References of "Remacle, Claire"
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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 detailLoss of mitochondrial ATP synthase subunit beta (Atp2) alters mitochondrial and chloroplastic function and morphology in Chlamydomonas.
Lapaille, M.; Thiry, Marc ULg; Perez, E. et al

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

Mitochondrial F(1)F(O) ATP synthase (Complex V) catalyses ATP synthesis from ADP and inorganic phosphate using the proton-motive force generated by the substrate-driven electron transfer chain. In this ... [more ▼]

Mitochondrial F(1)F(O) ATP synthase (Complex V) catalyses ATP synthesis from ADP and inorganic phosphate using the proton-motive force generated by the substrate-driven electron transfer chain. In this work, we investigated the impact of the loss of activity of the mitochondrial enzyme in a photosynthetic organism. In this purpose, we inactivated by RNA interference the expression of the ATP2 gene, coding for the catalytic subunit beta, in the green alga Chlamydomonas reinhardtii. We demonstrate that in the absence of beta subunit, complex V is not assembled, respiratory rate is decreased by half and ATP synthesis coupled to the respiratory activity is fully impaired. Lack of ATP synthase also affects the morphology of mitochondria which are deprived of cristae. We also show that mutants are obligate phototrophs and that rearrangements of the photosynthetic apparatus occur in the chloroplast as a response to ATP synthase deficiency in mitochondria. Altogether, our results contribute to the understanding of the yet poorly studied bioenergetic interactions between organelles in photosynthetic organisms. [less ▲]

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See detailSubunit-subunit interactions and overall topology of the dimeric mitochondrial ATP synthase of Polytomella sp.
Cano-Estrada, A.; Vazquez-Acevedo, M.; Villavicencio-Queijeiro, A. et al

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

Mitochondrial F(1)F(0)-ATP synthase of chlorophycean algae is a dimeric complex of 1600kDa constituted by 17 different subunits with varying stoichiometries, 8 of them conserved in all eukaryotes and 9 ... [more ▼]

Mitochondrial F(1)F(0)-ATP synthase of chlorophycean algae is a dimeric complex of 1600kDa constituted by 17 different subunits with varying stoichiometries, 8 of them conserved in all eukaryotes and 9 that seem to be unique to the algal lineage (subunits ASA1-9). Two different models proposing the topological assemblage of the nine ASA subunits in the ATP synthase of the colorless alga Polytomella sp. have been put forward. Here, we readdressed the overall topology of the enzyme with different experimental approaches: detection of close vicinities between subunits based on cross-linking experiments and dissociation of the enzyme into subcomplexes, inference of subunit stoichiometry based on cysteine residue labelling, and general three-dimensional structural features of the complex as obtained from small-angle X-ray scattering and electron microscopy image reconstruction. Based on the available data, we refine the topological arrangement of the subunits that constitute the mitochondrial ATP synthase of Polytomella sp. [less ▲]

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See detailKnock-down of the COX3 and COX17 gene expression of cytochrome c oxidase in the unicellular green alga Chlamydomonas reinhardtii.
Remacle, Claire ULg; Coosemans, Nadine ULg; Jans, Frédéric ULg et al

in Plant Molecular Biology (2010), 74(3), 223-2363

The COX3 gene encodes a core subunit of mitochondrial cytochrome c oxidase (complex IV) whereas the COX17 gene encodes a chaperone delivering copper to the enzyme. Mutants of these two genes were isolated ... [more ▼]

The COX3 gene encodes a core subunit of mitochondrial cytochrome c oxidase (complex IV) whereas the COX17 gene encodes a chaperone delivering copper to the enzyme. Mutants of these two genes were isolated by RNA interference in the microalga Chlamydomonas. The COX3 mRNA was completely lacking in the cox3-RNAi mutant and no activity and assembly of complex IV were detected. The cox17-RNAi mutant presented a reduced level of COX17 mRNA, a reduced activity of the cytochrome c oxidase but no modification of its amount. The cox3-RNAi mutant had only 40% of the wild-type rate of dark respiration which was cyanide-insensitive. The mutant presented a 60% decrease of H(2)O(2) production in the dark compared to wild type, which probably accounts for a reduced electron leakage by respiratory complexes III and IV. In contrast, the cox17-RNAi mutant showed no modification of respiration and of H(2)O(2) production in the dark but a two to threefold increase of H(2)O(2) in the light compared to wild type and the cox3-RNAi mutant. The cox17-RNAi mutant was more sensitive to cadmium than the wild-type and cox3-RNAi strains. This suggested that besides its role in complex IV assembly, Cox17 could have additional functions in the cell such as metal detoxification or Reactive Oxygen Species protection or signaling. Concerning Cox3, its role in Chlamydomonas complex IV is similar to that of other eukaryotes although this subunit is encoded in the nuclear genome in the alga contrary to the situation found in all other organisms. [less ▲]

