathways and a concomitant general down-regulation of enzymes of the main catabolic pathways. [less ▲]

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See detailPlasticity of the mitoproteome to nitrogen sources (nitrate and ammonium) in Chlamydomonas reinhardtii: the logic of Aox1 gene localization
Gérin, Stéphanie ULg; Mathy, Grégory ULg; Blomme, Arnaud ULg et al

in Biochimica et Biophysica Acta-Bioenergetics (2010), 1797

Nitrate and ammonium constitute primary inorganic nitrogen sources that can be incorporated into carbon skeletons in photosynthetic eukaryotes. In Chlamydomonas, previous studies and the present one ... [more ▼]

Nitrate and ammonium constitute primary inorganic nitrogen sources that can be incorporated into carbon skeletons in photosynthetic eukaryotes. In Chlamydomonas, previous studies and the present one showed that the mitochondrial AOX is up-regulated in nitrate-grown cells in comparison with ammonium-grown cells. In this work, we have performed a comparative proteomic analysis of the soluble mitochondrial proteome of Chlamydomonas cells growth either on nitrate or ammonium. Our results highlight important proteomics modifications mostly related to primary metabolism in cells grown on nitrate. We could note an up-regulation of some TCA cycle enzymes and a down-regulation of cytochrome c1 together with an up-regulation of l-arginine and purine catabolism enzymes and of ROS scavenging systems. Hence, in nitrate-grown cells, AOX may play a dual role: (1) lowering the ubiquinone pool reduction level and (2) permitting the export of mitochondrial reducing power under the form of malate for nitrate and nitrite reduction. This role of AOX in the mitochondrial plasticity makes logical the localization of Aox1 in a nitrate assimilation gene cluster. [less ▲]

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See detailThe onset of NPQ and Deltamu(H)+ upon illumination of tobacco plants studied through the influence of mitochondrial electron transport.
Cardol, Pierre ULg; De Paepe, Rosine; Franck, Fabrice ULg et al

in Biochimica et Biophysica Acta (2010), 1797(2), 177-88

The relationship between the development of photoprotective mechanisms (non-photochemical quenching, NPQ), the generation of the electrochemical proton gradient in the chloroplast and the capacity to ... [more ▼]

The relationship between the development of photoprotective mechanisms (non-photochemical quenching, NPQ), the generation of the electrochemical proton gradient in the chloroplast and the capacity to assimilate CO(2) was studied in tobacco dark-adapted leaves at the onset of illumination with low light. These conditions induce the generation of a transient NPQ, which relaxes in the light in parallel with the activation of the Calvin cycle. Wild-type plants were compared with a CMSII mitochondrial mutant, which lacks the respiratory complex I and shows a delayed activation of photosynthesis. In the mutant, a slower onset of photosynthesis was mirrored by a decreased capacity to develop NPQ. This correlates with a reduced efficiency to reroute electrons at the PSI reducing side towards cyclic electron flow around PSI and/or other alternative acceptor pools, and with a smaller ability to generate a proton motive force in the light. Altogether, these data illustrate the tight relationship existing between the capacity to evacuate excess electrons accumulated in the intersystem carriers and the capacity to dissipate excess photons during a dark to light transition. These data also underline the essential role of respiration in modulating the photoprotective response in dark-adapted leaves, by poising the cellular redox state. [less ▲]

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See detailEukaryotic algae: where lies the diversity of oxygenic photosynthesis.
Cardol, Pierre ULg; Franck, Fabrice ULg

in Photosynthesis Research (2010), 106(1-2), 1-2

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See detailHydrogen photo-evolution upon S deprivation stepwise: An illustration of microalgal photosynthetic and metabolic flexibility and a step stone for future biotechnological methods of renewable H2 production
Ghysels, Bart ULg; Franck, Fabrice ULg

in Photosynthesis Research (2010), 106

The metabolic flexibility of some photosynthetic microalgae enables them to survive periods of anaerobiosis in the light by developing a particular photofermentative metabolism. The latter entails ... [more ▼]

The metabolic flexibility of some photosynthetic microalgae enables them to survive periods of anaerobiosis in the light by developing a particular photofermentative metabolism. The latter entails compounds of the photosynthetic electron transfer chain and an oxygen-sensitive hydrogenase in order to reoxidise reducing equivalents and to generate ATP for maintaining basal metabolic function. This pathway results in the photo-evolution of hydrogen gas by the algae. A decade ago Melis and coworkers managed to reproduce such a condition in a laboratory context by depletion of sulfur in the algal culture media, making the photo-evolution by the algae sustainable for several days (Melis et al. 2000). This observation boosted research in algal H2 evolution. A feature, which due to its transient nature was long time considered as a curiosity of algal photosynthesis suddenly became a phenomenon with biotechnological potential. Although the Melis procedure has not been developed into a biotechnological process of renewable H2 generation so far, it has been a useful tool for studying microalgal metabolic and photosynthetic flexibility and a possible step stone for future H2 production procedures. Ten years later most of the critical steps and limitations of H2 production by this protocol have been studied from different angles particularly with the model organism C. reinhardtii, by introducing various changes in culture conditions and making use of mutants issued from different screens or by reverse genomic approaches. A synthesis of these observations with the most important conclusions driven from recent studies will be presented in this review. [less ▲]

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See detailPhotoacclimation responses of a symbiotic sea anemone reveal an important host cellular plasticity
Roberty, Stéphane ULg; Fransolet, David ULg; Ladrière, Ophélie ULg et al

Poster (2010)

The high productivity of coral reef ecosystems is largely attributed to the mutualistic symbiosis between reef-building corals and their intracellular dinoflagellate in the genus Symbiodinium commonly ... [more ▼]

The high productivity of coral reef ecosystems is largely attributed to the mutualistic symbiosis between reef-building corals and their intracellular dinoflagellate in the genus Symbiodinium commonly referred to as zooxanthellae. These photosynthetic algae translocate a majority of their photosynthetically fixed carbon to the host and contribute to their metabolic needs and the calcification process. <i>Symbiodinium</i> must maintain a balance between the energy derived from the light reactions in the chloroplast and the amount of energy used during dark reactions and other metabolic processes. Nevertheless, in the natural environment the holobiont have to cope with daily and seasonal changes in light intensity, upsetting that balance and creating a stress that induces a physiological response (photoacclimation) to optimize growth rates. After a ten day exposition to high and very low light intensity, morphological and photophysiological analysis conducted on the symbiotic sea anemone, Anemonia manjano, reveal significant modifications of the host tissues ultrastructure and the Symbiodinium metabolic processes (photosynthesis, respiration). Those results highlight particularly important gastrodermal and ectodermal plasticity in which symbiotic cnidarians acclimate to the Symbiodinium physiological status (mainly photosynthesis) by varying the density of particular cellular types (e.g.: cnidocytes, gastrodermal cells) contained in their tissues. [less ▲]

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See detailVitamin B6 deficient plants display increased sensitivity to high light and photo-oxidative stress
Havaux, Michel; Ksas, Brigitte; Szewczyk, Agnieszka et al

in BMC Plant Biology (2009), 9

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See detailImpaired respiration discloses the physiological significance of state transitions in Chlamydomonas.
Cardol, Pierre ULg; Alric, Jean; Girard-Bascou, Jacqueline et al

in Proceedings of the National Academy of Sciences of the United States of America (2009), 106(37), 15979-84

State transitions correspond to a major regulation process for photosynthesis, whereby chlorophyll protein complexes responsible for light harvesting migrate between photosystem II and photosystem I in ... [more ▼]

State transitions correspond to a major regulation process for photosynthesis, whereby chlorophyll protein complexes responsible for light harvesting migrate between photosystem II and photosystem I in response to changes in the redox poise of the intersystem electron carriers. Here we disclose their physiological significance in Chlamydomonas reinhardtii using a genetic approach. Using single and double mutants defective for state transitions and/or mitochondrial respiration, we show that photosynthetic growth, and therefore biomass production, critically depends on state transitions in respiratory-defective conditions. When extra ATP cannot be provided by respiration, enhanced photosystem I turnover elicited by transition to state 2 is required for photosynthetic activity. Concomitant impairment of state transitions and respiration decreases the overall yield of photosynthesis, ultimately leading to reduced fitness. We thus provide experimental evidence that the combined energetic contributions of state transitions and respiration are required for efficient carbon assimilation in this alga. [less ▲]

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See detailS13.45 Chlamydomonas reinhardtii mitoproteome adaptation in response to inactivation of the energy-dissipating alternative oxidase 1 by RNA interference
Cloes, Marie ULg; Mathy, Grégory ULg; Cardol, Pierre ULg et al

in Biochimica et Biophysica Acta (BBA) - Bioenergetics, Volume 1777, Supplement 1, 19 July 2008, Page S99 (2008, July 18), 1777(Supplement 1), 99

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See detailImportance of the alternative pathway of respiration for avoidance of ROS production and for optimisation of photosynthesis in Chlamydomonas
Franck, Fabrice ULg; Dinant, M.; Cardol, Pierre ULg et al

Conference (2008, June)

The physiological function of the alternative pathway of respiration has been investigated by analysing two RNAi C.reinhardtii lines deprived of alternative oxidase protein (AOX1). Compared to wild-type ... [more ▼]

The physiological function of the alternative pathway of respiration has been investigated by analysing two RNAi C.reinhardtii lines deprived of alternative oxidase protein (AOX1). Compared to wild-type, AOX1- lines exhibited modified growth curves and reduced maximal cell density. These differences were more pronounced at high irradiance and in nitrate-containing medium (TAP NO3) rather than in ammonium-containing medium (TAP NH4). Although the alternative pathway was inactive, respiration was not significantly altered in transgenics. Light-saturation curves of O2-evolution were only slightly modified. However, non-photochemical quenching of fluorescence (NPQ) was strongly reduced. Further analysis showed that AOX1- transgenics present a reduced ability to promote the change in energy distribution between photosystems, known as state transition. This effect, which explains low NPQ in the light, was most pronounced in high-light cells cultivated in TAP NO3 medium. Moreover, AOX1- transgenics exhibited higher levels of intracellular peroxides, which suggests that inhibition of state transition might result from higher ROS production. In support of this hypothesis, addition of millimolar-range concentrations of H2O2 to wild-type inhibited the state transition promoted by the reduction of the plastoquinone pool in darkness. [less ▲]

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See detailA type II NAD(P)H dehydrogenase mediates light-independent plastoquinone reduction in the chloroplast of Chlamydomonas
Jans, Frédéric ULg; Mignolet, Emmanuel ULg; Houyoux, Pierre-Alain et al

in Proceedings of the National Academy of Sciences of the United States of America (2008), 105(51), 20546-51

In photosynthetic eukaryotes, nonphotochemical plastoquinone (PQ) reduction is important for the regulation of photosynthetic electron flow. In green microalgae where this process has been demonstrated ... [more ▼]

In photosynthetic eukaryotes, nonphotochemical plastoquinone (PQ) reduction is important for the regulation of photosynthetic electron flow. In green microalgae where this process has been demonstrated, the chloroplastic enzyme that catalyses nonphotochemical PQ reduction has not been identified yet. Here, we show by an RNA interference (RNAi) approach that the NDA2 gene, belonging to a type II NAD(P)H dehydrogenases family in the green microalga Chlamydomonas reinhardtii, encodes a chloroplastic dehydrogenase that functions to reduce PQ nonphotochemically in this alga. Using a specific antibody, we show that the Nda2 protein is localized in chloroplasts of wild-type cells and is absent in two Nda2-RNAi cell lines. In both mutant cell lines, nonphotochemical PQ reduction is severely affected, as indicated by altered chlorophyll fluorescence transients after saturating illumination. Compared with wild type, change in light excitation distribution between photosystems ('state transition') upon inhibition of mitochondrial electron transport is strongly impaired in transformed cells because of inefficient PQ reduction. Furthermore, the amount of hydrogen produced by Nda2-RNAi cells under sulfur deprivation is substantially decreased compared with wild type, which supports previous assumptions that endogenous substrates serve as source of electrons for hydrogen formation. These results demonstrate the importance of Nda2 for nonphotochemical PQ reduction and associated processes in C. reinhardtii. [less ▲]

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See detailThe Mehler reaction in Chlamydomonas during photosynthetic induction and steady-state photosynthesis in wild-type and in a mitochondrial mutant
Franck, Fabrice ULg; Houyoux, Pierre-Alain

in Allen, J. F.; Gantt, E.; Golbeck, J. H. (Eds.) et al Photosynthesis: Energy from the Sun (2008)

The effects of fast O2-removal on electron transport rate (ETR), measured by chlorophyll fluorescence, were investigated in C. reinhardtii. In wild-type, O2-removal caused a strong decrease in ETR during ... [more ▼]

The effects of fast O2-removal on electron transport rate (ETR), measured by chlorophyll fluorescence, were investigated in C. reinhardtii. In wild-type, O2-removal caused a strong decrease in ETR during photosynthetic induction due to the absence of Mehler reaction. Light-saturation curves of the O2-dependent electron flow were established in different conditions. O2-dependent electron flow was triggered within a few seconds and amounted to up to 80 % of total electron flow during photosynthetic induction in algae grown on minimal medium, while it represented less than 15 % at steady-state. It was significantly weaker in the presence of acetate and in a mutant devoided of mitochondrial complexes I and III. Measurements of the light-induced changes of 820nm transmission in the s time-range show that in wild-type the O2-dependent electron flow leads to complete re-oxidation of PSI donor side in saturating light. [less ▲]

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