A invasão de células tumorais para o tecido cerebral sadio é a marca patológica dos gliomas e contribui para o fracasso das modalidades terapêuticas atuais (cirurgia, radioterapia e quimioterapia). As células da glia transformadas apresentam os padrões clássicos do processo invasivo, incluindo adesão celular aos componentes da matriz extracelular (MEC), locomoção celular e a capacidade de remodelar o espaço extracelular. As metaloproteases de matriz (MMPs) são essenciais para o remodelamento adequado da MEC e para a invasão. O proteína supressora de tumor e metástase RECK regula pelo menos três diferentes membros da família das MMPs, especificamente: MMP-2, MMP-9 e MMP-14. Com o propósito de mimetizar o processo invasivo in vivo, as linhagens celulares de glioma humano A172 e T98G, respectivamente não invasiva e invasiva, foram plaqueadas em plástico (controle), colágeno tipo 1 ou Matrigel - membrana basal reconstituída, e incubadas por 3 e 7 dias, para estabelecer o processo invasivo. Nossos resultados mostram uma diminuição das taxas de proliferação e alterações morfológicas quando estas células foram cultivadas na presença de colágeno ou Matrigel. Microscopia eletrônica de transmissão das células T98G, cultivadas por 7 dias em colágeno, evidenciam invaginações de membrana similares ao que foi recentemente descrito como podossomos. Este novo tipo de estrutura é encontrado tipicamente em células que precisam cruzar barreiras teciduais, já que são sítios de degradação da MEC. A presença destas estruturas reforça o caráter invasivo da linhagem T98G. Ensaios de PCR em tempo real revelaram maior expressão de mRNA de RECK nas células A172, quando comparadas às células T98G, nas três condições de cultivo. Interessantemente as células A172 apresentaram maior expressão de RECK no colágeno tipo 1, enquanto a T98G demonstra uma tendência de aumento na expressão de RECK para colágeno tipo 1 e Matrigel. As MMPs são mais expressas e possuem maior atividade nas células T98G, e também são induzidas pelo substrato de colágeno. Estes resultados sugerem: 1) a expressão de RECK é diminuída pelo caráter invasivo apresentado por T98G, 2) o uso de substratos de MEC, como colágeno tipo 1 (A172 e T98G) e Matrigel (T98G), permite modular a expressão de RECK nestas linhagens de glioma. Como foi estabelecida uma correlação positiva entre a expressão de RECK e a taxa de sobrevida de pacientes para vários tipos tumorais, nossos resultados podem contribuir para o esclarecimento dos complexos mecanismos do processo invasivo no modelo de gliomas / The invasion of neoplastic cells into healthy brain tissue is a pathologic hallmark of gliomas and contributes to the failure of current therapeutic modalities (surgery, radiation and chemotherapy). Transformed glial cells display the common attributes of the invasion process, including cell adhesion to extracellular matrix (ECM) components, cell locomotion, and the ability to remodel the extracellular space. Matrix metalloproteinases (MMPs) are essential for proper ECM remodeling and invasion. The tumor and metastasis suppressor RECK protein regulates at least three members of the matrix metalloproteinase (MMPs) family, namely: MMP-2, MMP-9 and MMP-14. In order to mimic the in vivo invasion process, A172 and T98G, respectively, non-invasive and invasive human glioma cell lines were plated onto either plastic (control), or collagen type I gel or Matrigel™-basement membrane reconstituted, and incubated for 3 and 7 days, to establish the invasion process. Our results indicate decreased growth rates and morphological alterations regarding the invasive phenotype when these cell lines are cultured onto collagen gel and Matrigel™. Electronic transmission microscopy of T98G cells, cultured for 7 days onto collagen, pointed out membrane invaginations that are similar to what was recently described as podosomes. These new structures are typically found in cells that have to cross tissue boundaries, since they are sites of ECM degradation. The presence of these structures reinforces the invasive behavior of T98G cell line. Real time PCR assays revealed higher RECK mRNA expression in A172 cells, when compared to T98G cells in all three different substrate conditions. Interestingly, A172 cells displayed the upregulation of RECK expression in collagen gel, while in T98G high RECK expression was observed both in collagen and in Matrigel™. MMPs appear more expressed and more active for T98G cells, and are also upregulated by collagen. These results suggest that: 1) RECK expression is downregulated in the invasion process displayed by T98G cells, 2) coating of the substrate with ECM elements, such as collagen in the case of A172, and Matrigel™ and Collagen for T98G cells, can modulate RECK expression in glioma cell lines. Since positive correlation between RECK expression and survival of patients has been noted in several types of tumors, our preliminary results can contribute to elucidate the complex mechanisms of the glioma invasiveness process.

