Thesis (B.A. (Hons.))--University of Adelaide, Dept. of English, 1994? / Includes bibliographical references (leaves 43-45).

Zugl.: Marburg, Univ., Magisterarbeit

Le virus de la dengue (DENV) et le virus Zika (ZIKV) sont deux virus transmis par le moustique et responsables de maladies importantes chez l’Homme. En absence de vaccin et de traitements antiviraux efficaces, ces pathogènes représentent des problèmes de santé publique majeurs. Les bases moléculaires des interactions qu’établissent le DENV et ZIKV et la cellule hôte lors de l’entrée virale sont peu connues. Notre laboratoire a récemment identifié, les protéines TIM (TIM-1 et TIM-4) et TAM (Tyro3 et Axl), deux familles de récepteurs à la phosphatidylsérine (PtdSer) impliqués dans la reconnaissance et l’élimination des cellules apoptotiques par phagocytose, comme de nouveaux facteurs d’entrée du DENV. Les récepteurs TIM et TAM permettent l’infection par le DENV en interagissant avec la PtdSer associée aux virions selon un mécanisme similaire à la reconnaissance des cellules apoptotiques (mimétisme apoptotique). L’objectif général de mon travail de thèse a été d’explorer les mécanismes moléculaires et cellulaires par lesquels TIM-1 et Axl médient l’entrée des flavivirus. A l’aide de techniques d’imagerie en temps réel nous avons montré que TIM-1 et DENV sont co-internalisés et que TIM-1 joue un rôle actif dans l’entrée du DENV. Notamment, nous avons montré que deux résidus lysine présentes dans le domaine cytoplasmique de TIM-1 sont importantes pour l’ubiquitination du récepteur et pour l’endocytose du virus. La recherche de partenaires de TIM-1 par des études de spectrométrie de masse a permis d’identifier STAM, un membre du complexe ESCRT-0 impliqué dans le trafic des récepteurs ubiquitinés, comme facteur important pour l’infection. Collectivement, nos résultats suggèrent très fortement que TIM-1 est le premier récepteur bona fide caractérisé pour le DENV.Identifier les facteurs d’entrée du ZIKV représente un enjeu majeur dans la compréhension du tropisme et de la pathogénèse associée à ce virus. Nous avons montré que le récepteur Axl est essentiel pour l’entrée du ZIKV dans les cellules microgliales, les astrocytes du cerveau humain en développement ainsi que dans les fibroblastes de la peau. Nos études ont démontré un double rôle du récepteur Axl dans l’infection par ZIKV. Axl lie et permet l’internalisation des particules virales, mais aussi, contribue à l’établissement d’un environnement favorable à la réplication virale en inhibant la réponse immunitaire innée. En conclusion, ce travail a contribué à améliorer notre compréhension des mécanismes d’entrée des virus DENV et ZIKV. Nos résultats indiquent que ces deux virus exploitent plusieurs récepteurs aux phospholipides pour initier leur cycle infectieux, ce qui pourrait contribuer à l’élargissement de leur tropisme. / Dengue virus (DENV) and ZIKA virus (ZIKV) are two mosquito-borne viruses responsible for important diseases in humans. Since there is currently no vaccine neither antiviral treatment available against these human pathogens, they are two major health concerns. The molecular basis of DENV and ZIKV host cell interactions leading to virus entry are poorly understood, hampering the discovery of new targets for antiviral intervention. Our laboratory recently discovered that TIM (TIM-1 and TIM-4) and TAM (Tyro3 and Axl) proteins, two receptor families that contribute to the phosphatidylserine (PtdSer)-dependent phagocytic removal of apoptotic cells, are DENV entry factors. TIM and TAM receptors mediate DENV infection by interacting with virion-associated PtdSer through a mechanism similar to the recognition and engulfment of apoptotic cells by phagocytes (viral apoptotic mimicry). The general objective of my PhD was to establish a detailed understanding of the molecular mechanisms by which TIM-1 and Axl mediated infection. Using live imaging, we demonstrated that TIM-1 and DENV are co-internalised and TIM-1 play an active role during DENV endocytosis. We showed that TIM-1 cytoplasmic domain is essential for DENV internalization, especially, we identified two lysine residues that are essential for TIM-1 ubiquitination and DENV endocytosis. Proteomic analysis of TIM-1 interacting partners identified STAM, a member of the ESCRT-0 complex involved in intracellular sorting of ubiquitinated cargos, as an essential host factor for DENV infection. Collectively our results establish TIM-1 as the first identified DENV bona fide receptor.Identifying ZIKV entry factors represents a major challenge in the understanding of ZIKV tropism and pathogenesis. We showed that Axl is responsible for ZIKV infection of microglial cells and astrocytes in the human developing brain and primary fibroblasts in human skin, suggesting an important role of this receptor during ZIKV life cycle. We also highlighted the dual role of the Axl receptor in ZIKV infection, which simultaneously promotes viral entry and dampens the innate immune response to facilitate a post entry step of the ZIKV life cycle. In conclusion, this work provided new insights in our understanding of the DENV and ZIKV entry program. Both viruses engage phospholipid receptors for their infectious entry, providing a rational to ascertain therapeutic strategies targeting virion-associated phospholipids.

Thesis (M.A.)--Villanova University, 2009. / English Dept. Includes bibliographical references. Mode of access: World Wide Web.

vi, 73 p. A print copy of this thesis is available through the UO Libraries. Search the library catalog for the location and call number. / At the beginning of the 20th century, several critics suggested that the genre of the novel was in a state of crisis. A vigorous discussion about whether the genre of the novel as a whole had to be done away with erupted and produced contradicting frameworks. A textual analysis of two recent books by Swiss writer Tim Krohn, Quatemberkinder (1998) and Vrenelis Gartli (2004), reveals that the time of great epics in the form of the novel has not yet come to an end. Krohn's novels portray a new approach to counter what Lukacs called "transcendental homelessness." By combining oral Swiss legends with historical facts and magic realism Krohn is able to achieve a new wholeness the modem novel lacked. His novels portray a possible solution for overcoming the "crisis of the novel." / Committee in Charge: Prof. Susan C. Anderson, Chair; Prof. Kenneth Scott Calhoon; Prof. Martin Klebes

T cell immunoglobulin and mucin (Tim) family molecules are cell membrane proteins with four functional Tim family members in mouse, and three in human. They are preferentially expressed on immune cells with Tim1 on Th2 cells, Tim3 on Th1 cells and Tim4 on antigen-presenting cells (APCs). They have several roles, including regulating immune responses and mediating phagocytosis of dead cells. However, little is known about them beyond these two species, and nothing outside mammals. To investigate the Tim family in the chicken, the genes were identified and cDNAs cloned. Differently to mammals, the chicken genome only contains genes for Tim1 and Tim4. Chicken Tim1 (chTim1) has similar mRNA expression patterns to those of mammalian Tim1 in lymphoid tissues and immune cells. Interestingly, chTim4 has at least four splice variants – an extra short isoform (chTimeS) lacking exons 5, 6, 7 and 8, a short isoform (chTim4S) without exons 3, 4 and 5, a long isoform (chTim4L) with all exons and an extra long isoform (chTim4eL), which is similar to chTim4L but with a longer exon 3. The chTim4S is a homologue of mammalian Tim4 with constitutive expression in lymphoid tissues and immune cells; other chTim4 variants showed inducible or cell-specific expression patterns. Like mammalian Tim4, chTim4S is expressed by APCs; but differently to mammals, chTim4S is also expressed by γδ T cells, suggesting a unique role for chTim4 in this population of T cells. The biological activities of the chicken Tim family molecules were also investigated using chTim-Ig fusion proteins. Like mammals, chTim1 and chTim4S fusion proteins can specifically recognise phosphatidylserine (PS), an indicator of apoptotic cells, suggesting they are PS receptors. Pre-incubation with PS blocked binding of the chTim4S fusion protein to PS-exposing apoptotic cells. Physiologically, recognition of PS by the chTim proteins mediates apoptotic cell clearance, which was demonstrated using chTim-transfected fibroblast cells (3T3), which significantly increased their uptake of apoptotic cells compared with untransfected cells. The chTim4-Ig fusion protein also had costimulatory activity on chicken T cells. Monoclonal antibodies against the chTim proteins were generated. They specifically recognise their own antigen tested intensively by different immunological assays. ChTim4L is expressed intracellularly in freshly-isolated splenocytes rather than on the surface, whereas PMA-stimulated splenocytes express chTim4S and chTim4L on the cell surface. Like mammals, chicken splenic macrophages also express chTim4S and chTim4L. Both of them are also expressed by bone marrow-derived macrophages but not bone marrow-derived DCs. The chTim1 protein was detected at high levels in bursal cells and splenocytes by western blot analysis using polyclonal anti-chTim1 serum, which is consistent with its mRNA expression pattern through qRT-PCR analysis, suggesting B and T cells may express chTim1, consistent with its expression in mammals. Mammalian Tim1 is expressed on Th2 cells, its ligand, Tim4, on APCs; the interaction between them drives Th2 cell proliferation. The knowledge from this study will help to further dissect how the chicken’s Th2 responses are regulated through cell surface molecules.

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Diss. Ph. D. : Philosophy : Uppsala university : 1998. / Notes bibliogr. Index.

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