Spécialité: Biologie cellulaire et moléculaire Identification des protéines PPR impliquées dans l... more Spécialité: Biologie cellulaire et moléculaire Identification des protéines PPR impliquées dans l’épissage des ARN messagers dans les chloroplastes et les mitochondries chez Arabidopsis Thaliana.
Pathogens deploy effector proteins that interact with host proteins to manipulate the host physio... more Pathogens deploy effector proteins that interact with host proteins to manipulate the host physiology to the pathogen’s own benefit. However, effectors can also be recognized by host immune proteins leading to the activation of defense responses. Effectors are thus essential components in determining the outcome of plant-pathogen interactions. Despite major efforts to decipher effector functions, our current knowledge on effector biology is scattered and often limited. In this study, we conducted two systematic large-scale yeast two-hybrid screenings to detect interactions between Arabidopsis thaliana proteins and effectors from two vascular bacterial pathogens: Ralstonia pseudosolanacearum and Xanthomonas campestris. We then constructed an interactomic network focused on Arabidopsis and effector proteins from a wide variety of bacterial, oomycete, fungal and animal pathogens. This network contains our experimental data and protein-protein interactions from 2,035 peer-reviewed publi...
The energy transfer from Nd3+ to Yb3+ has been demonstrated by timeresolved spectroscopy in glass... more The energy transfer from Nd3+ to Yb3+ has been demonstrated by timeresolved spectroscopy in glasses of composition 15Li2O-10BaO-(5 − x − y)-La2O3-xNd2O3-yYb2O3-3Al2O3-67B2O3 (0 < x + y ⩽ 5). The microparameter CDA (Nd3+ → Yb3+) characterizing the transfer probability is quite high: 60 × 10−40 cm6 s−1. From the experimental fluorescence decays of donor and acceptor the transfer yield calculated for the glass with x = 1, y = 2 is equal to 87%. For low Yb3+ concentrations a Yb3+ → Nd3+ back transfer exists.
L'invention concerne un regulateur negatif de la biosynthese d'ethylene dans des plantes.... more L'invention concerne un regulateur negatif de la biosynthese d'ethylene dans des plantes. Ce regulateur est mis en oeuvre par ETHYLENE-OVERPRODUCER1 (ETO1), un gene recessif regulant l'expression de la synthese d'ethylene hormonal vegetal. ETO1 est un membre d'une nouvelle famille de genes presentant un degre de conservation eleve dans le regne vegetal et comprenant des domaines BTB et TPR, d'ou son caractere unique. Des experiences a deux hybrides conjuguees a des experiences de liaison in vitro et a des analyses de double mutant in planta ont revele une interaction directe entre ETO1 et la 1-aminocyclopropane-1-carboxylate synthase (ACS), une enzyme jouant un role cle dans la synthese d'ethylene. Par ailleurs, un essai fonctionnel dans E. coli a montre que les membres de la famille des proteines ETO1-LIKE (EOL) associees inhibent aussi fortement la biosynthese d'ethylene. L'invention concerne en outre des methodes de regulation et de modulation ...
Philosophical Transactions of the Royal Society B: Biological Sciences
The early steps in germination and development of angiosperm seedlings often occur in the dark, i... more The early steps in germination and development of angiosperm seedlings often occur in the dark, inducing a special developmental programme called skoto-morphogenesis. Under these conditions photosynthesis cannot work and all energetic requirements must be fulfilled by mitochondrial metabolization of storage energies. Here, we report the physiological impact of mitochondrial dysfunctions on the skoto-morphogenic programme by using the Arabidopsis rpoTmp mutant. This mutant is defective in the T7-phage-type organellar RNA polymerase shared by plastids and mitochondria. Lack of this enzyme causes a mitochondrial dysfunction resulting in a strongly reduced mitochondrial respiratory chain and a compensatory upregulation of the alternative-oxidase (AOX)-dependent respiration. Surprisingly, the mutant exhibits a triple-response-like phenotype with a twisted apical hook and a shortened hypocotyl. Highly similar phenotypes were detected in other respiration mutants ( rug3 and atphb3) and in ...
