<p>Initiation of protein synthesis in <i>E.coli </i>involves several steps, which lead to the formation of the first peptide bond. This process requires three initiation factors: IF1, IF2 and IF3. Using a novel technique of combined light scattering and stopped-flow, we elucidated the importance of IF2•GTP conformation for the recruitment of 50S to 30S pre-initiation complex. Moreover, GTP hydrolysis is essential for IF2 release and later binding of ternary complex. Interestingly, a switch in IF2 affinity to G-nucleotides is induced during 30S pre-initiation complexes formation. </p><p>We found that IF1, previously with unknown functions in vitro, increases the rate of naked 70S dissociation by a factor 80 and acts as a fidelity factor in preventing 70S formation containing elongator tRNA instead of fMet-tRNA^fMet. We showed that RRF/EFG/IF3 split both naked and post-termination complexes while IF1/IF3 split only naked ribosomes. The mechanisms of action of RRF/ EFG, the order of their binding to 70S, as well as, the three different conformation of EF-G on the ribosomes are emphasized. Interestingly, 70S formation rate is dependent on the concentration of IF3 and not linear with 50S subunits concentration. We demonstrated that the rate-limiting step in 70S formation is IF3 dissociation from 30S complexes.</p><p>The interplay between initiation factors in the rate and accuracy of protein synthesis was thoroughly studied. Using fMet-tRNA^fMet (initiator tRNA), Met-tRNA^fMet(non-formylated initiator tRNA) and Phe-tRNA^Phe (elongator tRNA), we showed that the major player in the accuracy is IF2 through recognizing the formyl group on fMet-tRNA^fMet, while IF3 acts by increasing both the on- and off-rate of tRNA from 30S pre-initiation complexes.</p><p>Collectively, these novel results describe a comprehensive model of initiation of protein synthesis. In this model, initiation factors increase the rate of fMet-tRNA^fMet binding to 30S subunits, subsequently; the stabilization of fMet-tRNA^fMet by IF2 increases the rate of IF3 dissociation. Later, IF2 in GTP conformation allows 50S docking to 30S pre-initiation complex free of IF3 followed by GTP hydrolysis allowing IF2 release for ternary complex to bind and start elongation of protein synthesis. </p>

Initiation of protein synthesis in E.coli involves several steps, which lead to the formation of the first peptide bond. This process requires three initiation factors: IF1, IF2 and IF3. Using a novel technique of combined light scattering and stopped-flow, we elucidated the importance of IF2•GTP conformation for the recruitment of 50S to 30S pre-initiation complex. Moreover, GTP hydrolysis is essential for IF2 release and later binding of ternary complex. Interestingly, a switch in IF2 affinity to G-nucleotides is induced during 30S pre-initiation complexes formation. We found that IF1, previously with unknown functions in vitro, increases the rate of naked 70S dissociation by a factor 80 and acts as a fidelity factor in preventing 70S formation containing elongator tRNA instead of fMet-tRNAfMet. We showed that RRF/EFG/IF3 split both naked and post-termination complexes while IF1/IF3 split only naked ribosomes. The mechanisms of action of RRF/ EFG, the order of their binding to 70S, as well as, the three different conformation of EF-G on the ribosomes are emphasized. Interestingly, 70S formation rate is dependent on the concentration of IF3 and not linear with 50S subunits concentration. We demonstrated that the rate-limiting step in 70S formation is IF3 dissociation from 30S complexes. The interplay between initiation factors in the rate and accuracy of protein synthesis was thoroughly studied. Using fMet-tRNAfMet (initiator tRNA), Met-tRNAfMet (non-formylated initiator tRNA) and Phe-tRNAPhe (elongator tRNA), we showed that the major player in the accuracy is IF2 through recognizing the formyl group on fMet-tRNAfMet, while IF3 acts by increasing both the on- and off-rate of tRNA from 30S pre-initiation complexes. Collectively, these novel results describe a comprehensive model of initiation of protein synthesis. In this model, initiation factors increase the rate of fMet-tRNAfMet binding to 30S subunits, subsequently; the stabilization of fMet-tRNAfMet by IF2 increases the rate of IF3 dissociation. Later, IF2 in GTP conformation allows 50S docking to 30S pre-initiation complex free of IF3 followed by GTP hydrolysis allowing IF2 release for ternary complex to bind and start elongation of protein synthesis.