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See detailF1FO ATP synthase mutants in Chlamydomonas: Stability and oligomycin resistance mediated by atypical Asa7 protein; interaction between chloroplastic and mitochondrial bioenergetics
Lapaille, Marie ULg; Escobar-Ramírez, Adelma; Degand, Hervé et al

in Biochimica et Biophysica Acta (BBA) - Bioenergetics (2010), 1797(Supplement 1), 29

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See detailCharacterization of complex I mutants in Chlamydomonas reinhardtii : Role of structural subunits and identification of assembly factors.
Larosa, Véronique ULg; Barbieri, Rosario; Bonnefoy, Nathalie et al

Scientific conference (2009)

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See detailThe ARG9 Gene Encodes the Plastid-Resident N-Acetyl Ornithine Aminotransferase in the Green Alga Chlamydomonas reinhardtii
Remacle, Claire ULg; Cline, Sara; Boutaffala, Layla ULg et al

in Eukaryotic Cell (2009), 8(9), 1460-1463

Here we report the characterization of the Chlamydomonas reinhardtii gene ARG9, encoding the plastid resident N-acetyl ornithine aminotransferase, which is involved in arginine synthesis. Integration of ... [more ▼]

Here we report the characterization of the Chlamydomonas reinhardtii gene ARG9, encoding the plastid resident N-acetyl ornithine aminotransferase, which is involved in arginine synthesis. Integration of an engineered ARG9 cassette in the plastid chromosome of the nuclear arg9 mutant restores arginine prototrophy. This suggests that ARG9 could be used as a new selectable marker for plastid transformation. [less ▲]

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See detailSteady-state levels of imported tRNAs in Chlamydomonas mitochondria are correlated with both cytosolic and mitochondrial codon usages
Vinogradova, Elizaveta; Salinas, Thalia; Cognat, Valerie et al

in Nucleic Acids Research (2009), 37(5), 1521-1528

The mitochondrial genome of Chlamydomonas reinhardtii only encodes three expressed tRNA genes, thus most mitochondrial tRNAs are likely imported. The sharing of tRNAs between chloroplasts and mitochondria ... [more ▼]

The mitochondrial genome of Chlamydomonas reinhardtii only encodes three expressed tRNA genes, thus most mitochondrial tRNAs are likely imported. The sharing of tRNAs between chloroplasts and mitochondria has been speculated in this organism. We first demonstrate that no plastidial tRNA is present in mitochondria and that the mitochondrial translation mainly relies on the import of nucleus-encoded tRNA species. Then, using northern analysis, we show that the extent of mitochondrial localization for the 49 tRNA isoacceptor families encoded by the C. reinhardtii nuclear genome is highly variable. Until now the reasons for such variability were unknown. By comparing cytosolic and mitochondrial codon usage with the sub-cellular distribution of tRNAs, we provide unprecedented evidence that the steady-state level of a mitochondrial tRNA is linked not only to the frequency of the cognate codon in mitochondria but also to its frequency in the cytosol, then allowing optimal mitochondrial translation. [less ▲]

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See detailOxidative phosphorylation: Building blocks and related components
Cardol, Pierre ULg; Figueroa, Francisco; Remacle, Claire ULg et al

in Stern, David; Harris, Elizabeth; Witman, George (Eds.) The Chlamydomonas Sourcebook 3-vol set, 1-3 (2009)

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See detailThe mitochondrial genome
Cardol, Pierre ULg; Remacle, Claire ULg

in Stern, David; Harris, Elizabeth; Witman, George (Eds.) The Chlamydomonas Sourcebook 3-vol set, 1-3 (2009)

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See detailThe fully-active and structurally-stable form of the mitochondrial ATP synthase of Polytomella sp is dimeric
Villavicencio-Queijeiro, Alexa; Vazquez-Acevedo, Miriam; Cano-Estrada, Araceli et al

in Journal of Bioenergetics & Biomembranes (2009), 41(1), 1-13

Mitochondrial F1FO-ATP synthase of chlorophycean algae is a stable dimeric complex of 1,600 kDa. It lacks the classic subunits that constitute the peripheral stator-stalk and the orthodox polypeptides ... [more ▼]