C to U editing is one of the post-transcriptional steps which are required for the proper express... more C to U editing is one of the post-transcriptional steps which are required for the proper expression of chloroplast and mitochondrial genes in plants. It depends on several proteins acting together which include the PLS-class pentatricopeptide repeat proteins (PPR). DYW2 was recently shown to be required for the editing of many sites in both organelles. In particular almost all the sites associated with the E+ subfamily of PPR proteins are depending on DYW2, suggesting that DYW2 is required for the function of E+-type PPR proteins. Here we strengthened this link by identifying 16 major editing sites controlled by 3 PPR proteins: OTP90, a DYW-type PPR and PGN and MEF37, 2 E+-type PPR proteins. A re-analysis of the DYW2 editotype showed that the 49 sites known to be associated with the 18 characterized E+-type PPR proteins all depend on DYW2. Considering only the 288 DYW2-dependent editing sites as potential E+-type PPR sites, instead of the 795 known editing sites, improves the perfo...
The lack of resolution on the many different ubiquitin chain types found in eukaryotic cells has ... more The lack of resolution on the many different ubiquitin chain types found in eukaryotic cells has been a major hurdle to our understanding of their specific roles. We currently have very little insight into the cellular and physiological functions of lysine(K)63-linked ubiquitin chains, although they are the second most abundant forms of ubiquitin in plant cells. To overcome this, we developed several large-scale approaches aiming at characterizing i) the E2-E3 ubiquitination machinery driving K63-linked ubiquitin chain formation, and ii) K63 polyubiquitination targets to provide the most comprehensive picture of K63 polyubiquitin networks in plants. Our work identified the UBC35/36 E2s as the major driver of K63 polyubiquitin chain formation and highlights the major role of these E2s in plant growth and development. Interactome approaches allowed us to identify many proteins interacting with the K63 polyubiquitination-dedicated UBC35/36 E2s and their cognate E2 variants, including over a dozen of E3s and their putative targets. In parallel, we have improved the in vivo detection of proteins decorated with K63-linked ubiquitin chains by sensor-based proteomics, yielding new insight into K63 polyubiquitination roles in plant cells. Altogether, this work strongly increases our understanding of K63 polyubiquitination networks and functions in plants.
RNA editing is converting hundreds of cytosines into uridines during organelle gene expression of... more RNA editing is converting hundreds of cytosines into uridines during organelle gene expression of land plants. The pentatricopeptide repeat (PPR) proteins are at the core of this posttranscriptional RNA modification. Even if a PPR protein defines the editing site, a DYW domain of the same or another PPR protein is believed to catalyze the deamination. To give insight into the organelle RNA editosome, we performed tandem affinity purification of the plastidial CHLOROPLAST BIOGENESIS 19 (CLB19) PPR editing factor. Two PPR proteins, dually targeted to mitochondria and chloroplasts, were identified as potential partners of CLB19. These two proteins, a P-type PPR and a member of a small PPR-DYW subfamily, were shown to interact in yeast. Insertional mutations resulted in embryo lethality that could be rescued by embryo-specific complementation. A transcriptome analysis of these complemented plants showed major editing defects in both organelles with a very high PPR type specificity, indi...
Numerous statistical pipelines are now available for the differential analysis of gene expression... more Numerous statistical pipelines are now available for the differential analysis of gene expression measured with RNA-sequencing technology. Most of them are based on similar statistical frameworks after normalization, differing primarily in the choice of data distribution, mean and variance estimation strategy and data filtering. We propose an evaluation of the impact of these choices when few biological replicates are available through the use of synthetic data sets. This framework is based on real data sets and allows the exploration of various scenarios differing in the proportion of non-differentially expressed genes. Hence, it provides an evaluation of the key ingredients of the differential analysis, free of the biases associated with the simulation of data using parametric models. Our results show the relevance of a proper modeling of the mean by using linear or generalized linear modeling. Once the mean is properly modeled, the impact of the other parameters on the performance of the test is much less important. Finally, we propose to use the simple visualization of the raw P-value histogram as a practical evaluation criterion of the performance of differential analysis methods on real data sets.
