Biomassza-alapú energiák innovációjának genomikai megközelítései

A genomika valódi eszköz a biomassza-alapú energiák innovatív kutatása területén, egyes részterületei (metagenomika, transzkriptomika) ténylegesen alkalmazhatók a biomassza-alapanyag optimalizálása során, illetve a biomassza-konverzió hatékonyságának javításában, ezáltal a megújuló energiatípusok előállítási költségeinek csökkentésében. ------------------------------------------------ Genomics is a useful tool in innovative research into biomass energy. Certain fields of genomics (metagenomics, transcriptomics) have a practical application in biomass optimisation and in improving the efficiency of biomass, and can thus help reduce the costs of renewable energy production.genomika, biomassza, kutatás, innováció, biomass energy, genomics, innovation, research, Research and Development/Tech Change/Emerging Technologies,

Symbiotic plant peptides eliminate Candida albicans both in vitro and in an epithelial infection model and inhibit the proliferation of immortalized human cells.

The increasing number of multidrug-resistant microbes now emerging necessitates the identification of novel antimicrobial agents. Plants produce a great variety of antimicrobial peptides including hundreds of small, nodule-specific cysteine-rich NCR peptides that, in the legume Medicago truncatula, govern the differentiation of endosymbiotic nitrogen fixing bacteria and, in vitro, can display potent antibacterial activities. In this study, the potential candidacidal activity of 19 NCR peptides was investigated. Cationic NCR peptides having an isoelectric point above 9 were efficient in killing Candida albicans, one of the most common fungal pathogens of humans. None of the tested NCR peptides were toxic for immortalized human epithelial cells at concentrations that effectively killed the fungus; however, at higher concentrations, some of them inhibited the division of the cells. Furthermore, the cationic peptides successfully inhibited C. albicans induced human epithelial cell death in an in vitro coculture model. These results highlight the therapeutic potential of cationic NCR peptides in the treatment of candidiasis

Differentiation of Symbiotic Cells and Endosymbionts in Medicago truncatula Nodulation Are Coupled to Two Transcriptome-Switches

The legume plant Medicago truncatula establishes a symbiosis with the nitrogen-fixing bacterium Sinorhizobium meliloti which takes place in root nodules. The formation of nodules employs a complex developmental program involving organogenesis, specific cellular differentiation of the host cells and the endosymbiotic bacteria, called bacteroids, as well as the specific activation of a large number of plant genes. By using a collection of plant and bacterial mutants inducing non-functional, Fix− nodules, we studied the differentiation processes of the symbiotic partners together with the nodule transcriptome, with the aim of unravelling links between cell differentiation and transcriptome activation. Two waves of transcriptional reprogramming involving the repression and the massive induction of hundreds of genes were observed during wild-type nodule formation. The dominant features of this “nodule-specific transcriptome” were the repression of plant defense-related genes, the transient activation of cell cycle and protein synthesis genes at the early stage of nodule development and the activation of the secretory pathway along with a large number of transmembrane and secretory proteins or peptides throughout organogenesis. The fifteen plant and bacterial mutants that were analyzed fell into four major categories. Members of the first category of mutants formed non-functional nodules although they had differentiated nodule cells and bacteroids. This group passed the two transcriptome switch-points similarly to the wild type. The second category, which formed nodules in which the plant cells were differentiated and infected but the bacteroids did not differentiate, passed the first transcriptome switch but not the second one. Nodules in the third category contained infection threads but were devoid of differentiated symbiotic cells and displayed a root-like transcriptome. Nodules in the fourth category were free of bacteria, devoid of differentiated symbiotic cells and also displayed a root-like transcriptome. A correlation thus exists between the differentiation of symbiotic nodule cells and the first wave of nodule specific gene activation and between differentiation of rhizobia to bacteroids and the second transcriptome wave in nodules. The differentiation of symbiotic cells and of bacteroids may therefore constitute signals for the execution of these transcriptome-switches

Protection of Sinorhizobium against Host Cysteine-Rich Antimicrobial Peptides Is Critical for Symbiosis

A bacterial membrane protein, BacA, protects Sinorhizobium meliloti against the antimicrobial activity of host peptides, enabling the peptides to induce bacterial persistence rather than bacterial death

Conserved CDC20 Cell Cycle Functions Are Carried out by Two of the Five Isoforms in Arabidopsis thaliana