[pt] O Golfo Pérsico é responsável por aproximadamente trinta por cento da produção mundial de petróleo e detém mais da metade das reservas petrolíferas mundiais. A concentração geográfica do principal recurso enérgico, que alimenta o atual padrão tecnológico mundial, eleva essa região a um ponto de passagem obrigatório nas opções de geopolítica de todos os países do Sistema Internacional. O Golfo Pérsico é uma região de grande instabilidade política, e em menos de trinta anos, enfrentou três guerras internacionais: nos anos oitenta a Guerra Irã- Iraque, nos anos noventa a Guerra do Golfo e, mais recentemente a Invasão Americana ao Iraque. Tais conflitos foram marcados pelo uso, ou pela ameaça de uso, de armas de destruição em massa, e pelas perdas de um contingente imenso das populações dos países em conflito. Esta dissertação analisa a participação americana nestes três conflitos, tomando como referenciais conceitos de geopolítica, uma vez que a especificidade da região exige a retomada dessa disciplina que anda esquecida nas análises internacionais. A geopolítica procura enfatizar o impacto da geografia sobre a política; desta forma, a presença do petróleo no território do Golfo Pérsico, entendido como o Coração Energético Mundial, vai influir decisivamente nas suas relações com os outros Estados do Sistema Internacional. Este estudo analisa as opções de geopolítica dos EUA para a região durante os três conflitos, uma vez que assegurar o acesso às fontes de suprimento energético do Golfo Pérsico é um interesse nacional vital americano. / [en] The Persian Gulf produces about thirty per cent of the world's oil, while holding more than a half of the world's crude oil reserves. The geographical concentration of the most important energy resource that holds the world's contemporary technological standard, puts this region in a very important place for the geopolitical options for all countries in the International System. However, the Persian Gulf is a political unstable region in the world, in less than thirty years was involved in three international wars: in the eighties The Iran-Iraq War, in the nineties The Gulf War and recently The American Invasion of Iraq. These conflicts were known by the use or by the threat of use weapons of mass destruction (WMD), and by the heavy casualties in the countries involved in the war. This dissertation analyses the American participation in these three conflicts taking as referential geopolitical concepts, once the specificity of the region demands the rebirth of this discipline that was so often forgotten in the international analyses. The geopolitics emphasize the geographical impact over politics, so the oil reserves in the territory of the Persian Gulf, the energy heartland, will influence the relationship with the others States in the International System. This study examines the American geopolitical options for the region, once a secure access to Persian Gulf is America's national vital interest.

La mondialisation et les changements globaux actuels ont un impact considérable sur la distribution des espèces et la composition des communautés. Lorsque ces espèces sont impliquées dans une interaction hôte-parasite les changements dans leur répartition peuvent entraîner la (ré)émergence de maladies infectieuses. La fasciolose, maladie causée par les grandes douves du foie (Fasciola hepatica et Fasciola gigantica) est réémergente dans de nombreux points du globe. Il est difficile de mettre en place des programmes de contrôle parce que (1) les hôtes intermédiaires, des mollusques d’eau douce de la famille des Lymnaeidae, sont composés d’un groupe d’espèces cryptiques difficilement identifiables et (2) plusieurs espèces impliquées dans cette maladie sont invasives. L’objectif de cette thèse était d’étudier les facteurs écologiques et évolutifs à large échelle de l’interaction entre la grande douve du foie et ses hôtes intermédiaires susceptibles de favoriser une (ré)émergence de la fasciolose. J’ai, dans un premier temps, développé des approches moléculaires pour reconnaître les espèces cryptiques tant du côté de la grande douve du foie que du côté des Lymnaeidae. L’utilisation d’une des techniques développées sur Galba schirazensis, Galba cubensis et Galba truncatula, trois espèces de limnées, m’a permis d’identifier leur distribution respective et de modéliser leur niches bioclimatiques grâce à l’utilisation de modèles de niches écologiques. Cette approche de modélisation de niches permet d’inférer la distribution potentielle des trois espèces et nous amène à discuter des avantages potentiels de ces modèles dans la gestion de la fasciolose. Dans un second temps, je me suis intéressée à la structuration de la diversité génétique chez des espèces invasives du système grande douve du foie-limnée, par des approches de génétique des populations et de phylogénie. J’ai pu retracer l’histoire de colonisation, les dynamiques démographiques et le système de reproduction chez P. columella, G. schirazensis et G. cubensis. J’ai montré que ces trois espèces font préférentiellement de l’autofécondation entrainant des pertes drastiques de diversité génétiques sur le front d’invasion. G. cubensis présente cependant une coexistence de plusieurs génotypes dans les aires anciennement colonisées. Les différences génétiques entre ces trois espèces sont discutées à la lumière de ce qu’on sait de leur écologie. Enfin ces résultats m’ont permis de discuter de l’avantage d'être autofécondant en cas d’invasion biologique. Pour conclure l’invasion par ces populations d’hôtes intermédiaires dépourvus de diversité génétique pourrait représenter un risque épidémiologique. En effet un parasite a plus de probabilité de circuler dans une population hôte clonale que dans une population polymorphe. Cette thèse fait le lien entre écologie, interactions hôtes-parasites et génétique de l’invasion afin de mieux comprendre les facteurs d’expansion de la fasciolose à échelle globale. / Globalization and the current global change have significant impacts on species distribution and community composition. When these species are involved in a host-parasite interaction, changes in species range distribution can result in the (re)emergence of infectious diseases. Fasciolosis, a disease caused by the liver flukes (Fasciola hepatica and Fasciola gigantica) is reemerging in many parts of the world. It is difficult to implement control programs because (1) the intermediate hosts, freshwater molluscs of the Lymnaeidae family, are composed by a group of cryptic species (2) several species involved in this system are invasive. The objective of this thesis was to study the ecological and evolutionary factors at a large scale in the interaction between the liver fluke and its intermediate hosts that might drive to fasciolosis (re)emergence. First, I developed molecular approaches to quickly identify cryptic species on the two liver flukes and on three Lymnaeidae species. Using one of these techniques, I identify the respective distribution of Galba schirazensis, Galba cubensis and Galba truncatula and infer their respective climatic envelope by ecological niche modelling. We then modelled and projected the potential species distribution ranges. We discussed the contribution of models to predict the species distribution in space and time giving a considerable advantage to control fasciolosis. I then study the genetic diversity structuration in invasive snails involved in the transmission of F. hepatica, using population genetics and phylogeny. I could infer colonization history, population dynamics and reproductive system of Pseudosuccinea columella, G. schirazensis and G. cubensis. I showed that these three species preferentially make inbreeding causing drastic losses of genetic diversity in the invasion front. However G. cubensis presents a coexistence of several genotypes in formerly colonized areas. Genetic differences between these three species are discussed in the light of what we know about their ecology. Overall, these results illustrate how dramatic the reduction in genetic diversity can be for hermaphrodite animals. Finally, we discuss the epidemiological risk related to the invasion by intermediate hosts depleted in genetic diversity. Indeed, a parasite might circulate easily in a clonal host population than in a polymorphic population. In my thesis I linked ecology, host-parasite interactions with genetics of the invasion to better understand the expansion of fasciolosis at a global scale.