Spécialité: Biologie cellulaire et moléculaire Identification des protéines PPR impliquées dans l... more Spécialité: Biologie cellulaire et moléculaire Identification des protéines PPR impliquées dans l’épissage des ARN messagers dans les chloroplastes et les mitochondries chez Arabidopsis Thaliana.
Pathogens deploy effector proteins that interact with host proteins to manipulate the host physio... more Pathogens deploy effector proteins that interact with host proteins to manipulate the host physiology to the pathogen’s own benefit. However, effectors can also be recognized by host immune proteins leading to the activation of defense responses. Effectors are thus essential components in determining the outcome of plant-pathogen interactions. Despite major efforts to decipher effector functions, our current knowledge on effector biology is scattered and often limited. In this study, we conducted two systematic large-scale yeast two-hybrid screenings to detect interactions between Arabidopsis thaliana proteins and effectors from two vascular bacterial pathogens: Ralstonia pseudosolanacearum and Xanthomonas campestris. We then constructed an interactomic network focused on Arabidopsis and effector proteins from a wide variety of bacterial, oomycete, fungal and animal pathogens. This network contains our experimental data and protein-protein interactions from 2,035 peer-reviewed publi...
The energy transfer from Nd3+ to Yb3+ has been demonstrated by timeresolved spectroscopy in glass... more The energy transfer from Nd3+ to Yb3+ has been demonstrated by timeresolved spectroscopy in glasses of composition 15Li2O-10BaO-(5 − x − y)-La2O3-xNd2O3-yYb2O3-3Al2O3-67B2O3 (0 < x + y ⩽ 5). The microparameter CDA (Nd3+ → Yb3+) characterizing the transfer probability is quite high: 60 × 10−40 cm6 s−1. From the experimental fluorescence decays of donor and acceptor the transfer yield calculated for the glass with x = 1, y = 2 is equal to 87%. For low Yb3+ concentrations a Yb3+ → Nd3+ back transfer exists.
L'invention concerne un regulateur negatif de la biosynthese d'ethylene dans des plantes.... more L'invention concerne un regulateur negatif de la biosynthese d'ethylene dans des plantes. Ce regulateur est mis en oeuvre par ETHYLENE-OVERPRODUCER1 (ETO1), un gene recessif regulant l'expression de la synthese d'ethylene hormonal vegetal. ETO1 est un membre d'une nouvelle famille de genes presentant un degre de conservation eleve dans le regne vegetal et comprenant des domaines BTB et TPR, d'ou son caractere unique. Des experiences a deux hybrides conjuguees a des experiences de liaison in vitro et a des analyses de double mutant in planta ont revele une interaction directe entre ETO1 et la 1-aminocyclopropane-1-carboxylate synthase (ACS), une enzyme jouant un role cle dans la synthese d'ethylene. Par ailleurs, un essai fonctionnel dans E. coli a montre que les membres de la famille des proteines ETO1-LIKE (EOL) associees inhibent aussi fortement la biosynthese d'ethylene. L'invention concerne en outre des methodes de regulation et de modulation ...
Philosophical Transactions of the Royal Society B: Biological Sciences
The early steps in germination and development of angiosperm seedlings often occur in the dark, i... more The early steps in germination and development of angiosperm seedlings often occur in the dark, inducing a special developmental programme called skoto-morphogenesis. Under these conditions photosynthesis cannot work and all energetic requirements must be fulfilled by mitochondrial metabolization of storage energies. Here, we report the physiological impact of mitochondrial dysfunctions on the skoto-morphogenic programme by using the Arabidopsis rpoTmp mutant. This mutant is defective in the T7-phage-type organellar RNA polymerase shared by plastids and mitochondria. Lack of this enzyme causes a mitochondrial dysfunction resulting in a strongly reduced mitochondrial respiratory chain and a compensatory upregulation of the alternative-oxidase (AOX)-dependent respiration. Surprisingly, the mutant exhibits a triple-response-like phenotype with a twisted apical hook and a shortened hypocotyl. Highly similar phenotypes were detected in other respiration mutants ( rug3 and atphb3) and in ...
C to U editing is one of the post-transcriptional steps which are required for the proper express... more C to U editing is one of the post-transcriptional steps which are required for the proper expression of chloroplast and mitochondrial genes in plants. It depends on several proteins acting together which include the PLS-class pentatricopeptide repeat proteins (PPR). DYW2 was recently shown to be required for the editing of many sites in both organelles. In particular almost all the sites associated with the E+ subfamily of PPR proteins are depending on DYW2, suggesting that DYW2 is required for the function of E+-type PPR proteins. Here we strengthened this link by identifying 16 major editing sites controlled by 3 PPR proteins: OTP90, a DYW-type PPR and PGN and MEF37, 2 E+-type PPR proteins. A re-analysis of the DYW2 editotype showed that the 49 sites known to be associated with the 18 characterized E+-type PPR proteins all depend on DYW2. Considering only the 288 DYW2-dependent editing sites as potential E+-type PPR sites, instead of the 795 known editing sites, improves the perfo...
The lack of resolution on the many different ubiquitin chain types found in eukaryotic cells has ... more The lack of resolution on the many different ubiquitin chain types found in eukaryotic cells has been a major hurdle to our understanding of their specific roles. We currently have very little insight into the cellular and physiological functions of lysine(K)63-linked ubiquitin chains, although they are the second most abundant forms of ubiquitin in plant cells. To overcome this, we developed several large-scale approaches aiming at characterizing i) the E2-E3 ubiquitination machinery driving K63-linked ubiquitin chain formation, and ii) K63 polyubiquitination targets to provide the most comprehensive picture of K63 polyubiquitin networks in plants. Our work identified the UBC35/36 E2s as the major driver of K63 polyubiquitin chain formation and highlights the major role of these E2s in plant growth and development. Interactome approaches allowed us to identify many proteins interacting with the K63 polyubiquitination-dedicated UBC35/36 E2s and their cognate E2 variants, including over a dozen of E3s and their putative targets. In parallel, we have improved the in vivo detection of proteins decorated with K63-linked ubiquitin chains by sensor-based proteomics, yielding new insight into K63 polyubiquitination roles in plant cells. Altogether, this work strongly increases our understanding of K63 polyubiquitination networks and functions in plants.
RNA editing is converting hundreds of cytosines into uridines during organelle gene expression of... more RNA editing is converting hundreds of cytosines into uridines during organelle gene expression of land plants. The pentatricopeptide repeat (PPR) proteins are at the core of this posttranscriptional RNA modification. Even if a PPR protein defines the editing site, a DYW domain of the same or another PPR protein is believed to catalyze the deamination. To give insight into the organelle RNA editosome, we performed tandem affinity purification of the plastidial CHLOROPLAST BIOGENESIS 19 (CLB19) PPR editing factor. Two PPR proteins, dually targeted to mitochondria and chloroplasts, were identified as potential partners of CLB19. These two proteins, a P-type PPR and a member of a small PPR-DYW subfamily, were shown to interact in yeast. Insertional mutations resulted in embryo lethality that could be rescued by embryo-specific complementation. A transcriptome analysis of these complemented plants showed major editing defects in both organelles with a very high PPR type specificity, indi...
Numerous statistical pipelines are now available for the differential analysis of gene expression... more Numerous statistical pipelines are now available for the differential analysis of gene expression measured with RNA-sequencing technology. Most of them are based on similar statistical frameworks after normalization, differing primarily in the choice of data distribution, mean and variance estimation strategy and data filtering. We propose an evaluation of the impact of these choices when few biological replicates are available through the use of synthetic data sets. This framework is based on real data sets and allows the exploration of various scenarios differing in the proportion of non-differentially expressed genes. Hence, it provides an evaluation of the key ingredients of the differential analysis, free of the biases associated with the simulation of data using parametric models. Our results show the relevance of a proper modeling of the mean by using linear or generalized linear modeling. Once the mean is properly modeled, the impact of the other parameters on the performance of the test is much less important. Finally, we propose to use the simple visualization of the raw P-value histogram as a practical evaluation criterion of the performance of differential analysis methods on real data sets.
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