The CDC20 and Cdh1/CCS52 proteins are substrate determinants and activators of the Anaphase Promoting Complex/Cyclosome (APC/C) E3 ubiquitin ligase and as such they control the mitotic cell cycle by targeting the degradation of various cell cycle regulators. In yeasts and animals the main CDC20 function is the destruction of securin and mitotic cyclins. Plants have multiple CDC20 gene copies whose functions have not been explored yet. In Arabidopsis thaliana there are five CDC20 isoforms and here we aimed at defining their contribution to cell cycle regulation, substrate selectivity and plant development.Studying the gene structure and phylogeny of plant CDC20s, the expression of the five AtCDC20 gene copies and their interactions with the APC/C subunit APC10, the CCS52 proteins, components of the mitotic checkpoint complex (MCC) and mitotic cyclin substrates, conserved CDC20 functions could be assigned for AtCDC20.1 and AtCDC20.2. The other three intron-less genes were silent and specific for Arabidopsis. We show that AtCDC20.1 and AtCDC20.2 are components of the MCC and interact with mitotic cyclins with unexpected specificity. AtCDC20.1 and AtCDC20.2 are expressed in meristems, organ primordia and AtCDC20.1 also in pollen grains and developing seeds. Knocking down both genes simultaneously by RNAi resulted in severe delay in plant development and male sterility. In these lines, the meristem size was reduced while the cell size and ploidy levels were unaffected indicating that the lower cell number and likely slowdown of the cell cycle are the cause of reduced plant growth.The intron-containing CDC20 gene copies provide conserved and redundant functions for cell cycle progression in plants and are required for meristem maintenance, plant growth and male gametophyte formation. The Arabidopsis-specific intron-less genes are possibly "retrogenes" and have hitherto undefined functions or are pseudogenes

Différenciation des cellules symbiotiques au cours de l interaction Medicago truncatula-Sinorhizobium meliloti (implication de peptides sécrétés d origine végétale)

Les légumineuses s associent avec des bactéries du sol appelées rhizobia qui se différencient en bactéroïdes fixateurs d azote au sein de nodosités racinaires. Durant cette thèse, les mécanismes de différenciation des bactéroïdes ont été étudiés dans différentes interactions Rhizobium-légumineuse. Selon la plante hôte, les bactéroïdes sont semblables à des bactéries libres ou subissent une différenciation poussée comprenant un fort allongement cellulaire corrélé à une endoréplication massive, une peméabilisation membranaire et une perte de capacité à croître. Ce processus est caractéristique de Sinorhizobium meliloti dans les nodosités de Medicago truncatula et des symbiontes des plantes appartenant au Inverted Repeat-Lacking Clade. Au contraire, dans les autres légumineuses, la différenciation des bactéroïdes ne comprend aucune de ces étapes. Il a été montré que la différenciation irréversible des bactéroïdes est sous le contrôle de la plante et est corrélée à l expression de grandes familles de gènes codant des peptides sécrétés spécifiques des nodosités. Parmi eux, les Nodule-specific Cysteine-Rich et les Glycine-Rich Protein comprennent respectivement plus de 300 et 20 membres. Ces gènes sont exclusivement exprimés dans les cellules infectées des IRLC. Des études génomique et évolutive ont révélé que ces gènes sont regroupés dans le génome de M. truncatula et évoluent par duplication locale. Une étude de microsynténie a démontré qu ils sont absents du génome de Lotus japonicus. Une étude fonctionnelle a été initiée et indique que les NCRs sont adressés aux bactéroïdes. Finalement, ces résultats ont été discutés dans le contexte de l évolution des endosymbioses.Legume plants associate with soil bacteria called rhizobia that differentiate into nitrogen-fixing bacteroids in a symbiotic organ: the root nodule. During this thesis, the mechanisms of bacteroid differentiation have been studied in different Rhizobium-legume interactions. Depending on the host plant, bacteroids were either similar to cultured bacteria or showed striking differentiation comprising cell enlargement, massive endoreduplication, membrane permeabilization and loss of growth capacity. Such a differentiation is observed for Sinorhizobium meliloti in Medicago truncatula nodules, and for those rhizobia that interact with plants related to Medicago and belonging to the Inverted Repeat-Lacking Clade (IRLC). In the non-IRLC legumes, bacteroids do not show any of these features and bacteroids are similar to free-living bacteria. It has been demonstrated that the terminal differentiation of bacteroids is under plant control and correlated to the evolution and expression of large gene families encoding nodule-specific secreted peptides resembling antimicrobial peptides. Among them are the Nodule-specific Cysteine-Rich and the Glycine-Rich Protein families, which comprise more than 300 and 20 members respectively. They are exclusively expressed in infected cells of IRLC legumes. Genomic and evolutionary studies revealed that these genes are clustered in M. truncatula genome and evolve by local duplication. A microsynteny study showed that they are absent from Lotus japonicus genome. Functional studies of these genes have been initiated and indicate that NCRs are targeted to the bacteroids. These new results have been discussed in the frame of endosymbioses evolution.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

Analyse fonctionnelle des gènes précoces impliqués dans la symbiose fixatrice d'azote entre Medicago truncatula et Sinorhizobium meliloti