No description available.

Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2017-10-03T11:41:38Z No. of bitstreams: 2 Dissertação - Óscar Morais Fernando Namuholopa - 2017.pdf: 2659597 bytes, checksum: c4b704da3d80eac474b0654f92fac768 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-10-03T11:41:57Z (GMT) No. of bitstreams: 2 Dissertação - Óscar Morais Fernando Namuholopa - 2017.pdf: 2659597 bytes, checksum: c4b704da3d80eac474b0654f92fac768 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-10-03T11:41:57Z (GMT). No. of bitstreams: 2 Dissertação - Óscar Morais Fernando Namuholopa - 2017.pdf: 2659597 bytes, checksum: c4b704da3d80eac474b0654f92fac768 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-02-20 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / This dissertation, entitled The role of initiation rites in the yaawo community: case of the city of Lichinga-Mozambique has the aim: to analyze the social role of initiation rites in the yaawo community; to characterize the practice of initiation rites in this community in present times Of cultural flow; to differentiate male and female initiation related to practice and thematic and to describe the role of initiation rites in the preservation of the socio-cultural values of the region. Its execution has become possible through research developed within the community, involving initiates as witness, teachers and managers of the schools where these initiates seek their learning. The research with schools and its agents aimed to analyze the relationship level between the practice of initiation rites and school institutions. The findings showed that initiation rites have the role of reinforcing internal social cohesion among members of the community, disseminating among their participants the spirit of solidarity and awakening the sense of belonging. They also have the role of socialization between initiates and the community at large. They create moral guidelines and outline the community's rules of social conduct. Initiation in the yaawo context means the attainment of social majority. By initiation the individual passes from childhood to adulthood, thus becoming part of the active part of the life of his community and enjoying full rights. They also mean the highest moment of the exaltation of culture and community identity. / Esta dissertação, intitulada O papel dos ritos de iniciação na comunidade yaawo: caso da cidade de Lichinga-Moçambique centra o seu objetivo em analisar o papel social dos ritos de iniciação na comunidade yaawo, caracterizar a prática dos ritos de iniciação nesta comunidade em tempos atuais de fluxo cultural, diferenciar a iniciação masculina e feminina quanto à prática e temática e descrever o papel dos ritos de iniciação na preservação dos valores socioculturais da região. A suaexecução tornou-se possível através da pesquisa desenvolvida dentro da comunidade, envolvendo iniciados, mestres e gestores das escolas onde estes iniciados buscam o seu aprendizado. A pesquisa junto destes últimos visava aferir o nível de relacionamento entre a prática dos ritos de iniciação e as instituições escolares. Da pesquisa, constatei que os ritos de iniciação têm o papel de reforçar a coesão social interna entre os membros da comunidade, disseminar entre os seus participantes o espírito de solidariedade e despertar o sentimento de pertença. Têm igualmente, o papel de socialização entre os iniciados e a comunidade em geral. Eles criam linhas mestras da moral e traçam as regras de conduta social da comunidade. A iniciação no contexto yaawo significa a conquista da maioridade social. Pela iniciação o indivíduo passa da infância para a vida adulta, passando assim a integrar a parte ativa da vida da sua comunidade e gozando de plenos direitos. Significam ainda, o momento mais alto da exaltação da cultura e identidade da comunidade.

Cette thèse présente une ethnologie des phénomènes migratoires en Afrique de l’Ouest qui promeut une étude de la mobilité, localement baptisée « aventure », en tant qu’anthropologie de l’incertitude dont le dessein premier a été de saisir les identités labiles et vagabondes de migrants se désignant aventuriers – identité ramifiée à des pratiques, des désirs et des conceptions spécifiques de l’Ailleurs. En serrant de près le sens des discours des acteurs et des témoins rencontrés, en analysant des biographies fouillées d’aventuriers afin de restituer la dimension subjective du vécu migratoire, et en observant in vivo les creusets sociaux et les foyers de significations où naissent les désirs d’extraversion, nous donnons à voir et à comprendre les usages que font les aventuriers de l’Ici et de l’Ailleurs, ainsi que les rapports à eux-mêmes, aux autres et au monde qu’ils produisent. À rebours du poncif misérabiliste, nous dévoilons ainsi une philosophie de la mobilité qui nous incline à penser l’aventurier comme sujet-auteur de son existence et l’aventurisme comme un éthos et une forme rituelle d’esthétisation, de narration, de conquête et d’exposition de soi. / Ethnology of the adventure. Contemporary practices of male mobility and identity buildings in West Africa (Burkina Faso) – This PhD thesis features an ethnology of migratory phenomena in West Africa which promotes a study of the mobility locally called “adventure” as an anthropology of the uncertainty. The initial purpose was to grasp labile and roaming identities of migrants who refer to themselves as adventurers – identity branched to specific practices, desires and conceptions of the Elsewhere. In approaching as closely as possible the meaning of the speeches of the actors and the witnesses, in analyzing thick biographies of adventurers in order to reproduce the subjective dimension of their migratory experience, and in observing in vivo the social contexts and the crucibles of meanings where the desires of extroversion arise, we give to see and understand the adventurers’ uses of the Here and the Elsewhere, as well as their relationships to themselves, to others and to the world they generate. Contrary to the cliché painting migrations in the most miserable way, thus we unveil a philosophy of the mobility that makes us inclined to think the adventurer as a subject-author of his existence and the adventurism as an éthos and a ritual form of self-aestheticization, self-narrative, self-conquest and self-exposure.

Ce projet de recherche souhaite démontrer qu’une nouvelle ère s’instaure (la postmodernité) où l’imaginaire initiatique réinvente une nouvelle forme de socialisation. Il s’agit de questionner les pratiques sociales liées à l’imaginaire initiatique (cosplay, japan expo, jeux en ligne), ainsi que l’impact provoqué dans la vie quotidienne des individus. En fait, on se demandera si l’imaginaire initiatique entraîne des changements de comportements, crée des habitus. Ce qu’il est nécessaire de souligner, c’est que l’homme postmoderne accède au domaine du sacré, entreprend une démarche initiatique, à partir de ce qu’il vit dans les mondes fictionnels. Cette manière d’être est tout à fait légitime, et s’apparente sans aucun doute à une nouvelle façon de vivre l’initiation des sociétés archaïques, l’essentiel étant de retrouver un sens magique du monde. Nous serons ainsi amener à interroger la fonction fantastique de l’imaginaire initiatique, c’est-à-dire sa capacité à ordonner symboliquement un sens authentique à l’existence sociale et permettre à l’individu de dépasser l’aspect tragique de la réalité opaque. / This research project would like to show that a new era is established (postmodernity) where initiatic imaginary reinvents a new form of socialization. It’s a matter of questioning social practices related to initiatic imaginary (cosplay, japan expo, plays on line), as well as the impact caused in the everyday life of individuals. In fact, we shall wonder whether the initiatic imaginary involves changes of behaviors, creates habitus. What it is necessary to underline, it is that the postmodern man reaches the field of sacred, takes an initiatic step, from what he lives in the fictional worlds. This way of being is completely legitimate, and is connected without any doubt with a new way of living archaic initiation, the main thing being to find a magic direction of the world. Thus we shall be brought to question fantastic function of initiatic imaginary, that is its capacity symbolically to order an authentic direction to social existence and to allow individual to overtake the tragic aspect of opaque reality.

Ces dernières décennies ont été marquées par la découverte de la transcription dite « cachée » ou « pervasive ». Il a été en effet montré que la majeure partie du génome des eucaryotes est transcrite, donnant naissance à la formation de nombreux ARNs non-codants. La délimitation des unités de transcription apparait essentielle dans le contrôle de l’expression des gènes mais également dans le maintien de l’intégrité des processus associés à l’ADN en limitant notamment l’apparition de conflits avec la transcription. Dans ce contexte, l’initiation et la terminaison de la transcription représentent des étapes clés dans le partitionnement du génome et le métabolisme des ARNs. Nous avons montré que certains facteurs de transcription, appelés GRFs (General Regulatory Factors) chez la levure S. cerevisiae, jouent un rôle important dans le contrôle de la transcription pervasive à la fois au niveau de l’initiation mais également de la terminaison de la transcription et sont également requis pour assurer la fidélité de la transcription des gènes codant les ARN messagers. Nous avons prouvé que les GRFs liés au niveau des régions promotrices sont capables d’induire la terminaison de la transcription en bloquant physiquement la progression d’ARN polymérases issues de la translecture des terminateurs situés en amont. D’après nos études, cette voie de terminaison appelée « roadblock » est très répandue à l’échelle du génome et joue un rôle important dans la protection des promoteurs contre l’interférence transcriptionnelle. Nous avons également découvert que les GRFs limitent la transcription pervasive en obstruant les sites d’initiations ectopiques situés à proximité de leur site de fixation sur l’ADN. Ces facteurs sont aussi impliqués dans le contrôle de l’expression des gènes codants en favorisant l’utilisation de sites d’initiations les plus appropriés, c’est-à-dire, permettant la synthèse d’ARNs ayant un fort potentiel codant. Le rôle des GRFs dans le contrôle de l’initiation apparait intimement lié à leur capacité à correctement positionner les nucléosomes au niveau des promoteurs en collaboration avec les facteurs de remodelage de la chromatine. / The last decades have been marked by the discovery of pervasive transcription. Indeed, many studies have shown that transcription by RNA polymerase II is not restricted to annotated regions but is widespread in eukaryotic genomes, leading to the production of a plethora of non-coding RNAs. Precise delimitation of transcriptional units appears to be essential to ensure robust fidelity of gene expression and to maintain the integrity of DNA-associated events by preventing the occurrence of conflicts with transcription. In this respect, accurate transcription initiation and termination represent crucial mechanisms to partition the genome and define the correct processing of RNA molecules. Here, we show that yeast general regulatory factors (GRFs), a class of highly expressed transcription regulators, control pervasive transcription at the level of initiation and termination and are also involved in the fidelity of initiation of mRNA-coding genes. We demonstrate that GRFs bound at promoter regions can elicit transcription termination by physically impeding the progression of polymerases mainly deriving from readthrough transcription at upstream canonical termination sites. We provide evidence that this termination pathway named roadblock is widespread throughout the yeast genome and protects promoter regions from transcriptional interference. Furthermore, we establish that the presence of general regulatory factors also limits pervasive transcription at the level of initiation, notably by occluding spurious transcription start sites present in the vicinity of their binding sites. We also unveil the importance of these factors in promoting correct transcription start site selection at mRNA-coding genes thus favouring the synthesis of transcripts with an appropriate coding potential. Finally, we determine that the role of GRFs in controlling proper initiation is intimately linked to their ability to correctly position nucleosomes in promoters, a role that occurs independently from but in cooperation with chromatin remodelers.

Résumé : L’immunothérapie anti-PD1 est à l’origine de résultats cliniques impressionnants dans le traitement de certains cancers comme le mélanome métastatique ou le lymphome Hodgkinien. Néanmoins, les rechutes sont fréquentes et certaines tumeurs y sont d’emblée résistantes. Par ailleurs, l’étude du complexe d’initiation de la traduction eIF4F gagne de plus en plus d’intérêt dans le domaine du cancer. En effet, eIF4F joue un rôle fondamental dans la biologie des cancers grâce au contrôle sélectif de la synthèse de protéines impliquées dans le développement tumoral.Dans cette étude, nous montrons que l’inhibition du complexe eIF4F, en plus d’avoir un effet antitumoral directe via l’inhibition de la croissance tumorale in-vitro, a une action indirecte grâce à l’inhibition de l’expression de PD-L1 sous IFN-g, évitant ainsi le blocage des lymphocytes cytotoxiques suite à l’engagement PD-1/PD-L1. Dans un modèle murin de mélanome, nous avons montré une inhibition de la croissance tumorale grâce à l’inhibition de l’expression de PD-L1, uniquement dans des souris immunocompétentes, montrant ainsi le rôle fondamental du système immunitaire. Nous avons ensuite identifié la voie de régulation de PD-L1 par eIF4F via une régulation traductionnelle de l’ARNm de STAT1, principal facteur de transcription de PD-L1 sous IFN-g.Cette étude apporte une nouvelle preuve de l’intérêt des inhibiteurs d’eIF4F dans le cancer en démontrant leur effet immunothérapeutique via l’inhibition de PD-L1, évitant ainsi l’interaction PD-1/PD-L1 qui conduit à l’échappement des tumeurs. Ces résultats ouvrent la voie vers de nouvelles stratégies dans la lutte contre le cancer. / The eukaryotic translation initiation complex eIF4F is subject of an increased interest in the field of cancer. This heterotrimeric complex, comprising the RNA helicase eIF4A, the cap-binding protein eIF4E and the scaffold protein eIF4G, is known to be more abundant and active in tumor cells than non-malignant counterparts.In a previous work, we showed that this complex is implicated in the resistance to melanoma-targeted therapies (Boussemart et al, Nature 2014). Furthermore, it is implicated in the resistance to various chemotherapies. Thus, agents targeting the eIF4F complex appear as promising tools in the field of cancer therapy.On the other hand, immunotherapy, by (re)stimulating and enhancing the host immune system against tumors is giving good clinical results in oncology treatment and appears nowadays as the most promising approach to fight cancer, especially anti-PD1 treatment. Even though immunotherapy has demonstrated remarkable results in curing some established cancers, such as advanced melanoma or Hodgkin’s lymphoma, many tumors relapse or fail to respond. It is thus important to still look for a new strategy enhancing the efficacy of actual treatments. Here, we propose to study the impact of inhibiting the eIF4F complex on the tumor-specific immune response.

