Michel Zivy

Publications (32)124.73 Total impact

• Source

  Available from: arxiv.org

  Article: MassChroQ: a versatile tool for mass spectrometry quantification.

  Benoît Valot, Olivier Langella, Edlira Nano, Michel Zivy
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  ABSTRACT: Recently, many software tools have been developed to perform quantification in LC-MS analyses. However, most of them are specific to either a quantification strategy (e.g. label-free or isotopic labelling) or a mass-spectrometry system (e.g. high or low resolution). In this context, we have developed MassChroQ (Mass Chromatogram Quantification), a versatile software that performs LC-MS data alignment and peptide quantification by peak area integration on extracted ion chromatograms. MassChroQ is suitable for quantification with or without labelling and is not limited to high-resolution systems. Peptides of interest (for example all the identified peptides) can be determined automatically, or manually by providing targeted m/z and retention time values. It can handle large experiments that include protein or peptide fractionation (as SDS-PAGE, 2-D LC). It is fully configurable. Every processing step is traceable, the produced data are in open standard formats and its modularity allows easy integration into proteomic pipelines. The output results are ready for use in statistical analyses. Evaluation of MassChroQ on complex label-free data obtained from low and high-resolution mass spectrometers showed low CVs for technical reproducibility (1.4%) and high coefficients of correlation to protein quantity (0.98). MassChroQ is freely available under the GNU General Public Licence v3.0 at http://pappso.inra.fr/bioinfo/masschroq/.
  Proteomics 09/2011; 11(17):3572-7. · 4.43 Impact Factor
• Article: The ZmASR1 protein influences branched-chain amino acid biosynthesis and maintains kernel yield in maize under water-limited conditions.

  Laetitia Virlouvet, Marie-Pierre Jacquemot, Denise Gerentes, Hélène Corti, Sophie Bouton, Françoise Gilard, Benoît Valot, Jacques Trouverie, Guillaume Tcherkez, Matthieu Falque, Catherine Damerval, Peter Rogowsky, Pascual Perez, Graham Noctor, Michel Zivy, Sylvie Coursol
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  ABSTRACT: Abscisic acid-, stress-, and ripening-induced (ASR) proteins were first described about 15 years ago as accumulating to high levels during plant developmental processes and in response to diverse stresses. Currently, the effects of ASRs on water deficit tolerance and the ways in which their physiological and biochemical functions lead to this stress tolerance remain poorly understood. Here, we characterized the ASR gene family from maize (Zea mays), which contains nine paralogous genes, and showed that maize ASR1 (ZmASR1) was encoded by one of the most highly expressed paralogs. Ectopic expression of ZmASR1 had a large overall impact on maize yield that was maintained under water-limited stress conditions in the field. Comparative transcriptomic and proteomic analyses of wild-type and ZmASR1-overexpressing leaves led to the identification of three transcripts and 16 proteins up- or down-regulated by ZmASR1. The majority of them were involved in primary and/or cellular metabolic processes, including branched-chain amino acid (BCAA) biosynthesis. Metabolomic and transcript analyses further indicated that ZmASR1-overexpressing plants showed a decrease in BCAA compounds and changes in BCAA-related gene expression in comparison with wild-type plants. Interestingly, within-group correlation matrix analysis revealed a close link between 13 decreased metabolites in ZmASR1-overexpressing leaves, including two BCAAs. Among these 13 metabolites, six were previously shown to be negatively correlated to biomass, suggesting that ZmASR1-dependent regulation of these 13 metabolites might contribute to regulate leaf growth, resulting in improvement in kernel yield.
  Plant physiology 08/2011; 157(2):917-36. · 6.53 Impact Factor
• Article: Oil body proteins sequentially accumulate throughout seed development in Brassica napus.