Les légumineuses utilisent une voie de signalisation commune pour établir l association symbiotique avec les rhizobiums et les champignons endomycorhiziens. La perception du microorganisme a lieu à la membrane plasmique et active une cascade de transduction du signal jusqu au noyau pour la transcription de gènes spécifiques. Le gène NORK chez les espèces de Medicago code pour un récepteur kinase-like, composant de la voie de signalisation des facteurs Nod et essentiel pour le développement de la nodosité. Pour comprendre le rôle de NORK, nous avons cherché des protéines partenaires agissant en aval par double hybride et nous avons identifié une 3-hydroxy-3-méthylglutaryl Coenzyme A réductase (MtHMGR1). MtHMGR1 fait partie d une famille multigénique chez M. truncatula contrôlant la biosynthèse de mévalonate et en testant d autres membres nous avons trouvé une spécificité pour cette unique protéine. La répression spécifique de l expression de MtHMGR1 entraîne une inhibition de la nodulation. Ainsi, MtHMGR1 est un nouveau composant de la voie de signalisation symbiotique qui pourrait contrôler la production de cytokine et la biosynthèse de stérols. Des études surd autres mutants, présentant le même phénotype, ont révélé l existence du gène DM11 codant un canal ionique potentiel. Alors que ses homologues chez L. japonicus, CASTOR et POLLUX, sont localisés dans les plastres, nous avons localisé la protéine de fusion DM11 :GFP, fonctionnelle, à la membrane nucléaire des cellules de racines de M.truncatula lorsque elle est exprimée sous promoteur 35S ou natif. Il se peut que la région N-terminal de DM11 soit en partie responsable de sa localisation. Ces nouvelles informations placent DM11 dans l enveloppe nucléaire, proche du lieu d origine des oscillations de la concentration en calcium cytoplasmique induits par les facteurs Nod.Legumes utilize a common signaling pathway to form symbiotic associations both with rhizobial bacteria and arbuscular mycorrhizal fungi. The microbe perception is believe to take place at the plasma membrane, activating a cascade that converges on the nucleus where transcriptional reprogramming takes place. The NORK gene in Medicago species encodes a receptor-like kinase, component of the Nod signal transduction pathway and essential for root nodule development. NORK is required for the generation of nucleus associated calcium spikes in response to Nod factors as well as for infection of plant cells with rhizobia. To understand the mode of NORK action, we searched for its downstream targets by yeast two-hybrid screens. We identified 3-hydroxy-3-methylglutaryl Coenzyme A reductase (MtHMGR1) as a interacting partner of NORK. MtHMGR1 belong to a multigene family in M.truncatula whose members are key component of the mevalonate pathway. Testing other members, the interaction specificity was revealed for MtHMGR1. Downregulation of MtHMGR1 expression affect dramatically the nodulation process. Thus, MtHMGR1 is a novel component of the symbiotic signaling pathway that could control biochemical pathway such as cytokinin and sterol synthesis for protein isoprenylation. Genetics studies on other mutants with same phenotypes have identified in M truncatula DM11 gene that encodes a putative ion channel. While the DM11 homologs from Lotus japonicus, XASTOR and POLLUX, were recently reported to localize in plastids, we report here that a functional DM11:GFP fusion localizes to the nuclear envelope in M.truncatula roots when espressed both a constitutive 35S promoter and from a native DM11 promoter. Localization may be mediated in part by sequences located within the amino-terminus of DM11. These new data place DM11 in the nuclear envelope in close proximity to the origin of Nod factor induced calcium spiking.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

le rôle des activateurs APC/C dans la croissance et le développement chez arabidopsis thaliana

La protéolyse médiée par ubiquitinylation régule de nombreux processus cellulaires. Les protéines cibles sont dans un premier temps polyubiquitinylées, pour être ensuite dégradées. Le cycle cellulaire est contrôlé par deux Ubiquitine ligases, appelées complexes SCF et APC/C dont les actions consécutives et complémentaires contrôlent sa dynamique en dégradant une variété de régulateurs protéiques de ce cycle cellulaire. Ce travail de thèse est centré sur les implications de l APC/C dans le cycle cellulaire et dans un contexte développemental chez Arabidopsis thaliana, notamment en étudiant des activateurs contrôlant l activité temporelle et la spécificité de substrat du complexe APC/C, CDC20 et CDH1, connue chez les plantes sous le nom de CCS52. De façon différente des autres organismes, Arabidopsis thaliana possède plusieurs isoformes activatrices de ce complexe : cinq CDC20s et trois CCS52s. Ce travail a eu pour objectif de caractériser fonctionnellement les isoformes de CDC20 ainsi que CCS52A1 et CCS52A2.Ubiquitin Médiated Proteolysis (UMP) regulates many cellular processes by polyubiquitination and consequent degradation of target proteins. The cell cycle is controlled by two ubiquitin ligases :the Skp, Cullin, F-box containing (SCF) and Anaphase-Promoting Complex/ Cyclosome (APC/C) who s complementary and consecutive actions drive the cell cycle by degradation of various cell cycle regulators. This thesis is focused on the cell and developmental role of the APC/C in Arabidopsis thaliana via studying its activators that control the temporal activity and substrate specificity of the APC/C. These are CDC20 and CDH1, the latter described in plants as CCS52. Unlike most organisms, Arabidopsis possesses multiple activator isoforms : five CDC20s and three CCS52s. This work aimed at exploring the functions of the CDC20 isoforms and that CCS52A1 and CCS52A2.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF