One of the key processes in the carbon cycle on our planet is the degradation of dissolved organic matter (DOM) in aquatic environments. The use of organic matter by bacteria links energy from DOM to higher trophic levels of the ecosystem when bacteria are consumed by other organisms. This is referred to as the microbial loop. In this thesis I examined if the communities were functionally redundant in their ability to utilize organic matter, or if variation in bacterial composition and richness is of importance. To test this overarching question several experiments were conducted that include methods such as illumina sequencing of the 16S rRNA gene for taxonomic identification of bacterial communities, flow cytometry to follow the growth of communities and spectroscopic measurement to describe the composition of the organic matter pool. Initially we demonstrated how to optimally sterilize organic matter for experimental studies in order to preserve its natural complexity. In further experiments we found that bacterial communities are redundant in their utilization of organic matter and can maintain optimal performance towards a range of organic matter pools. Related to this we found that pre-adaptation to organic matter played a small role as communities performed equally well regardless of their environmental history. We saw a small effect of richness and composition of bacterial communities on the efficiency of organic matter use, but conclude that this is of minor importance relative to abiotic factors. Still, we also show that organic matter can put strong selection pressure on bacterial communities with regards to richness and composition. Additionally we found that the supply rate of a carbon compound greatly influenced the energy utilization of the compound, i.e. a higher growth rate can be maintained if substrate is delivered in pulses relative to a continuous flow. Finally we conclude that the variation in bacterial communities is unlikely to have a major influence on carbon cycling in boreal lakes, but to enable a finer understanding, the genetics underlying the carbon utilization needs to be further explored.

La productivité des processus d'usinage robotisé est très souvent limitée par le manque de rigidité des robots et les problèmes vibratoires relatifs à l’instabilité de la coupe. L’analyse de l’instabilité de la coupe en usinage robotisé est un problème difficile en raison de la variabilité du comportement dynamique du robot dans son espace de travail. Par conséquent, le phénomène de broutement en usinage robotisé dépend non seulement des paramètres de coupe mais également de la configuration du robot. Le premier objectif de cette thèse est de déterminer une méthode de modélisation dynamique du robot, adaptée du point de vue de l’analyse des vibrations et de la stabilité en usinage robotisé. Cette approche a été réalisée sur le robot d’usinage industriel ABB IRB6660. Une démarche de recalage a été mise en place afin de déterminer les paramètres du modèle numérique du robot. Ensuite, une présentation 3D de la limite de stabilité en usinage robotisé prenant en compte les variations du comportement dynamique du robot est réalisée. Le deuxième objectif consiste à optimiser le procédé vis-à-vis de la stabilité de la coupe. Les variations du comportement dynamique du robot sont exploitées par la gestion des redondances fonctionnelles afin d’optimiser la configuration du robot du point de vue de la stabilité. L’analyse numérique a montré et les essais expérimentaux d’usinage ont confirmé la possibilité de passer de la zone instable à la zone stable par la gestion de la redondance fonctionnelle sans modifier les paramètres de coupe. / Productivity in robotic machining processes can be limited by the low rigidity of the overall structure and vibration instability (chatter). The robot’s dynamic behavior, due to changes in its posture along a machining trajectory, varies within its workspace. Chatter in robotic machining therefore depends not only on the cutting parameters but also on the robot configuration. The first objective of this thesis is to determine a dynamic modeling approach of the robot in order to analyze the vibration and the stability in robotic machining. This modeling approach has been realized to dynamic modeling of an ABB IRB6660 industrial robot. The numerical model parameters are adjusted on the basis of experimental modal identifications. Then, a three-dimensional representation of stability lobes diagram for the prediction to take into account the robot dynamic behavior variations in machining trajectory is established. The second objective is to optimize the robot configurations regarding stability. The dynamic behavior variations of the robot in the workspace are exploited through functional redundancy management in order to optimize robot configurations with respect to machining stability. The numerical analyze demonstrated and experimental machining tests confirmed that stability conditions in machining operations can be achieved by managing functional redundancy without changing the cutting parameters.