  Pascale Jolivet, Céline Boulard, Annick Bellamy, Benoît Valot, Sabine d'Andréa, Michel Zivy, Nathalie Nesi, Thierry Chardot
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  ABSTRACT: Despite the importance of seed oil bodies (OBs) as enclosed compartments for oil storage, little is known about lipid and protein accumulation in OBs during seed formation. OBs from rapeseed (Brassica napus) consist of a triacylglycerol (TAG) core surrounded by a phospholipid monolayer embedded with integral proteins which confer high stability to OBs in the mature dry seed. In the present study, we investigated lipid and protein accumulation patterns throughout seed development (from 5 to 65 days after pollination [DAP]) both in the whole seed and in purified OBs. Deposition of the major proteins (oleosins, caleosins and steroleosins) into OBs was assessed through (i) gene expression pattern, (ii) proteomics analysis, and (iii) protein immunodetection. For the first time, a sequential deposition of integral OB proteins was established. Accumulation of oleosins and caleosins was observed starting from early stages of seed development (12-17 DAP), while steroleosins accumulated later (~25 DAP) onwards.
  Journal of plant physiology 07/2011; 168(17):2015-20. · 2.50 Impact Factor
• Article: Analysis of the xylem sap proteome of Brassica oleracea reveals a high content in secreted proteins.

  Laetitia Ligat, Emmanuelle Lauber, Cécile Albenne, Hélène San Clemente, Benoît Valot, Michel Zivy, Rafael Pont-Lezica, Matthieu Arlat, Elisabeth Jamet
  [show abstract] [hide abstract]
  ABSTRACT: Xylem plays a major role in plant development and is considered part of the apoplast. Here, we studied the proteome of Brassica oleracea cv Bartolo and compared it to the plant cell wall proteome of another Brassicaceae, the model plant Arabidopsis thaliana. B. oleracea was chosen because it is technically difficult to harvest enough A. thaliana xylem sap for proteomic analysis. We studied the whole proteome and an N-glycoproteome obtained after Concanavalin A affinity chromatography. Altogether, 189 proteins were identified by LC-MS/MS using Brassica EST and cDNA sequences. A predicted signal peptide was found in 164 proteins suggesting that most proteins of the xylem sap are secreted. Eighty-one proteins were identified in the N-glycoproteome, with 25 of them specific of this fraction, suggesting that they were concentrated during the chromatography step. All the protein families identified in this study were found in the cell wall proteomes. However, proteases and oxido-reductases were more numerous in the xylem sap proteome, whereas enzyme inhibitors were rare. The origin of xylem sap proteins is discussed. All the experimental data including the MS/MS data were made available in the WallProtDB cell wall proteomic database.
  Proteomics 03/2011; 11(9):1798-813. · 4.43 Impact Factor
• Source

  Available from: ibpc.fr

  Article: Dynamics of post-translational modifications and protein stability in the stroma of Chlamydomonas reinhardtii chloroplasts.

  Willy V Bienvenut, Christelle Espagne, Aude Martinez, Wojetec Majeran, Benoît Valot, Michel Zivy, Olivier Vallon, Zach Adam, Thierry Meinnel, Carmela Giglione
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  ABSTRACT: The proteome of any system is a dynamic entity dependent on the intracellular concentration of the entire set of expressed proteins. In turn, this whole protein concentration will be reliant on the stability/turnover of each protein as dictated by their relative rates of synthesis and degradation. In this study, we have investigated the dynamics of the stromal proteome in the model organism Chlamydomonas reinhardtii by characterizing the half-life of the whole set of proteins. 2-DE stromal proteins profiling was set up and coupled with MS analyses. These identifications featuring an average of 26% sequence coverage and eight non-redundant peptides per protein have been obtained for 600 independent samples related to 253 distinct spots. An interactive map of the global stromal proteome, of 274 distinct protein variants is now available on-line at http://www.isv.cnrs-gif.fr/gel2dv2/. N-α-terminal-Acetylation (NTA) was noticed to be the most frequently detectable post-translational modification, and new experimental data related to the chloroplastic transit peptide cleavage site was obtained. Using this data set supplemented with series of pulse-chase experiments, elements directing the relationship between half-life and N-termini were analyzed. Positive correlation between NTA and protein half-life suggests that NTA could contribute to protein stabilization in the stroma.
  Proteomics 03/2011; 11(9):1734-50. · 4.43 Impact Factor
• Source

  Available from: Cécile Albenne

  Article: Combining various strategies to increase the coverage of the plant cell wall glycoproteome.