All previous taxonomic studies on the Ectemnorhinus group of weevils have been based primarily on morphological data. While these studies are invaluable, some questions can only be addressed adequately through molecular studies. This is especially true when studying the genetic relationships and phylogeograpic patterns of taxa endemic to the South Indian Ocean Province (SIP) biotas that have long been controversial. The Ectemnorhinus group of genera is a monophyletic unit of weevils endemic to the region. The present study focused mainly on the Ectemnorhinus group of weevils found on the Prince Edward Islands archipelago (PEIA). The mitochondrial cytochrome oxidase I gene was targeted when investigating relationships among members of this weevil group. On the PEIA, it is important to note that Marion Island (MI) and Prince Edward Island (PEI) differ in terms of alien invasive species, such as the introduced house mouse Mus musculus and in conservation management strategies. Since emergence, a series of volcanic and glaciation events have occurred on Marion Island, whilst Prince Edward Island has remained largely unaffected by glaciation. Phylogenetic analyses revealed the presence of two genetically and morphometrically distinct species of Ectemnorhinus weevils on PEI, whilst evidence for a single species, comprising diverse genetically discrete populations was found on MI. Based on these results, the species unique to PEI has been designated E. kuscheli n. sp., whilst the present study confirmed the synonymy between E. similis and E. marioni, the two species originally described from MI. Ectemnorhinus kucheli appears to be restricted to PEI, whereas E. similis occurs on both MI and PEI. When investigating the population dynamics of the Ectemnorhinus weevils on the PEIA, the data indicated that PEI was the first of the two islands of the PEIA to be colonized by Ectemnorhinus weevils, at an estimated time of coalescence of approximately 0.3116 million years ago (MYA). The PEI population then acted as the source population for the colonization of MI by Ectemnorhinus weevils some time before the last glaciation, approximately 10 000 to 35 000 years ago. The separation by distance of the PEI Ectemnorhinus weevils from those on MI then gave rise to two species by allopatric speciation on MI. During the last glaciations, MI was extensively glaciated with only the southwestern corner of the island being free of ice. This extensive glaciation of MI would have resulted in the eradication of all E. similis on MI except for those occurring on the ice-free southwestern corner of the island. At the end of the last glacial maximum, when the ice started to melt, the coastal areas of MI emerged first from beneath the ice and were available for re-colonization by weevils. The movement of weevils that were isolated in the south-western corner of MI, along the coastal areas of the island, was assisted by strong, frequent south-western winds. Subsequent, post-glacial volcanism during the Holocene was then responsible for the fragmentation of the new migrants, resulting in small population pockets surrounded by fresh, uninhabitable lava and subsequent divergence of each populations. When the Holocene black lava became re-colonizeable, the weevils from the different isolated populations migrated to the remainder of the island. Currently, members of the different genetically-identified populations occur in sympatry and in some cases even on the same plant, but no noticeable geneflow was detected between them. It is thus suggested that the time of isolation, before the post-glacial black lava during Holocene became hospitable, was sufficiently long and the populations sufficiently small that a number of genetically-discrete populations arose. Consequently, the present study recognises two genetically discrete populations of E. kucheli on PEI and seven discrete E. similis populations on MI that are morphologically indistinct. When examining the relationships among 13 species from five different islands within the South Indian Ocean Province (SIP) that are representative of 22 populations within the genera Palirhoeus, Bothrometopus and Ectemnorhinus, there was little support for separating the genus Palirhoeus from Bothrometopus, and no support for the morphologically-delineated species groups currently recognized within Bothrometopus. The present study shows that colonization of the Prince Edward Islands is likely to have occurred repeatedly from other islands within the SIP and that Bothrometopus parvulus on the PEIA comprises two species that are not sister taxa. The second novel con-generic species was therefore designated Bothrometopus huntleyi n. sp. and examination of the genetically identified specimens resulted in the indentification of distinguishing morphological characteristics. The analyses indicated that B. huntleyi arose approximately 0.5 million years ago from a high-altitude population that is still present on MI. The first major intra- and inter-island dispersal event occurred ~0.338 MYA, coinciding with the glaciation-free second volcanic stage on MI. Apart from this early inter-island colonisation, only one other between-island dispersal event, corresponding with the glaciation-free seventh volcanic stage, was detected. Genetically discrete weevil complexes on each of the islands of the PEIA together with the low levels of inter-island gene flow reaffirm the need to control alien invasive mice, which are restricted to MI, and which prey on these weevil species. / Thesis (PhD)--University of Pretoria, 2012. / Zoology and Entomology / unrestricted