Land-use change is the single most important global driver of changes in biodiversity. Such changes in biodiversity, in turn, are expected to influence the functioning of ecosystems and their resilience to environmental perturbations and disturbances. It is widely recognised that the use of functional traits and functional diversity is best for understanding the causes and functional consequences of changes in biodiversity, but conceptual development has outpaced empirical applications. This thesis explores these ideas in grazing systems, which are expected to undergo rapid intensification of fertiliser use and grazing pressure to meet the growing global demand for livestock products. First, a flexible framework for measuring different facets of functional diversity is described, and a new multidimensional functional diversity index, called functional dispersion (FDis), is presented. Second, two vegetation sampling methods are compared with regard to their ability to detect changes in vegetation composition. Third, shifts in plant trait distributions following land-use changes are quantified and compared to null models, and a maximum entropy approach is used to quantify the direction and strength of selection on each trait. Fourth, it is shown that these shifts in trait distributions have cascading effects on primary production, litter decomposition, soil respiration, and ultimately soil carbon sequestration. Finally, data from 18 land-use intensity gradients are used to show that land-use intensification reduces functional redundancy and response diversity, two components of biodiversity that are thought to influence ecosystem resilience to future disturbances. This study illustrates (i) the importance of considering species functional differences to understand how plant communities react to changes in soil resource availability and grazing pressure, and (ii) how such changes directly, indirectly, and interactively control ecosystem functioning, as well as (iii) increasing the vulnerability of ecosystems to future disturbances.

Le drainage minier acide de Carnoulès est caractérisé par un pH très acide et une forte concentration en métaux et en arsenic. Par différentes approches moléculaires, des études précédentes ont montré une faible biodiversité et ont permis d'établir un modèle de fonctionnement de la communauté bactérienne. Le but de ce travail de thèse a été de préciser la composition et le fonctionnement de cette communauté bactérienne, en utilisant pour cela des approches culturales et fonctionnelles, en se focalisant particulièrement sur le recyclage de la matière organique. L'élaboration de différents milieux a permis l'isolement de 49 souches bactériennes appartenant à 19 genres, augmentant ainsi de 10 % la diversité bactérienne détectée à Carnoulès par rapport aux approches métagénomiques précédentes. Parmi les 19 genres, 3 sont nouveaux dont un, inféodé aux écosystèmes acides, a été caractérisé taxonomiquement et dénommé Acidiminas carnoulesii. La capacité de l'isolat Q8 appartenant au genre Paenibacillus à dégrader I'amidon et la xylane, dans de larges gammes de pH et de concentrations en arsenic, a permis d'attribuer à Paenibacillus un rôle dans la résilience de la communauté pour ces fonctions. Un criblage fonctionnel de I'ADN de Q8 dans Escherichia coli apermis d'isoler les gènes codant les protéines de dégradation de ces polymères. Par ailleurs, un criblage de 80000 clones de la banque d'ADN métagénomique de Carnoulès a permis la détection de 28 clones positifs pour l'activité amylolytique. Deux protéines ne présentant aucune similarité de séquence avec des amylases connues ont été caractérisées in vitro, confirmant leurs activités amylolyiques et démontrant que la bioprospection dans des sites a priori incongrus, autorise des découvertes insoupçonnées. Ces travaux ont ainsi permis de montrer que les approches culturales et fonctionnelles apportent des informations nouvelles par rapport à celles obtenues par les approches moléculaires. La complémentarité de ces approches est vérifiée, et elle apparaît indispensable dans l'analyse de la complexité des écosystèmes. Cependant, la compréhension de leur fonctionnement exigera des efforts redoublés. / The Carnoulès acid mine drainage is characterized by an acidic pH and high metal and arsenic concentrations. Based on the low bacterial biodiversity, molecular approaches allowed the determination of a bacterial community functioning model. The aim of the PhD work was to clariff both the composition andthe functioning