  Yu Zhang, Aurélie Giboulot, Michel Zivy, Benoît Valot, Elisabeth Jamet, Cécile Albenne
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  ABSTRACT: Glycoproteomics recently became a very active field, mostly in mammals. The first part of this paper consists of a mini-review on the strategies used in glycoproteomics, namely methods for enrichment in glycoproteins and mass spectrometry (MS) techniques currently used. In a second part, these strategies are applied to the cell wall glycoproteome of etiolated hypocotyls of Arabidopsis thaliana, showing their complementarity. Several sub-glycoproteomes were obtained by: (i) affinity chromatography on concanavaline A (ConA) and analysis of glycoproteins by MALDI-TOF MS; (ii) multidimensional lectin chromatography (using AIL, PNA, ConA and WGA lectins) and subsequent identification of glycoproteins by MALDI-TOF MS and LC-MS/MS; (iii) boronic acid chromatography followed by identification of glycoproteins by MALDI-TOF MS. Altogether, 127 glycoproteins were identified. Most glycoproteins were found to be putative N-glycoproteins and N-glycopeptides were predicted from MS data using the ProTerNyc bioinformatics software.
  Phytochemistry 11/2010; 72(10):1109-23. · 3.35 Impact Factor
• Article: Phospho-site mapping, genetic and in planta activation studies reveal key aspects of the different phosphorylation mechanisms involved in activation of SnRK2s.

  Florina Vlad, Marie-Jo Droillard, Benoît Valot, Mehdi Khafif, Americo Rodrigues, Mathias Brault, Michel Zivy, Pedro L Rodriguez, Sylvain Merlot, Christiane Laurière
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  ABSTRACT: Snf1-related protein kinases 2 (SnRK2s) are major positive regulators of drought stress tolerance. The kinases of this family are activated by hyperosmotic stress, but only some of them are also responsive to abscisic acid (ABA). Moreover, genetic evidence has indicated the ABA-independence of SnRK2 activation in the fast response to osmotic stress. Although phosphorylation was demonstrated to be crucial for the activation or activity of the kinases of both subgroups, different phosphorylation mechanisms were suggested. Here, using one kinase from each subgroup (SnRK2.6 and SnRK2.10), two phosphorylation sites within the activation loop were identified by mass spectrometry after immunoprecipitation from Arabidopsis cells treated by ABA or osmolarity. By site-directed mutagenesis, the phosphorylation of only one of the two sites was shown to be necessary for the catalytic activity of the kinase, whereas both sites are necessary for the full activation of the two SnRK2s by hyperosmolarity or ABA. Phosphoprotein staining together with two-dimensional PAGE followed by immunoblotting indicated distinct phosphorylation mechanisms of the two kinases. While SnRK2.6 seems to be activated through the independent phosphorylation of these two sites, a sequential process occurs in SnRK2.10, where phosphorylation of one serine is required for the phosphorylation of the other. In addition, a subgroup of protein phosphatases 2C which interact and participate in the regulation of SnRK2.6 do not interact with SnRK2.10. Taken together, our data bring evidence for the involvement of distinct phosphorylation mechanisms in the activation of SnRK2.6 and SnRK2.10, which may be conserved between the two subgroups of SnRK2s depending on their ABA-responsiveness.
  The Plant Journal 09/2010; 63(5):778-90. · 6.16 Impact Factor
• Source

  Available from: PubMed Central

  Article: The Arabidopsis ABA-activated kinase OST1 phosphorylates the bZIP transcription factor ABF3 and creates a 14-3-3 binding site involved in its turnover.

  Caroline Sirichandra, Marlène Davanture, Benjamin E Turk, Michel Zivy, Benoît Valot, Jeffrey Leung, Sylvain Merlot
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  ABSTRACT: Genetic evidence in Arabidopsis thaliana indicates that members of the Snf1-Related Kinases 2 family (SnRK2) are essential in mediating various stress-adaptive responses. Recent reports have indeed shown that one particular member, Open Stomata (OST)1, whose kinase activity is stimulated by the stress hormone abscisic acid (ABA), is a direct target of negative regulation by the core ABA co-receptor complex composed of PYR/PYL/RCAR and clade A Protein Phosphatase 2C (PP2C) proteins. Here, the substrate preference of OST1 was interrogated at a genome-wide scale. We phosphorylated in vitro a bank of semi-degenerate peptides designed to assess the relative phosphorylation efficiency on a positionally fixed serine or threonine caused by systematic changes in the flanking amino acid sequence. Our results designate the ABA-responsive-element Binding Factor 3 (ABF3), which controls part of the ABA-regulated transcriptome, as a genuine OST1 substrate. Bimolecular Fluorescence Complementation experiments indicate that ABF3 interacts directly with OST1 in the nuclei of living plant cells. In vitro, OST1 phosphorylates ABF3 on multiple LXRXXpS/T preferred motifs including T451 located in the midst of a conserved 14-3-3 binding site. Using an antibody sensitive to the phosphorylated state of the preferred motif, we further show that ABF3 is phosphorylated on at least one such motif in response to ABA in vivo and that phospho-T451 is important for stabilization of ABF3. All together, our results suggest that OST1 phosphorylates ABF3 in vivo on T451 to create a 14-3-3 binding motif. In a wider physiological context, we propose that the long term responses to ABA that require sustained gene expression is, in part, mediated by the stabilization of ABFs driven by ABA-activated SnRK2s.
  PLoS ONE 01/2010; 5(11):e13935. · 4.09 Impact Factor
• Article: Cotranslational proteolysis dominates glutathione homeostasis to support proper growth and development.

  Frédéric Frottin, Christelle Espagne, José A Traverso, Caroline Mauve, Benoît Valot, Caroline Lelarge-Trouverie, Michel Zivy, Graham Noctor, Thierry Meinnel, Carmela Giglione
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  ABSTRACT: The earliest proteolytic event affecting most proteins is the excision of the initiating Met (NME). This is an essential and ubiquitous cotranslational process tightly regulated in all eukaryotes. Currently, the effects of NME on unknown complex cellular networks and the ways in which its inhibition leads to developmental defects and cell growth arrest remain poorly understood. Here, we provide insight into the earliest molecular mechanisms associated with the inhibition of the NME process in Arabidopsis thaliana. We demonstrate that the developmental defects induced by NME inhibition are caused by an increase in cellular proteolytic activity, primarily induced by an increase in the number of proteins targeted for rapid degradation. This deregulation drives, through the increase of the free amino acids pool, a perturbation of the glutathione homeostasis, which corresponds to the earliest limiting, reversible step promoting the phenotype. We demonstrate that these effects are universally conserved and that the reestablishment of the appropriate glutathione status restores growth and proper development in various organisms. Finally, we describe a novel integrated model in which NME, protein N-alpha-acylation, proteolysis, and glutathione homeostasis operate in a sequentially regulated mechanism that directs both growth and development.
  The Plant Cell 10/2009; 21(10):3296-314. · 8.99 Impact Factor
• Article: Phosphorylation of the Arabidopsis AtrbohF NADPH oxidase by OST1 protein kinase.

  Caroline Sirichandra, Dan Gu, Heng-Cheng Hu, Marlène Davanture, Sangmee Lee, Michaël Djaoui, Benoît Valot, Michel Zivy, Jeffrey Leung, Sylvain Merlot, June M Kwak
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  ABSTRACT: The plant hormone abscisic acid (ABA) triggers production of reactive oxygen species (ROS) in guard cells via the AtrbohD and AtrbohF NADPH oxidases, leading to stomatal closure. The ABA-activated SnRK2 protein kinase open stomata 1 (OST1) (SRK2E/SnRK2.6) acts upstream of ROS in guard cell ABA signaling. Here, we report that OST1 phosphorylates Ser13 and Ser174 on AtrbohF. In addition, substitution of Ser174 to Ala results in a approximately 40% reduction in the phosphorylation of AtrbohF by OST1. We also show that OST1 physically interacts with AtrbohF. These results provide biochemical evidence suggesting that OST1 regulates AtrbohF activity.
  FEBS letters 09/2009; 583(18):2982-6. · 3.54 Impact Factor
• Source

  Available from: Warren Albertin

  Article: Comparative proteomics of leaf, stem, and root tissues of synthetic Brassica napus.

  Warren Albertin, Olivier Langella, Johann Joets, Luc Négroni, Michel Zivy, Catherine Damerval, Hervé Thiellement
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  ABSTRACT: Comparative proteomics was applied to three vegetative organs of Brassica napus, the leaf, stem, and root using 2-DE. Among the >1600 analyzed spots, 43% were found to be common to all three organs, suggesting the existence of a "basal" or ubiquitous proteome composed of housekeeping proteins. The green organs, leaf, and stem, were closely related (approximately 80% common spots) while the root displayed more organ-specific polypeptides (approximately 10%). Reference maps were established using MS, allowing the identification of 93, 385, and 266 proteins in leaf, stem, and root proteomes, respectively. Bioinformatic analyses were also performed; in silico functional categorization and cellular localization allow obtaining a precise picture of the cell molecular network within vegetative organs. These proteome maps can be explored using the PROTICdb software at the following address: http://bioinformatique.moulon.inra.fr/proticdb/web_view/.
  Proteomics 02/2009; 9(3):793-9. · 4.43 Impact Factor
• Article: A method based on bead flows for spot detection on 2-D gel images.

  Olivier Langella, Michel Zivy
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  ABSTRACT: A method has been developed to detect spots on 2-D gel images. It is based on an analogy with beads flowing uphill on the surface of the gel image and on the analysis of their paths. This method does not need image smoothing, thereby allowing for conservation of the resolution of the original image. In addition, it allows for the detection of overlapping spots, even when there is no valley between their centres.
  Proteomics 12/2008; 8(23-24):4914-8. · 4.43 Impact Factor
• Article: Extent of N-terminal modifications in cytosolic proteins from eukaryotes.

  Aude Martinez, José A Traverso, Benoît Valot, Myriam Ferro, Christelle Espagne, Geneviève Ephritikhine, Michel Zivy, Carmela Giglione, Thierry Meinnel
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  ABSTRACT: Most proteins in all organisms undergo crucial N-terminal modifications involving N-terminal methionine excision, N-alpha-acetylation or N-myristoylation (N-Myr), or S-palmitoylation. We investigated the occurrence of these poorly annotated but essential modifications in proteomes, focusing on eukaryotes. Experimental data for the N-terminal sequences of animal, fungi, and archaeal proteins, were used to build dedicated predictive modules in a new software. In vitro N-Myr experiments were performed with both plant and animal N-myristoyltransferases, for accurate prediction of the modification. N-terminal modifications from the fully sequenced genome of Arabidopsis thaliana were determined by MS. We identified 105 new modified protein N-termini, which were used to check the accuracy of predictive data. An accuracy of more than 95% was achieved, demonstrating (i) overall conservation of the specificity of the modification machinery in higher eukaryotes and (ii) robustness of the prediction tool. Predictions were made for various proteomes. Proteins that had undergone both N-terminal methionine (Met) cleavage and N-acetylation were found to be strongly overrepresented among the most abundant proteins, in contrast to those retaining their genuine unblocked Met. Here we propose that the nature of the second residue of an ORF is a key marker of the abundance of the mature protein in eukaryotes.
  Proteomics 07/2008; 8(14):2809-31. · 4.43 Impact Factor
• Chapter: Proteomics for genetic and physiological studies in plants

  Hervé Thiellement, Nasser Bahrman, Catherine Damerval, Christophe Plomion, Michel Rossignol, Véronique Santoni, Dominique De Vienne, Michel Zivy
  12/2007: pages 38 - 51; , ISBN: 9783527613489
• Article: Total protein extraction with TCA-acetone.