MA (Psychology) / Department of Psychology

Les paradigmes expérimentaux de l'initiation de la marche (IM) et du rattrapage de l'équilibre (RE) consécutif à une chute-avant ont été choisis pour identifier les adaptations motrices éventuelles chez les enfants souffrant de lésions cérébrales (PC) bilatérales et unilatérales vs. les enfants avec un développement normal (DN). Les enfants PC et DN étaient âgés entre 5 et 16 ans. Les enfants PC avaient une marche autonome sans aide technique. Les résultats montraient que, dans l'IM, les différents groupes d'enfants présentent des patterns biomécaniques et d'activités EMG comparables lorsque l'appui est sain. Plus particulièrement, la présence d'un freinage de la chute pendant la phase pendulaire, qui est l'indice qui caractérise la maturation du processus de la marche, chez les plus jeunes PC indique l'absence de retard. Lorsque l'appui est sur le côté lésé, le freinage est absent voire faible. Cette absence de freinage peut être imputée à la modification de l'appui en équin. De même, les tracés biomécaniques et EMG dans RE montraient des patterns comparables entre les différents groupes d'enfants. Toutefois une différence remarquable peut être distinguée par rapport à l'activité EMG des muscles fléchisseurs plantaires et dorsaux des adultes. Chez les adultes, la chute provoque une réponse précoce bilatérale des Soleus accompagnée parallèlement d'une activité de faible amplitude des TA. Chez les enfants, les TA montraient une importante bouffée EMG en même temps que la bouffée du Soleus. La suppression de cette bouffée précoce lorsque la chute est rapidement arrêtée annihilant l'exécution du pas suggère que les enfants DN et PC déclencheraient en même temps la réaction à la chute et le programme d'initiation du pas. Si les enfants PC pouvaient réalisés les deux tâches motrices, exécution du pas normal et exécution du pas provoqué, c'est semble-t-il grâce à la toxine botulique qui avait permis à l'enfant PC de se mettre debout libre, apprendre à contrôler son équilibre postural et la marche. / Experimental paradigms of gait initiation (GI) and balance recovery (BR) following a forward-fall were chosen to study motor adaptation in cerebral palsy (CP) children as compared to typical development children (TD). Children age ranged between 5 and 16 y.o. PC children walked independently. Results in GI showed that biomechanical and EMG pattern are comparable between the different groups when stance foot was on sound side. More particularly, the presence of fall braking during swing phase, which is a maturation index of gait process, in young CP indicates that there is no delay. When stance foot is on affected side, fall braking is absent that can be explained by equines foot. In BR, biomechanical and EMG traces are comparable between the different groups. However, if compare to adults, there is a striking difference in the EMG patterns. In adults, the fall elicited a bilateral burst of EMG in Soleus muscle, in parallel Tibialis anterior (TA) showed concomitant EMG activity but with lower amplitude. In children, TA showed a burst of EMG activity in parallel to SOL. This TA EMG burst was suppressed whether the fall was arrested annihilating the stepping. This result suggests that children trigger at the time the fall reaction program and the stepping program. If CP children can carry out both motor tasks, normal stepping and provoked stepping, this was thanks to use of botulinum toxin which help children standing up. Then, the mechanism of balance control can operate, and supporting gait acquisition.