of this community, using cultural and functional approaches, focusing on the organic matter recycling. Different media were designed that allowed the isolation of 49 bacterial strains belonging to 19 genera,leading to a 70 o  increase in the bacterial diversity compared with previous metagenomic approaches. Among the 19 genera,3 are new, one of which, a previously uncultured genus frequently detected in acidic environments, has been taxonomically characterized and named Acidiminos cornoulesii.The ability of strain Q8, belonging to the genus Paenibacillus, to degrade starch and xylan over a wide rangeof pH values as well as 4rsenic concentrations allows to assign to Paenibacillus a role in the resilience of the community for these functions. A function-based screening of the Q8-DNA in Escherichia coli led to isolatethe genes encoding the polymer-degrading proteins.Moreover, a function-based screening of 80,000 clones from a Carnoulès metagenomic DNA library led to the detection of 28 positive clones for the amylolyic activity. Two proteins sharing no sequence similarity with known amylases were characterized in vitro, demonstrating that bioprospecting in a priori incongruous sites can lead to unsuspected discoveries.These works show that new informations using cultural and functional approaches can be obtained compared to those gained with molecular approaches. Results confirmed the complementarity between both approachesis crucial to analyze complex ecosystems. Nevertheless, more research efforts still have to be undertaken tounderstand their functioning.

Arabidopsis thaliana, s’est adaptée à la variété des niveaux de Pi dans le sol en développant 9transporteurs de phosphates, membres de la famille PHT1, intervenant dans l’acquisition de cet ionpar les racines et sa translocation. Ces protéines révèlent une très forte homologie de séquence entre elles (plus que 61%). La présence de certains transporteurs de type PHT1 dans différents types d’organes ainsi que le chevauchement fréquent entre les divers membres de la famille PHT1 témoigne de la complexité de leurs rôles. De plus, leur redondance génétique et fonctionnelle empêche l’analyse de leur rôle spécifique. En vue de s’affranchir de ces obstacles, notre approche combine plusieurs stratégies génétiques avec l’insertion d’une construction RNAi inactivant plusieurs membres de la famille PHT1 et en particulier le cluster localisé sur le chromosome 5 (PHT1;1/1;2/1;3). Ces outils génétiques ont révélé aussi le fonctionnement des protéines PHT1 à la fois en tant que transporteurs à basse et à haute affinité, ce qui suggère que leur activité est contrôlée au niveau post-traductionnel. En cas de carence en Pi, ces lignées affichent des modifications physiologiques (biomasse, rendement,…) dues à une forte réduction affectant l’activité de l’influx en phosphate (80 à 96%). Ce travail suggère que la redondance génétique et les mécanismes de compensations pourraient protéger la plante de l’inactivation de PHT1. Il a aussi révélé que la perception systémique du Pi est déclenchée par des mécanismes en aval de l’activité des PHT1. / Arabidopsis thaliana absorb inorganic phosphate (Pi) from the soil through an active transport process mediated by the 9 members of the PHT1 family. These proteins share a high level of similarity (greater than 61%), with overlapping expression patterns. The resulting genetic and functional redundancy prevents the analysis of their specific roles. To overcome this difficulty, our approach combined several mutations with gene silencing to inactivate multiple members of the PHT1 family, including a cluster of genes localized on chromosome 5 (PHT1;1, PHT1;2 and PHT1;3). Physiological analyses of these lines established that these three genes, along with PHT1;4, are the main contributors to Pi uptake. Furthermore, PHT1;1 plays an important role in translocation from roots to leaves in high phosphate conditions. These genetic tools also revealed that some PHT1 transporters likely exhibit a dual affinity for phosphate, suggesting that their activity is posttranslationally controlled. These lines display significant phosphate deficiency-related phenotypes (e.g. biomass and yield) due to a massive (80 to 96%) reduction in phosphate uptake activities. These defects limited the amount of internal Pi pool, inducing compensatory mechanisms triggered by the systemic Pi starvation response. Such reactions have been uncoupled from PHT1 activity suggesting that systemic Pi sensing is most probably acting downstream of PHT1.