  Valérie Méchin, Catherine Damerval, Michel Zivy
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  ABSTRACT: We describe a procedure allowing extraction of total proteins that performs efficiently with a large variety of plant tissues, based on simultaneous precipitation and denaturation with TCA and 2ME in cold acetone. We also describe protein solubilization prior to IEF, either in classical rod gels or in IPGs, using two different solutions. The procedure is easy to carry out. The major caveats are (1) keep samples at low temperature during extraction, and then (2) manage protein samples at about 22 to 25 degrees C to avoid urea precipitation.
  Methods in molecular biology (Clifton, N.J.) 02/2007; 355:1-8.
• Article: Quantitative analysis of 2D gels.

  Michel Zivy
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  ABSTRACT: The use of 2D electrophoresis for comparative proteomics allows the revelation of variations of protein relative amounts according to various physiological or genetic criteria. The statistical significance of these results is related to different factors, from the experimental design to the statistical tests. In this chapter we describe different parameters that should be taken into account in the experimental design and during image acquisition, and we present different programs for the normalization of quantitative data, the selection of reproducible spots according to user-defined criteria, and the selection of spots showing qualitative and significant quantitative variations.
  Methods in molecular biology (Clifton, N.J.) 02/2007; 355:175-94.
• Article: The PROTICdb database for 2-DE proteomics.

  Olivier Langella, Michel Zivy, Johann Joets
  [show abstract] [hide abstract]
  ABSTRACT: PROTICdb is a web-based database mainly designed to store and analyze plant proteome data obtained by 2D polyacrylamide gel electrophoresis (2D PAGE) and mass spectrometry (MS). The goals of PROTICdb are (1) to store, track, and query information related to proteomic experiments, i.e., from tissue sampling to protein identification and quantitative measurements; and (2) to integrate information from the user's own expertise and other sources into a knowledge base, used to support data interpretation (e.g., for the determination of allelic variants or products of posttranslational modifications). Data insertion into the relational database of PROTICdb is achieved either by uploading outputs from Mélanie, PDQuest, IM2d, ImageMaster(tm) 2D Platinum v5.0, Progenesis, Sequest, MS-Fit, and Mascot software, or by filling in web forms (experimental design and methods). 2D PAGE-annotated maps can be displayed, queried, and compared through the GelBrowser. Quantitative data can be easily exported in a tabulated format for statistical analyses with any third-party software. PROTICdb is based on the Oracle or the PostgreSQLDataBase Management System (DBMS) and is freely available upon request at http://cms.moulon.inra.fr/content/view/14/44/.
  Methods in molecular biology (Clifton, N.J.) 02/2007; 355:279-303.
• Article: A sub-proteome of Arabidopsis thaliana mature stems trapped on Concanavalin A is enriched in cell wall glycoside hydrolases.

  Zoran Minic, Elisabeth Jamet, Luc Négroni, P Arsene der Garabedian, Michel Zivy, Lise Jouanin
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  ABSTRACT: N-glycosylated proteins were isolated from Arabidopsis thaliana mature stems using affinity chromatography on Concanavalin A Sepharose, separated by 2D-electrophoresis and identified using nanoHPLC-MS/MS and MALDI-TOF MS. 102 glycoproteins were identified. 94% of these proteins were predicted by bioinformatics to be targeted to the secretory pathway and 87% of them were predicted to be localized in the cell wall or at the plasma membrane. 30% of these proteins belong to glycoside hydrolase (GH) families with some of them possibly involved in the hydrolysis of cell wall polysaccharides. The second major class of identified proteins comprises aspartyl and serine proteases. Other proteins are predicted to be oxido-reductases, contain interacting domains, are potentially involved in signalling or have an unknown function. This is, to our knowledge, the first survey of plant cell wall N-glycosylated proteins.
  Journal of Experimental Botany 02/2007; 58(10):2503-12. · 5.36 Impact Factor
• Source

  Available from: Bertrand Hirel

  Article: Two cytosolic glutamine synthetase isoforms of maize are specifically involved in the control of grain production.

  Antoine Martin, Judy Lee, Thomas Kichey, Denise Gerentes, Michel Zivy, Christophe Tatout, Frédéric Dubois, Thierry Balliau, Benoît Valot, Marlène Davanture, [......], María-Begoña Gonzalez-Moro, Linda Bethencourt, Dimah Z Habash, Peter J Lea, Alain Charcosset, Pascual Perez, Alain Murigneux, Hitoshi Sakakibara, Keith J Edwards, Bertrand Hirel
  [show abstract] [hide abstract]
  ABSTRACT: The roles of two cytosolic maize glutamine synthetase isoenzymes (GS1), products of the Gln1-3 and Gln1-4 genes, were investigated by examining the impact of knockout mutations on kernel yield. In the gln1-3 and gln1-4 single mutants and the gln1-3 gln1-4 double mutant, GS mRNA expression was impaired, resulting in reduced GS1 protein and activity. The gln1-4 phenotype displayed reduced kernel size and gln1-3 reduced kernel number, with both phenotypes displayed in gln1-3 gln1-4. However, at maturity, shoot biomass production was not modified in either the single mutants or double mutants, suggesting a specific impact on grain production in both mutants. Asn increased in the leaves of the mutants during grain filling, indicating that it probably accumulates to circumvent ammonium buildup resulting from lower GS1 activity. Phloem sap analysis revealed that unlike Gln, Asn is not efficiently transported to developing kernels, apparently causing reduced kernel production. When Gln1-3 was overexpressed constitutively in leaves, kernel number increased by 30%, providing further evidence that GS1-3 plays a major role in kernel yield. Cytoimmunochemistry and in situ hybridization revealed that GS1-3 is present in mesophyll cells, whereas GS1-4 is specifically localized in the bundle sheath cells. The two GS1 isoenzymes play nonredundant roles with respect to their tissue-specific localization.
  The Plant Cell 12/2006; 18(11):3252-74. · 8.99 Impact Factor
• Article: A mass spectrometric approach to identify arbuscular mycorrhiza-related proteins in root plasma membrane fractions.

  Benoît Valot, Luc Negroni, Michel Zivy, Silvio Gianinazzi, Eliane Dumas-Gaudot
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  ABSTRACT: One of the most important morphological changes occurring in arbuscular mycorrhizal (AM) roots takes place when the plant plasma membrane (PM) invaginates around the fungal arbuscular structures resulting in the periarbuscular membrane formation. To investigate whether AM symbiosis-specific proteins accumulate at this stage, two complementary MS approaches targeting the root PM from the model legume Medicago truncatula were designed. Membrane extracts were first enriched in PM using a discontinuous sucrose gradient method. The resulting PM fractions were further analysed with (i) an automated 2-D LC-MS/MS using a strong cation exchange and RP chromatography, and (ii) SDS-PAGE combined with a systematic LC-MS/MS analysis. Seventy-eight proteins, including hydrophobic ones, were reproducibly identified in the PM fraction from non-inoculated roots, representing the first survey of the M. truncatula root PM proteome. Comparison between non-inoculated and Glomus intraradices-inoculated roots revealed two proteins that differed in the mycorrhizal root PM fraction. They corresponded to an H(+)-ATPase (Mtha1) and a predicted glycosylphosphatidylinositol-anchored blue copper-binding protein (MtBcp1), both potentially located on the periarbuscular membrane. The exact role of MtBcp1 in AM symbiosis remains to be investigated.
  Proteomics 05/2006; 6 Suppl 1:S145-55. · 4.43 Impact Factor