Mitochondrial F1FO-ATP synthase of chlorophycean algae is a stable dimeric complex of 1,600 kDa. It lacks the classic subunits that constitute the peripheral stator-stalk and the orthodox polypeptides involved in the dimerization of the complex. Instead, it contains nine polypeptides of unknown evolutionary origin named ASA1 to ASA9. The isolated enzyme exhibited a very low ATPase activity (0.03 Units/mg), that increased upon heat treatment, due to the release of the F-1 sector. Oligomycin was found to stabilize the dimeric structure of the enzyme, providing partial resistance to heat dissociation. Incubation in the presence of low concentrations of several non-ionic detergents increased the oligomycin-sensitive ATPase activity up to 7.0-9.0 Units/mg. Incubation with 3% (w/v) taurodeoxycholate monomerized the enzyme. The monomeric form of the enzyme exhibited diminished activity in the presence of detergents and diminished oligomycin sensitivity. Cross-linking experiments carried out with the dimeric and monomeric forms of the ATP synthase suggested the participation of the ASA6 subunit in the dimerization of the enzyme. The dimeric enzyme was more resistant to heat treatment, high hydrostatic pressures, and protease digestion than the monomeric enzyme, which was readily disrupted by these treatments. We conclude that the fully-active algal mitochondrial ATP synthase is a stable catalytically active dimer; the monomeric form is less active and less stable. Monomer-monomer interactions could be mediated by the membrane-bound subunits ASA6 and ASA9, and may be further stabilized by other polypeptides such as ASA1 and ASA5. [less ▲]

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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 detailIn Chlamydomonas, the Loss of Nd5 Subunit Prevents the Assembly of Whole Mitochondrial Complex I and Leads to the Formation of a Low Abundant 700 Kda Subcomplex
Cardol, Pierre ULg; Boutaffala, Layla ULg; Memmi, S. et al

in Biochimica et Biophysica Acta-Bioenergetics (2008), 1777

In the green alga Chlamydomonas reinhardtii, a mutant deprived of complex I enzyme activity presents a 1T deletion in the mitochondrial nd5 gene. The loss of the ND5 subunit prevents the assembly of the ... [more ▼]

In the green alga Chlamydomonas reinhardtii, a mutant deprived of complex I enzyme activity presents a 1T deletion in the mitochondrial nd5 gene. The loss of the ND5 subunit prevents the assembly of the 950 kDa whole complex I. Instead, a low abundant 700 kDa subcomplex, loosely associated to the inner mitochondrial membrane, is assembled. The resolution of the subcomplex by SDS-PAGE gave rise to 19 individual spots, sixteen having been identified by mass spectrometry analysis. Eleven, mainly associated to the hydrophilic part of the complex, are homologs to subunits of the bovine enzyme whereas five (including gamma-type carbonic anhydrase subunits) are specific to green plants or to plants and fungi. None of the subunits typical of the beta membrane domain of complex I enzyme has been identified in the mutant. This allows us to propose that the truncated enzyme misses the membrane distal domain of complex I but retains the proximal domain associated to the matrix arm of the enzyme. A complex I topology model is presented in the light of our results. Finally, a supercomplex most probably corresponding to complex I-complex III association, was identified in mutant mitochondria, indicating that the missing part of the enzyme is not required for the formation of the supercomplex. [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 detailOn the evolution and expression of Chlamydomonas reinhardtii nucleus-encoded transfer RNA genes
Cognat, Valerie; Deragon, Jean*-Marc; Vinogradova, Elizaveta et al

in Genetics (2008), 179(1), 113-123

In Chlamydomonas reinhardtii, 259 tRNA genes were identified and classified into 49 tRNA isoaccepting families. By constructing phylogenetic trees, we determined the evolutionary history for each tRNA ... [more ▼]

In Chlamydomonas reinhardtii, 259 tRNA genes were identified and classified into 49 tRNA isoaccepting families. By constructing phylogenetic trees, we determined the evolutionary history for each tRNA gene family. The majority of the IRNA sequences are more closely related to their plant counterparts than to animals ones. Northern experiments also permitted LIS to show that at least one member of each IRNA isoacceptor family is transcribed and correctly processed in vivo. A short stretch of T residues known to be a signal for termination of polymerase III transcription was found downstream of most IRNA genes. It allowed us to propose that the vast majority of the IRNA genes are expressed and to confirm that numerous IRNA genes separated by short spacers are indeed cotranscribed. Interestingly, in silico analyses and hybridization experiments show that the cellular IRNA abundance is correlated with the number of tRTNA genes and is adjusted to the codon usage to optimize translation efficiency. Finally, we studied the origin of SINEs, short interspersed elements related to tRNAs, whose presence in Chlamydomonas is exceptional. Phylogenetic analysis strongly suggests that tRNA(Asp)-related SINEs originate front a prokaryotic-type IRNA either horizontally transferred from a bacterium or originally present in mitochondria or chloroplasts. [less ▲]

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See detailEukaryotic complex I: functional diversity and experimental systems to unravel the assembly process
Remacle, Claire ULg; Barbieri, Rosario; Cardol, Pierre ULg et al

in Molecular Genetics & Genomics (2008), 280

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