Les légumineuses sont capables de s’associer en symbiose avec des bactéries du sol Rhizobium. Cette interaction culmine par la formation d’un nouvel organe racinaire appelé nodule, à l’intérieur duquel les bactéries différentiées fixent l’azote atmosphérique sous une forme assimilable par la plante hôte. La mise en place de cette association repose sur un dialogue moléculaire entre les deux partenaires, faisant intervenir des signaux bactériens lipo-chitooligosaccharidiques appelés Facteurs Nod (FNods). Chez la légumineuse modèle Medicago truncatula, la perception de ces signaux symbiotiques au niveau de l’épiderme racinaire, initie une voie de signalisation qui conduit à des oscillations calciques nécessaires pour l`activation de gènes de la plante hôte, tel le gène marqueur ENOD11. Il a été montré que les facteurs de transcription ERF ERN1/ERN2, étroitement apparentés, agissent comme des activateurs directs de la transcription d’ENOD11, via leur liaison à la séquence cis régulatrice NFbox. Le mutant ern1 est de manière cohérente requis pour l’activation d’ENOD11 en réponse aux FNods mais également au cours des étapes suivantes d’infection et de développement nodulaire. Cependant, ce mutant présente un phénotype symbiotique partiel soulevant la question d’une redondance fonctionnelle, qui pourrait être attribuée à la présence du facteur ERN2, étroitement apparenté. Ainsi, au cours de ma thèse, j’ai étudié la relation fonctionnelle entre les facteurs ERN1/ERN2 par de approches diverses visant à déterminer leur expression et fonctions relatives au cours de la symbiose rhizobienne. Mon travail de thèse a dans tout d’abord porté sur l’étude des profils d’expression spatio-temporels de gènes ERN au cours de la symbiose rhizobienne, corroborée par la dynamique de localisation cellulaire des protéines de fusions ERN. Ces facteurs sont exprimés de manière séquentielle mais aussi conjointe aux cours de la signalisation FNods et l’infection rhizobienne. Par la suite, des expériences de complémentation croisée, dans le fond mutant ern1, ont montré qu’ERN2 peut remplacer ERN1 pour l’induction d’ENOD11 en réponse aux FNods et pour la formation de nodules, dès lors qu’il est exprimé sous le contrôle du promoteur d’ERN1. Ceci indique que ces facteurs ont des activités biologiques similaires et suggère que l’absence de complémentation d’ern1 par le facteur endogène ERN2 est probablement liée à une régulation transcriptionnelle différentielle de la part de leurs promoteurs. Enfin, nous avons initié la caractérisation phénotypique de nouvelles lignées mutées au niveau du gène ERN2, dans le but d’identifier les fonctions spécifiques de ce facteur au cours de la nodulation. A travers l’analyse d’une lignée Tilling (ern2.1) présentant une mutation ponctuelle dans le domaine de liaison à l’ADN, nous avons mis en évidence un rôle d’ERN2 dans la progression des cordons d’infection au niveau du cortex racinaire. Des études moléculaires ont permis de montrer que l’acide aminé muté est un résidu conservé et important pour la topologie du domaine de liaison à l’ADN, mais également pour l’activité transcriptionnelle d’ERN2 sur ENOD11. Contrairement à ern1, le mutants ern2.1 et ern2.2 (mutant d’insertion) sont capables de former des nodules. Néanmoins, l’infection nodulaire apparait dans les deux cas parfois défectueuse, conduisant à une sénescence précoce. Ces résultats démontrent qu’ERN2 remplit aussi des rôles spécifiques au cours de la nodulation, qui ne sont pas entièrement complémentés par ERN1. Il semblerait donc que les facteurs ERN contrôlent des étapes communes et divergentes de l‘infection rhizobienne, ERN1 ayant un rôle prépondérant dans l`initiation et progression de l’infection tandis qu’ERN2 aurait un rôle secondaire, plus centré dans la progression des cordons. La lignée double mutant ern1ern2.1, ouvre de nouvelles perspectives pour