HARP (Heparin Affin regulatory peptide) est un facteur de croissance qui constitue avec la midkine une sous-famille des Heparin Binding Growth Factors (HBGFs). HARP est impliqué dans de nombreux processus physiologiques comme la neurogenèse et la vasculogenèse mais aussi dans des processus physiopathologiques comme l’angiogenèse et la progression tumorale. HARP interagit avec différents récepteurs (N-syndécan, RPTPβ/ζ, et ALK). Plus récemment, il a été montré au laboratoire que la nucléoline, protéine navette entre le noyau, le cytoplasme, et la surface cellulaire est un nouveau récepteur à HARP. Malgré les avancées dans ce domaine, l’interaction de HARP avec ses récepteurs n’est pas totalement élucidée. L'objectif de ce projet de thèse était la recherche de nouveaux partenaires moléculaires qui interagissent avec HARP, de comprendre le mécanisme de leurs interaction et d’analyser les effets biologiques. A ce titre, j’ai participé à l’étude de l’interaction de HARP avec le facteur de croissance des fibroblastes, le FGF-2. Ce facteur liant l’héparine est également mitogène et angiogène. En utilisant des techniques de biocapteurs optiques et d’interaction protéine-protéine, nous avons montré une interaction directe entre HARP et le FGF-2 et qui implique les domaines C-TSR-I et C-terminale de HARP. De plus, cette interaction inhibe la migration et la prolifération des cellules endothéliales, induites par le FGF-2 ou par HARP seuls. En parallèle, j’ai mis en évidence l’interaction entre HARP et la NRP-1. NRP-1 est une protéine transmembranaire, ayant comme ligands principaux, les sémaphorines de classe 3 (SEMA 3A), le facteur de croissance endothélial vasculaire (VEGF) et le FGF-2. En plus de son rôle crucial dans le développement des systèmes nerveux et cardiovasculaires, la NRP-1 est impliquée dans les processus physiopathologiques tels que l’angiogenèse et l’invasion tumorale. Ainsi, la NRP-1 présente un profil biologique similaire à HARP. En utilisant des tests d’ELISA, d’immunoprécipitation et de « pull-down », nous avons montré que HARP interagit avec la NRP-1. Cette interaction semble être directe et s’effectue via les domaines de liaison à l’héparine TSR-I. HARP induit l’internalisation de la NRP-1 au bout de 15 minutes et son recyclage partiel à la surface cellulaire au bout d’une heure. L’internalisation de la NRP-1 s’accompagne par la phosphorylation des voies MAPK (ERK1/2), Akt et FAK. L’interaction HARP/NRP-1 est cruciale pour la migration des cellules endothéliales et l’invasion des cellules tumorales. En conclusion, ces résultats apportent de nouvelles avancées concernant les partenaires moléculaires de HARP en particulier et montrent également la complexité des interactions des facteurs de croissance entre eux et avec leurs récepteurs. Plus généralement, cette étude permet d'envisager des stratégies thérapeutiques ciblant l’interaction de la NRP-1 avec HARP et aussi les autres facteurs de croissance. / HARP (Heparin Affin regulatory peptide) is a growth factor that constitutes with midkine a subfamily of Heparin Binding Growth Factors (HBGFs). HARP is involved in many physiological processes such as neurogenesis and vasculogenesis but also in pathophysiological processes such as angiogenesis and tumor progression. HARP interacts with different receptors (N-syndecan, RPTPβ / ζ and ALK). More recently, it has been shown in the laboratory that nucleolin, a protein shuttle between the nucleus, cytoplasm, and cell surface, is a new HARP receptor. Despite the advances in this field, the interaction of HARP with its receptors is not fully understood. The aim of this thesis was the search for new molecular partners that interact with HARP, to understand the mechanism of their interaction and analyze the biological effects. My work was firstly to participate to the study of the interaction of HARP with the fibroblast growth factor-2, FGF-2. This factor is also an heparin-binding factor, with mitogenic and angiogenic activities. Using techniques of optical biosensors and protein-protein interaction, we have shown a direct interaction between HARP and FGF-2 that involves C-TSR-I and C-terminus domains of HARP. In addition, HARP inhibits the migration and proliferation of endothelial cells induced by FGF-2. In parallel, I highlighted the interaction between HARP and NRP-1. NRP-1 is a transmembrane protein having as main ligands, semaphorins class 3 (SEMA 3A), the vascular endothelial growth factor (VEGF) and FGF-2. In addition to its crucial role in the development of the nervous and cardiovascular systems, the NRP-1 is involved in physiopathological processes such as angiogenesis and tumor invasion. Thus, NRP-1 has a biological profile similar to HARP. Using ELISA, immunoprecipitation and "pull-down" tests, we have shown that HARP interacts with NRP-1. This interaction appears to be direct and occurs via heparin binding domains of HARP: TSR-I. HARP induces internalization of NRP-1 after 15 minutes and partial recycling to the cell surface after one hour. The internalization of the NRP-1 is accompanied by the phosphorylation of MAPK pathways (ERK1 / 2), Akt and FAK. HARP/NRP-1 interaction is crucial for endothelial cell migration and invasion of tumor cells. In conclusion, these results provide new advances on molecular partners of HARP in particular and also show the complexity of the interactions between these growth factors and their receptors. More generally, this study allows considering therapeutic strategies targeting the interaction of NRP-1 with HARP as well as other growth factors.

La compréhension de l'émergence des maladies infectieuses végétales causées par les bactéries passe par l'identification des populations sources, des routes d'invasion et des voies de dissémination, ainsi que par l'estimation des paramètres biotiques et abiotiques associés. Xanthomonas citri pv. citri (Xcc) est l'agent pathogène responsable du chancre Asiatique des agrumes. La bactérie est distribuée dans plusieurs pays agrumicoles du monde et listée comme organisme de quarantaine par ceux où elle est absente. Nous avons abordé l'épidémiologie moléculaire de Xcc à deux échelles spatio-temporelles grâce à un schéma de 14 microsatellites (MLVA-14) et un schéma de 31 marqueurs minisatellites (MLVA-31). Le typage MLVA-14 s'est montré adapté au génotypage d'une bactérie monomorphe comme Xcc. Le typage MLVA-31 a permis de diviser le pathovar Xcc en quatre groupes génétiques distincts correspondant aux différences de gammes d'hôtes mise en évidence chez cette bactérie. Le pathotype A (souches à large gamme d'hôtes parmi les rutacées) est séparé en deux groupes génétiques, tandis que les pathotypes A* et Aw (souches à gamme d'hôtes restreinte au limettier Mexicain et quelques espèces proches) constituent chacun un groupe génétique. Alors que l'expansion géographique de Xcc depuis son aire d'origine dans la première moitié du XXème siècle a quasi exclusivement concerné un seul groupe génétique, trois des quatre groupes décrits ont contribué à l'émergence de Xcc en Afrique au cours de la dernière décennie. La bactérie est pré-adaptée et a été introduite avec son hôte depuis sa population d'origine, faisant de la barrière migratoire la seule étape à franchir pour rencontrer un succès d'invasion. L'objectif de cette thèse a été de décrire les différentes populations émergentes grâce à des approches d'épidémiologie moléculaire et inférentielles, et identifier les différentes origines, routes et acteurs de la dissémination. Nous avons dans un premier temps montré la coexistence de deux groupes génétiques distincts au Mali : DAPC1 qui est dispersé dans quatre provinces du pays et DAPC2 qui est resté cantonné à l'espace péri-urbain de Bamako. L'analyse de l'émergence de Xcc au Sénégal a révélé le succès invasif de DAPC2 dans un autre environnement. La structure des populations DAPC1 du Mali et DAPC2 du Sénégal suggèrent que les plants de pépinières constituent une voie de dissémination majeure dans ces pays. À l'opposé, DAPC2 de Bamako n'est pas détecté en pépinières au Mali et n'a pas montré de caractère invasif. L'existence d'une population « tête de pont » invasive de souches DAPC1 au Mali donnant lieu à une épidémie au Burkina Faso a été mise en évidence par une approche ABC (calcul Bayésien approché). Les populations DAPC2 du Mali et du Sénégal ne présentent pas de lien épidémiologique direct mais partagent des liens de parenté avec des souches présentes dans le sous-continent Indien. Dans un deuxième temps, l'analyse d'une population de souches appartenant au pathotype A* en Ethiopie nous a permis de procéder à des estimations de paramètres démographiques, tels que les tailles efficaces. Nous avons montré que l'approche inférentielle nous permettait d'éclairer l'histoire démographique de Xcc dans un cas d'émergence, et de mettre en avant une dynamique saisonnière accentuant probablement le déséquilibre mutation-dérive lié à la situation d'émergence. L'émergence de Xcc en Afrique est principalement associée aux activités humaines. Sa dissémination locale et globale peut alors être considérablement limitée par des mesures de gestion plus stricte au niveau des pépinières et des flux de commerces. / Several plant emerging infectious diseases are caused by bacteria. To improve our understanding of their emergence, a description of the emerging populations, the reconstruction of invasion routes and pathways, as well as the identification of involved biotic and abiotic parameters are fundamental. The bacterium Xanthomonas citri pv. citri (Xcc) is responsible for Asiatic citrus canker. It is present in many citrus producing countries and listed as a quarantine organism in canker-free countries.Two MLVA schemes were used for molecular epidemiological analyses of Xcc. The first one, MLVA-14, targeted 14 microsatellite markers, and is useful to describe the genetic diversity of this monomorphic bacterium. The second, MLVA-31, targeted 31 minisatellite markers, is suited to global epidemiology analyses. Based on MLVA-31 data, Xcc is divided in four genetic groups (referred to as DAPC clusters) corresponding to Xcc pathotype classification based on host range. Three pathotypes were described: pathotype A strains are able to infect most citrus species, while pathotypes A* and AW strains are naturally restricted to fewer host species.DAPC 1 is responsible for almost all cases of geographical expansion of Xcc over the first half of the 20th century, we show that three Xcc genetic clusters have emerged in Africa over the last decade. Xcc is pre-adapted to its host species. Invasive success of Xcc is then mostly conditioned by migration events. Our objectives were to describe these invasive populations using a molecular epidemiology approach and to assess the origin, routes and actors of this dissemination in Africa. Two genetic clusters were found in Mali: DAPC1 is present in four provinces while DAPC2 is restricted to the Bamako urban environment. In contrast, DAPC2 emerging populations in Senegal showed an invasive succes in an other environment. Populations structures of DAPC1 in Mali and DAPC2 in Senegal highlighted the role of nurseries in Xcc dissemination. On the contrary, DAPC2 strains in Bamako were not detected in Malian nurseries and showed a limited invasive success. Approximate Bayesian Computation highlighted an invasive bridgehead scenario between DAPC1 in Mali and Burkina Faso. DAPC2 populations in Mali and Senegal were not found epidemiologically related but were genetically related to strains previously reported from the Indian subcontinent.Demographic parameters inference, such as effective population sizes, were inferred from Ethiopian pathotype A* populations. The inference approach was useful to decipher the demographic history of this emerging population, and suggested seasonal fluctuations in population sizes over time.Emergence of Xcc in Africa was found strongly related to human activities. Therefore, the local and global dispersion could be limited by a better management of nurseries and trade.

Transforming growth factor β (TGF-β) and bone morphogenetic protein (BMP) signaling pathways are involved in many physiological processes during embryonic and adult life. TGF-β promotes epithelial to mesenchymal transition (EMT). We identified a gene target of TGF-β signaling, encoding the salt-inducible kinase 1 (SIK1). A potential substrate of this kinase, the polarity protein Par3, is an established regulator of tight junction assembly. SIK1 associates with Par3, can potentially phosphorylate Par3 and leads to its degradation, contributing to tight junction disassembly. Glioblastoma multiforme (GBM) is a common malignancy in the central nervous system, characterized by high heterogeneity, invasiveness, and resistance to therapy. One of the causes of heterogeneity and therapy-resistance is the existence of glioblastoma stem cells (GSCs). TGF-β signaling promotes self-renewal while BMP signaling induces differentiation of GSCs. Snail is a potent inducer of the EMT in carcinomas. However, in the context of GBM, Snail induces BMP signaling and represses TGF-β signaling through interaction with SMADs, the signaling mediators of TGF-β and BMP. In conclusion, Snail differentially regulates the activity of the opposing BMP and TGF-β pathways, thus promoting an astrocytic fate switch and repressing stemness in GSCs. Although profound changes in cell polarity is a hallmark of invasive malignancies, little is known about the role of the polarity machinery in tumor suppression. Patient transcriptomic data suggested low Par3 expression, correlating with poor survival of the GBM patients. Par3 silencing decreased the GSC self-renewal capacity and enhanced their invasiveness. Transcriptomic analysis indicates that loss of Par3 leads to downregulation of genes encoding mitochondrial enzymes that generate ATP. These results support a novel role of Par3 in GBM, beyond its contribution to junctional contacts between cells. Another regulator of TGF-β and BMP signaling is the liver kinase B1 (LKB1). According to GBM patient mRNA analysis, high levels of LKB1 correlate with poor prognosis. Silencing of LKB1 in GSCs impairs invasion and self-renewal capacity due to downregulation of genes involved in these processes. Moreover, loss of LKB1 induces mitochondrial dysfunction, leading to decreased ATP levels. Collectively, this thesis has delivered a group of novel regulatory pathways that control critical aspects of cancer cell polarity, invasion and stemness.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-04-19T12:54:17Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO DE MESTRADO - SILAS VALENTE.pdf: 3356138 bytes, checksum: 5efc9b5c2ec967aba4a0eded24e8fd9a (MD5) / Made available in DSpace on 2016-04-19T12:54:17Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO DE MESTRADO - SILAS VALENTE.pdf: 3356138 bytes, checksum: 5efc9b5c2ec967aba4a0eded24e8fd9a (MD5) Previous issue date: 2015-07-31 / As linhas de transmissão de energia elétrica (LT) são construções que se destinam ao transporte de energia elétrica das fontes geradoras aos centros de consumo. Uma LT possui como elementos básicos: cabos, isoladores e torres. O espaço no solo ocupado por esses elementos e entorno deve ser respeitado frente às diversas formas de ocupação. Esta área protegida é denominada Faixa de Servidão. A faixa de servidão, como território ocupado com potencial de geração de riscos à população, têm inúmeras restrições quanto à sua utilização, dada sua destinação: proteger o sistema elétrico e a sociedade. Sua largura varia em função da tensão dos cabos. Para tensões de 230 e 500 kV (kilovolts) a largura da faixa de servidão é respectivamente de 50 e 65 metros. A ocupação e conservação da faixa de servidão de forma adequada possibilitam maior segurança para as pessoas e bens localizados na vizinhança, bem como maior rapidez na localização de anomalias e na manutenção na LT. Ocupações das faixas de servidão, que não estejam contempladas no projeto de instalação da LT, são consideradas ocupação irregular ou invasão e devem ser identificadas e removidas. Atualmente existem dados de sensoriamento remoto de média e alta resolução espacial que podem auxiliar no estudo do uso e cobertura do solo. Neste contexto, esta pesquisa tem como objetivo desenvolver uma metodologia, a partir de dados de sensoriamento remoto (CBERS-2B/HRC e SRTM), para identificar áreas invadidas e suscetíveis à ocupação irregular em faixa de servidão de LT. Para isso foram aplicadas técnicas de segmentação, classificação de imagem e desenvolvido modelos matemáticos usando parâmetros definidos nesta pesquisa. A área de estudo é a faixa da LT 230 kV Ibicoara/Brumado, localizada no estado da Bahia. A pesquisa caracterizou de forma abrangente e eficaz, a temática de LT e faixa de servidão, envolvendo aspectos técnicos, ambientais e de segurança, e contextualizou o sensoriamento remoto relacionando-o com o tema, envolvendo sistemas sensores e aplicações. Os resultados do estudo mostraram-se eficientes na observação/fiscalização do quadro de ocupação da faixa de servidão em locais específicos, sendo possível antecipar ocorrência de invasões, auxiliando na gestão de rede transmissão de energia elétrica. / The transmission lines (TL) are constructions intended for the transport of electricity from the generating sources to consumption centers. A TL has as basic elements: cables, insulators and towers. The land space occupied by these elements and surroundings must be respected across the various forms of occupation. This protected area is called right of way. The right of way as a potential risk area for population has numerous restrictions for its use, given their destination: protect the electrical system. Its width varies depending on the tension of the cables. For voltages of 230 and 500 kV (kilovolts) the width of the right of way is respectively 50 and 65 meters. The occupation and conservation of the right of way enables greater security for people and property located in the surroundings, as well as greater speed in detecting anomalies and for the maintenance of the TL. Occupations in the right of way, which are not included in the TL installation project, are considered illegal occupation or invasion and must be identified and removed. Currently there are remote sensing data of medium and high spatial resolution that can support the study of land use and land cover. In this context, this research aims to develop a methodology, based on remote sensing data (CBERS-2B / HRC and SRTM) to identify areas invaded and susceptible to irregular occupation in the right of way. Therefore, segmentation techniques, image classification and developed mathematical models using parameters defined for this search were applied. The study area is the right of way of the TL 230 kV Ibicoara / Brumado, located in the state of Bahia. This study used a comprehensive and effective way to characterize the issues related to TL and its right of way, considering technical, environmental and safety aspects, and contextualized remote sensing relating it to the subject, considering sensor systems and applications. The results of the study showed to be efficient in the observation / surveillance for the status of occupation of the right of way, in specific locations, making possible the anticipation of occurrence of invasions, giving more support for the management of power grid.

A causa de morte da maioria das pacientes com câncer de mama se deve à doença metastática desenvolvida a partir do tumor primário. A degradação dos componentes da matriz extracelular (MEC), um dos principais eventos do processo metastático, é regulada pelo balanço entre as atividades das metaloproteinases de matriz (MMPs) e dos seus inibidores, tanto os inibidores teciduais (TIMPs) como o inibidor associado à membrana (RECK). Contudo, ainda existe pouca informação sobre os mecanismos moleculares responsáveis pela manutenção deste balanço. No presente trabalho, foi investigado o envolvimento de TGF-β1 (Transforming Growth Factor-β1), uma citocina multifuncional é capaz tanto de inibir o crescimento celular, quanto de promover invasão e metástase, dependendo do estadiamento e do tipo de tumor, na regulação da expressão de MMPs, TIMPs e RECK, em modelo de câncer de mama. Primeiramente, examinou-se os níveis de expressão de mRNA das isoformas e receptores de TGF-β, em um painel de cinco linhagens de carcinoma mamário humano, com diferentes potenciais invasivos e metastáticos, por qRT-PCR. Os resultados obtidos demonstraram uma correlação positiva entre a expressão dessas moléculas, e a progressão do caráter invasivo e metastático celular. Em seguida, a linhagem altamente invasiva, MDA-MB-231, foi tratada com diferentes concentrações de TGF-β1 recombinante. Esta citocina foi capaz de modular a expressão gênica de MMPs (MMP-2 e MMP-9) e de seus inibidores (TIMP- 2 e RECK). Tanto ERK½, quanto p38MAPK mostraram-se envolvidas neste mecanismo. Foi demonstrado que a inibição da atividade de ERK½ alterou a expressão das proteínas MMP-9, TIMP-2 e RECK, enquanto o bloqueio de p38 MAPK afetou os níveis protéicos de MMP-2 e TIMP-2. O aumento do potencial migratório e invasivo da linhagem MDA-MB-231, induzido por TGF-β1, mostrou-se também dependente da atividade de MMPs, ERK½ e p38MAPK. Dada a ausência de informações sobre o papel de RECK em modelo mamário, a função deste inibidor de MMPs também foi investigada. Primeiramente, analisou-se a expressão de RECK ao longo do desenvolvimento da mama e, posteriormente, em 1040 amostras tumorais de mama humana, através da metodologia de Tissue Microarray, tendo sido possível demonstrar que a alta expressão de RECK associa-se a menor tempo de sobrevida global e livre de doença em 10 anos. Os resultados obtidos indicaram que a expressão da proteína RECK, em oposição ao verificado em outros tipos de tumores, está relacionada ao fenótipo mais agressivo de tumores de mama. Entretanto, a análise funcional de RECK, realizada por meio da utilização de vetores shRNA específicos para a inibição desta proteína, demonstrou que RECK também atua como um inibidor de invasão celular e da expressão de MMP-9, na linhagem MDA-MB-231. Em conjunto, os resultados obtidos neste trabalho contribuíram para a elucidação dos mecanismos moleculares de regulação de RECK, por clássicas moléculas associadas ao processo de tumorigênese (TGF-β1 e MAPKs), bem como para o esclarecimento de suas funções em modelo mamário, sugerindo-o como mais um promissor candidato a marcador prognóstico e alvo molecular para a terapia do câncer de mama. / The metastatic disease is the main mortality cause of breast cancer patients. The metastatic process involves a complex cascade of events, including the organized breakdown of the extracellular matrix (ECM) compounds. The degradation of ECM is tightly regulated by the balance between the activities of matrix metalloproteinases (MMPs) and their inhibitors, the tissue inhibitors (TIMPs) and the membrane-associated inhibitor (RECK). Among the several molecules released and activated by ECM remodeling, TGF-β1 (Transforming Growth Factor-β1) is a multifunctional cytokine able to regulate both cell growth inhibition and invasion and metastasis promotion, depending on the tumor stage and type. Since the molecular mechanisms involved in the ECM remodeling control are still not completed understood, in this study, we investigated the involvement of TGF-β1 in regulating of MMPs, TIMPs and RECK expression, in the breast cancer model. By qRT-PCR, we first examined the gene expression levels of TGF-β isoforms and receptors, in a panel of five human breast cancer cell lines displaying different degrees of invasiveness and metastatic potential. Our results suggest a positive correlation between the mRNA expression of these molecules and the breast cancer progression. Moreover, the highly invasive breast cancer cell line MDA-MB-231 was treated with different concentrations of recombinant TGF-β1. We described that this cytokine was able to modulate the gene expression of MMPs (MMP-2 and MMP-9) and MMPs inhibitors (TIMP-2 and RECK) at both the mRNA and protein levels, with ERK½ and p38 MAPK being involved in this molecular mechanism. However, while ERK½ activity inhibition altered MMP-9, TIMP-2 and RECK expression, the p38 MAPK blockage affected the protein levels of MMP-2 and TIMP-2. Finally, we reposted that the TGF-β1-enhanced migration and invasion capacities of MDA-MB- 231 cells were blocked by MMPs, ERK½ and p38 MAPK inhibitors. Analysis of the RECK function in the breast model was also an objective of this study. We analyzed RECK expression during mammary gland development. We evaluated the RECK protein profile in 1040 breast tumor tissue samples using Tissue Microarray assays. We demonstrated that high expression levels of RECK were associated with shorter overall and disease-free survival in 10 years. Moreover, we verified that RECK is a biomarker of poor prognosis mainly for patients diagnosed with less aggressive breast tumor. Therefore, in contrast to other tumor types, our results indicate that high protein expression levels of RECK are related to a more aggressive phenotype. In fact, the RECK functional analysis, performed by using of shRNA vectors, showed that RECK function remains as an inhibitor of cellular invasion and MMP-9 expression, in MDA-MB-231 cells. Taken together, our results contribute to better understanding of the molecular mechanisms associated to RECK regulation by TGF-β1 and MAPK as well as to clarify its role in breast model. Thus, we suggests RECK as a new and promising prognostic marker and molecular target candidate for breast cancer therapy.

Bien moins médiatisées que la déforestation ou le changement climatique, les invasions biologiques n’en demeurent pas moins problématiques pour la conservation de nombreux écosystèmes ou le bon déroulement de nombreuses activités socio-économiques.Comprendre la façon dont les espèces exotiques envahissantes s’étendent dans les différents milieux qu’elles colonisent c’est permettre à la fois de mieux appréhender leurs impacts, de pouvoir de prédire leur expansion, et de trouver les endroits stratégiques où agir pour contrer ou atténuer les effets indésirables de ces espèces. Le problème est que dynamiques d’invasions d’une espèce regroupent en fait une hiérarchie de processus se produisant simultanément à différentes échelles spatiotemporelles et qui sont contrôlés par des facteurs qui changent en fonction du temps et du contexte d’invasion. Pour acquérir une compréhension profonde de ces dynamiques, il faut donc étudier leurs manifestations et leurs causes aux diverses échelles spatiales et temporelles auxquelles elles se produisent.Présentes sur les cinq continents et très envahissante sur au moins deux, les renouées asiatiques (Reynoutria spp.) sont reconnues comme faisant partie des espèces les plus invasives de la planète. Dotées d’une grande tolérance environnementale et d’importantes capacités de multiplication végétatives, ces plantes herbacées géantes peuvent former rapidement de grandes monocultures clonales qui excluent les autres espèces végétales, modifient les cycles biogéochimiques des zones envahies et perturbent diverses activités anthropiques. Cibles de campagnes de gestion depuis des décennies, elles affichent cependant une résilience insolente qui désespère de nombreux gestionnaires. Bien que très étudiées, de nombreux détails concernant leurs dynamiques nous échappent toujours. Dans ce travail de thèse, nous avons donc cherché à identifier quels sont les facteurs qui contrôlent les dynamiques spatiotemporelles des renouées à différentes échelles, et comment une meilleure compréhension de ces dynamiques clés pourrait profiter à leur gestion ?Pour ce faire, nous nous sommes donc d’abord intéressés à leur dynamiques clonales et à leurs variations en fonction de différentes conditions de stress (ombrage) ou de perturbation (fauchage répétée). Nous avons montré que si les renouées sont bien affectées par des conditions stressantes ou perturbées, cela ne les empêche ni de s’établir, ni de croître. En fait, les renouées semblent pouvoir adopter différentes stratégies de croissance pour pallier ces contraintes, soulevant différentes questions liées à la gestion. Dans une deuxième étude, nous avons cherché quels étaient les variables qui expliquaient l’expansion des taches de renouées le long d’un gradient altitudinal. Si nous avons montré que les dynamiques d’expansion des taches étaient principalement contrôlées par leur taille (et donc potentiellement leur âge) ainsi que par la proximité de routes et de rivières, nous avons également apporté des indices qui suggèrent que les renouées pourraient être potentiellement capables d’envahir les montagnes. Ensuite, pour tenter d’aider à résoudre le problème de l’acquisition des données de distribution des renouées à large échelle, nous avons développé une méthode pour détecter et cartographier les populations de renouées à partir d’images satellites et issues de drones. Notre méthode a montré des résultats encourageants et celle-ci pourrait être utile à l’étude des invasions ainsi qu’à la détection des nouveaux foyers d’invasion pour une gestion plus précoce et efficace des renouées.Enfin, nous avons tenté de dresser un portrait global de la compréhension actuelle des dynamiques d’invasion des renouées en réalisant, en guise de discussion, une grande revue de littérature sur les mouvements de ces plantes à travers les échelles spatiales et temporelles et en y intégrant les apports des autres travaux de cette thèse. / Although less mediatized than deforestation or climate change, biological invasions remain problematic for the conservation of many ecosystems and the maintenance of various socio-economical activities.Understanding the way invasive non-native species spread in their introduced range is enabling a better apprehension of their impacts, the possibility to predict their expansion, and the development of better management strategies. A problem is that a species invasion dynamics are actually composed of a hierarchy of processes occurring simultaneously at various spatiotemporal scales and which are controlled by drivers that are time- or context-dependent. To gain more profound insights into these dynamics, one has to study the patterns they create and their underlying processes at all relevant scales.Occurring on the five continents and highly invasive on at least two of them, Asian knotweeds (Reynoutria spp.) are acknowledged as being among the most invasive species in the world. Gifted with a large environmental tolerance and high abilities for vegetative multiplication, these giant herbaceous plants can quickly form large clonal monocultures that exclude the other plant species, modify biogeochemical cycles, and affect various anthropic activities. Target of management campaigns for decades, knotweeds display an insolent resilience to the great despair of many stakeholders. Despite having been extensively studied, many aspects of their dynamics are still elusive. Consequently, in this thesis, we have tried to highlight what the drivers of the spatiotemporal dynamics of knotweeds across scales are, and how their management could benefit from a better understanding of these dynamics?To answer these questions, we first focused our attention on the clonal dynamics of knotweeds and on their variations when they experience differing conditions in terms of light availability and disturbance (repeated mowing). We have shown that although shade or mowing affects the vigour and the development of clones, it does not prevent their establishment or their growth. In fact, knotweeds seem able to adopt different growth strategies to cope with environmental heterogeneity, suggesting some management avenues. In a second study, we tried to identify the drivers that controlled the expansion of knotweed stands along an elevational gradient. If we showed that the lateral expansion of stands is mostly constrained by their size (and thus, possibly their age) and the vicinity of roads and rivers, we also brought evidences that knotweeds could have the potential to invade mountain regions. Then, to help for the acquisition of large-scale datasets on knotweeds’ distribution, we developed a procedure to accurately detect and map knotweeds using satellite or drone imagery. Our results were quite encouraging and could be useful to both the basic research and to the detection of newly invaded areas, enabling earlier control operations and more efficient management.Finally, we tried to draw a full picture of the current knowledge on the invasion dynamics of knotweeds by reviewing the literature on the movement of these plants across spatial and temporal scales, to discuss and expand the reach of the insights gained in the various chapters of this thesis.