l’étude de la redondance fonctionnelle entre ces deux facteurs au cours des symbioses racinaires / Legumes are able to associate in symbiosis with Rhizobia bacteria in the soil, which culminates in the formation of a new organ referred to as the root nodule, within which differentiated bacteria fix nitrogen to the benefit of the host plant. The establishment of this association relies on a molecular dialogue between the two partners, involving bacterial lipo-chitooligosaccharide signals called Nod factors (NF). In the model legume Medicago truncatula, the perception of these symbiotic signals in the root epidermis, initiates a signaling pathway that leads to calcium oscillation responses required for the activation of downstream genes such as the well-characterized ENOD11. Previously, ERN1 and the closely-related ERN2 transcription factors (TFs) were reported as direct activators of ENOD11 via binding to the NFbox regulatory unit. In addition, phenotypic analysis of the ern1 knockout mutant has confirmed the importance of ERN1 not only during NF signaling but also throughout subsequent infection and nodule development stages. Nevertheless, the ern1 mutant displays a less severe phenotype compared to plants mutated in other NF signaling genes, raising the question of a possible functional redundancy with the endogenous closely-related ERN2 factor. My PhD project was focused on the study of the functional relationship between ERN1 and ERN2 TFs. By using a variety of strategies we aimed at determining both ERN expression profiles and relative functions during nodulation. We first examined the spatio-temporal expression profiles of these genes during rhizobial symbiosis and correlated this with the dynamics of cellular localization of ERN fusion proteins. These analyses revealed that these factors possess both common and distinct expression profiles, correlated with cell-type specific and dynamic in vivo protein accumulation, tightly associated with rhizobial pre-infection and subsequent infection stages in M. truncatula. Further cross-complementation studies in the ern1 mutant background showed that, when ERN2 is expressed under the control of the ERN1 promoter, it can fully restore the ern1 phenotype regarding NF-elicited ENOD11 activation and nodule formation. This indicates that these factors have similar biological activities and suggests that the incapacity of endogenous ERN2 to complement the ern1 mutant is mainly due to differences in their promoter activities. Finally, we also initiated a phenotypic characterization of M. truncatula ern2 mutant lines, in order to get a better insight into ERN2 specific functions during nodulation. The phenotypic analysis of a Tilling line (ern2.1) carrying a point mutation in a conserved amino acid in the ERN2 DNA binding domain, revealed a role for ERN2 during infection thread progression in the root cortex. Further molecular studies demonstrated that this mutated amino acid in the Tilling line is conserved and required for optimal DNA binding domain topology and transcriptional activity of ERN2 on its target ENOD11 gene. In addition, the ern2.1 line and a second ern2.2 insertional mutant line are both capable of forming nodules, in contrast to the ern1 mutant. Nevertheless, these nodules are partly infection defective leading to premature senescence. These findings provide evidence that ERN2 possesses specialized functions during nodulation that cannot be fully complemented by ERN1. This suggests that ERN possess common and divergent functions, ERN1 having a predominant role in rhizobial infection initiation and progression while ERN2 having a secondary and more centered role during infection thread progression. The ern1ern2.1 double mutant line, recently generated during my PhD, opens new perspectives to further study the functional relationship between ERN TFs during root endosymbioses.

Caenorhabditis elegans possui quatro genes de kpcs (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Em C. elegans, dois dos quatro polipeptídeos de vitelogenina encontrados dentro dos ovócitos, YP115 e YP88, se originam a partir de um precursor polipeptídico (VIT-6) clivado pós-traducionalmente após o motivo RGKR. Nematóides transgênicos foram produzidos com construções repórteres transcricionais de GFP. Foi verificada expressão de kpc-1 tanto em neurônios quanto em células musculares e intestinais. Esses dados, aliados aos dados da literatura para os outros genes kpc de C. elegans, sugerem o envolvimento de KPC-1 no processamento de VIT-6, que é secretada por células intestinais. Ensaios de Western-blot compararam o processamento de VIT-6 em nematóides selvagens, mutantes e knock-down por RNAi para os diferentes genes kpc. A análise de nematóides mutantes e knock-down por RNAi combinado para os outros três genes de convertase de C. elegans confirmou a redundância da atividade dessas enzimas no processamento de VIT-6. / Four kpc genes are found in the Caenorhabditis elegans genome: (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Two of the four vitellogenin polypeptides, YP115 and YP88, originate from a precursor, VIT-6. VIT-6 is cleaved post-translationally after the RGKR motif. Transgenic worms carrying GFP transcription reporter constructs were produced. Expression of kpc-1 has been localized to neurons as well as muscular and intestinal cells. These data, together with the ones available from the literature for the other kpc genes, suggest the involvement of KPC-1 in the processing of VIT-6, which is secreted from intestinal cells. Western-blot analysis compared the pattern of VIT-6 processing in wild-type, mutants and RNAi-treated worms for the other kpcs. Analysis of worms treated by combined RNAi confirmed the redundancy of KPCs in VIT-6 processing.

Caenorhabditis elegans possui quatro genes de kpcs (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Em C. elegans, dois dos quatro polipeptídeos de vitelogenina encontrados dentro dos ovócitos, YP115 e YP88, se originam a partir de um precursor polipeptídico (VIT-6) clivado pós-traducionalmente após o motivo RGKR. Nematóides transgênicos foram produzidos com construções repórteres transcricionais de GFP. Foi verificada expressão de kpc-1 tanto em neurônios quanto em células musculares e intestinais. Esses dados, aliados aos dados da literatura para os outros genes kpc de C. elegans, sugerem o envolvimento de KPC-1 no processamento de VIT-6, que é secretada por células intestinais. Ensaios de Western-blot compararam o processamento de VIT-6 em nematóides selvagens, mutantes e knock-down por RNAi para os diferentes genes kpc. A análise de nematóides mutantes e knock-down por RNAi combinado para os outros três genes de convertase de C. elegans confirmou a redundância da atividade dessas enzimas no processamento de VIT-6. / Four kpc genes are found in the Caenorhabditis elegans genome: (kex2/subtilisin-like proprotein convertases): kpc-1, kpc-2/egl-3, kpc-3/aex-5, kpc-4/bli-4. Two of the four vitellogenin polypeptides, YP115 and YP88, originate from a precursor, VIT-6. VIT-6 is cleaved post-translationally after the RGKR motif. Transgenic worms carrying GFP transcription reporter constructs were produced. Expression of kpc-1 has been localized to neurons as well as muscular and intestinal cells. These data, together with the ones available from the literature for the other kpc genes, suggest the involvement of KPC-1 in the processing of VIT-6, which is secreted from intestinal cells. Western-blot analysis compared the pattern of VIT-6 processing in wild-type, mutants and RNAi-treated worms for the other kpcs. Analysis of worms treated by combined RNAi confirmed the redundancy of KPCs in VIT-6 processing.

L’Adrénoleucodystrophie liée à l’X est une maladie neurodégénérative rare due à des mutations dans le gène ABCD1. Ce gène code un demi-transporteur ABC peroxysomal, impliqué dans l’importation d’acides gras à très longue chaîne. Deux autres demi-transporteurs sont localisés dans la membrane peroxysomale : ABCD2 et ABCD3. La surexpression d’ABCD2 permet de compenser la déficience en ABCD1, ouvrant ainsi des perspectives thérapeutiques. Dans cette optique, l’objectif principal de ma thèse était d’étudier la fonction et la structure d’ABCD2, et plus largement des transporteurs ABC peroxysomaux.Les demi-transporteurs doivent au minimum se dimériser pour constituer un transporteur fonctionnel. Leur dimérisation alternative pourrait moduler leur spécificité de substrat. Afin de tester cette hypothèse, nous avons réalisé des constructions plasmidiques codant différents dimères chimériques, dont la fonctionnalité a été vérifiée par transfection transitoire dans deux modèles cellulaires (fibroblastes humains et levures). D’après nos résultats, ABCD1 et ABCD2 seraient fonctionnels quel que soit leur agencement dimérique. De plus, comme d’autres transporteurs ABC, les transporteurs ABC peroxysomaux pourraient s’oligomériser. En utilisant différentes techniques biochimiques (co-immunoprécipitation, sédimentation sur gradient de sucrose et électrophorèse en conditions natives), sur un modèle cellulaire surexprimant ABCD2-EGFP, nous démontrons qu’ABCD2-EGFP interagit avec ABCD1 et ABCD3, et que les transporteurs ABC peroxysomaux sont capables de s’oligomériser. Il reste désormais à déterminer les facteurs qui contrôlent cette oligomérisation et comprendre la valeur fonctionnelle de ces interactions. / X-linked Adrenoleukodystrophy (X-ALD) is a rare neurodegenerative disease caused by deficiency of the peroxisomal half-transporter ABCD1, implicated in very long chain fatty acids import. Two additional half-transporters are located in the peroxisomal membrane: ABCD2 and ABCD3. Over-expression of ABCD2 is known to compensate for ABCD1 deficiency, making ABCD2 a therapeutic target for X-ALD treatment. In this context, the main objective of my thesis was to investigate the function and the structure of ABCD2, and more broadly, of peroxisomal ABC transporters.Half-transporters must at least dimerize to form a functional transporter. Alternative dimerization could modulate substrate specificity. In order to test this hypothesis, we engineered plasmidic constructs encoding chimeric ABCD dimers, whose functionality has been evaluated by transient transfection in two cell models (human fibroblasts and yeasts). Our results show that, ABCD1 and ABCD2 are functional whatever their dimeric organization. Besides, like other ABC transporters, peroxisomal ABC transporters could oligomerize. By using a multi-technical approach (co-immunoprecipitation, velocity sucrose gradient and native polyacrylamide gel electrophoresis experiments) on stably transfected hepatoma cells expressing ABCD2-EGFP, we demonstrate that ABCD2-EGFP interacts with ABCD1 and ABCD3, and that peroxisomal ABC transporters oligomerize. The perspectives will consist in determining which factors control the oligomerization process and understanding the functional value of these interactions.

Made available in DSpace on 2014-12-17T14:33:06Z (GMT). No. of bitstreams: 1 IgorGB_DISSERT.pdf: 695705 bytes, checksum: 0611f2165b1e05afa1e64ceaedeacebb (MD5) Previous issue date: 2012-08-27 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / High levels of local, regional, and global extinctions has progressively simplified communities in terms of both species and ecosystem functioning. Theoretical models demonstrated that the degree of functional redundancy determines the rates of functional group loss in response to species extinctions. Here, we improve the theoretical predictions by incorporating in the model interactions between species and between functional groups. In this study, we tested the effect of different scenarios of interspecific interactions and effects between functional groups on the resistance to loss of community functional groups. Virtual communities have been built with different distribution patterns of species in functional groups, both with high and low evenness. A matrix A was created to represent the net effect of interspecific interactions among all species, representing nesting patterns, modularity, sensitive species, and dominant species. Moreover, a second matrix B was created to represent the interactions between functional groups, also exhibiting different patterns. The extinction probability of each species was calculated based on community species richness and by the intensity of the interspecific interactions that act upon it and group to which it belongs. In the model, successive extinctions decrease the community species richness, the degree of functional redundancy and, consequently, the number of functional groups that remain in the system. For each scenario of functional redundancy, A, and B, we ran 1000 simulations to generate an average functional extinction curve. Different model assumptions were able to generate remarkable variation on functional extinction curves. More extreme variations occurred when the matrix A and B caused a higher heterogeneity in the species extinction probability. Scenarios with sensitive species, positive or negative, showed a greater variation than the scenarios with dominant species. Nested interactions showed greater variation than scenarios where the interactions were in modules. Communities with maximal functional evenness can only be destabilized by the interactions between species and functional groups. In contrast, communities with low functional evenness can have its resistance either increased or decreased by the interactions. The concentration of positive interactions in low redundancy groups or negative interactions in high redundancy groups was able to decrease the functional extinction rates. In contrast, the concentration of negative interactions in low redundancy groups or positive interactions in high redundancy groups was able to increase the functional extinction rates. This model shows results that are relevant for species priorization in ecosystem conservation and restoration / Os n?veis elevados de extin??es locais, regionais e globais t?m simplificado progressivamente comunidades em termos de esp?cies e funcionamento do ecossistema. Modelos te?ricos demonstraram que o grau de redund?ncia funcional determina as taxas de perda de grupos funcionais ? medida que as comunidades sofrem extin??o de esp?cies. Aqui n?s aprimoramos as predi??es te?ricas pela incorpora??o no modelo de intera??es entre esp?cies e entre grupos funcionais. Neste estudo, testamos o efeito de diferentes cen?rios de intera??es interespec?ficas e de efeitos entre grupos funcionais sobre a resist?ncia das comunidades ? perda de grupos funcionais. Comunidades virtuais foram constru?das com diferentes padr?es de distribui??o de esp?cies nos grupos funcionais, tanto com alta quanto com baixa equitabilidade. Uma matriz A foi criada para representar o efeito l?quido das intera??es interespec?ficas entre todas as esp?cies, representando padr?es de aninhamento, modularidade, esp?cies sens?veis ou dominantes. Al?m disto, uma segunda matriz B foi criada para representar as intera??es entre grupos funcionais, tendo tamb?m diferentes padr?es. A probabilidade de extin??o de cada esp?cie foi calculada com base na riqueza de esp?cie da comunidade e pela intensidade das intera??es interespec?ficas que atuam sobre ela e sobre o grupo funcional ao qual ela pertence. No modelo, extin??es de esp?cies sucessivas diminuem a riqueza da comunidade, o grau de redund?ncia funcional e consequentemente o n?mero de grupos funcionais que permanecem no sistema. Para cada cen?rio de redund?ncia funcional, A e B, n?s rodamos 1000 simula??es para gerar uma curva de extin??o funcional m?dia. Diferentes suposi??es do modelo foram capazes de gerar varia??es not?veis nas curvas de extin??o funcional. Varia??es mais extremas ocorreram quando as matrizes A e B definem um efeito diferencial acentuado na probabilidade de extin??o das esp?cies dos grupos funcionais. Cen?rios com esp?cies sens?veis, positivas ou negativas, apresentaram uma maior varia??o que os cen?rios com esp?cies dominantes. Intera??es aninhadas apresentaram maior varia??o do que cen?rios em que as intera??es s?o modulares. Comunidades com redund?ncia funcional m?xima podem somente ser fragilizadas pelas intera??es entre esp?cies e grupos funcionais. Em contraste, comunidades com baixa riqueza funcional pode ter sua resist?ncia aumentada ou diminu?da pelas intera??es. A concentra??o de intera??es positivas in grupos de baixa redund?ncia ou intera??es negativas em grupos de alta redund?ncia foi capaz de diminuir as taxas de extin??o funcional. Em contraste, a concentra??o de intera??es negativas em grupos de baixa redund?ncia ou de intera??es positivas em grupos de alta redund?ncia foi capaz de aumentar as taxas de extin??o funcional. Este modelo apresenta resultados relevantes para prioriza??o de esp?cies em trabalhos de conserva??o e restaura??o dos ecossistemas