Gametogenesis is a highly complex process that requires stringent control of gene expression, in which translational regulation plays an essential role. Deleted in Azoospermia-like (DAZL) belongs to the DAZ family of RNA-binding proteins, which are restricted to germ cells, and regulate mRNA translation. Importantly, loss of function of these proteins results in infertility in both males and females in a wide variety of organisms. A model for the mechanism by which DAZL stimulates translation has been proposed based on work in Xenopus laevis (X. laevis) oocytes. In this model, DAZL functions by recruiting the translation initiation factor poly(A)-binding protein (PABP) to the 3’ untranslated region (UTR) of messenger RNAs. Simultaneous binding of PABP to Dazl and factors at the 5’ end confers a “closed-loop” mRNA conformation, which promotes translation initiation. To examine whether DAZL plays a similar role in mammals, co-expression of Dazl and PABP family members was investigated in fetal and adult mouse gonads. In contrast to X. laevis, mammals encode four cytoplasmic PABPs which share a similar domain organisation: PABP1, tPABP, ePABP and PABP4, of which PABP1 and PABP4 appear to be expressed in a wide range of tissues. Immunohistochemistry revealed that Dazl, Pabp1 and Pabp4 are all expressed in primordial germ cells (PGCs) but these show different expression patterns following germ cell sex differentiation. In adult testes Dazl is expressed in spermatogonia and spermatocytes, coinciding with the peak of Pabp4 expression. In contrast, the peak of Pabp1 expression occurs later than that of Dazl, with these proteins only being co-expressed in late pachytene and secondary spermatocyte phases. In adult ovaries, Pabp1, Pabp4 and Dazl are all expressed in the oocytes of primordial and primary follicles. Since both PABP family members are co-expressed with Dazl, the ability of DAZL to interact with PABP1 and PABP4 was investigated in vitro and in vivo. Surprisingly, these studies showed that DAZL discriminates between different PABP family members, only interacting with PABP1, providing the first report of a PABP-specific protein partner. Several putative DAZL mutations have been identified in patients with impaired fertility. Two of these mutations, I37A and R115G, are located in the RNA recognition motif (RRM), a domain which is found in many RNA-binding proteins and mediates both RNA and protein interactions. Thus, the role of these mutations in the ability of DAZL to stimulate translation was investigated. To this end, a translational target of human DAZL (hDAZL) was sought. The 3’UTR of growth differentiation factor 9 (hGDF9) mRNA was found to confer regulation by hDAZL and thus the ability of mutant DAZLs to stimulate reporter mRNAs containing this 3’UTR was examined. This revealed that both mutations compromised the ability of hDAZL to stimulate hGDF9 translation, suggesting a causative effect. These results were further confirmed in assays in which hDAZL is artificially tethered to mRNAs. The ability of mutant hDAZLs to stimulate translation in this assay was compromised suggesting that loss of function is, at least in part, due to impaired protein-protein interactions rather than altered RNA-binding. This work provides insights into the molecular mechanism by which DAZL stimulates the translation of specific mRNAs during mammalian gametogenesis and provides evidence that this function may play an important physiological role in human reproduction.

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Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

Les Entérovirus humains du groupe B (EV-B) et plus spécifiquement les virus Coxsackie B sont considérés comme une cause majeure des myocardites infectieuses aigues et chroniques dont 10% peuvent évoluer vers la cardiomyopathie dilatée (CMD). Les mécanismes moléculaires viraux impliqués dans la progression de la myocardite aiguë vers le stade de la CMD ne sont pas élucidés.L’analyse par séquençage NGS a montré chez 8 (33%) des 24 patients atteints de CMD inexpliquée l’existence de populations majoritaires tronquées de 19 à 50 nucléotides associées à des formes virales minoritaires complètes. La proportion de populations tronquées s’est révélée négativement corrélée au ratio ARN+/ARN- et à la charge virale. Des études immuno-histologiques et par hybridation in situ des tissus cardiaques ont montré que le clivage de la dystrophine était uniquement retrouvé dans les cardiomyocytes infectés par les EV-B. Pour étudier les activités de réplication des populations d’EV-B persistants, un réplicon (CVB3-emGFP) a été généré à partir d’une souche cardiotrope (CV-B3/28). La transfection d’ARN de synthèse complets et tronqués (d50) dans des cultures de cardiomyocytes humains primaires a mis en évidence des mécanismes de recombinaison et/ou de trans-complémentation entre ces 2 formes virales induisant de faibles activités de réplication.Nos résultats démontrent l’existence de mécanismes de coopération moléculaire entre des populations d’EV-B tronquées et complètes qui pourraient expliquer la mise en place du mécanisme de persistance virale observée au cours de la phase clinique de CMD. Ces résultats pourraient contribuer au développement de nouvelles stratégies thérapeutiques pour prévenir et traiter les infections cardiaques à EV-B. / Enteroviruses group B (EV-B) and more specifically Coxsackievirus B are recognized as major causes of acute and chronic infectious myocarditis, which 10% may progress towards dilated cardiomyopathy (DCM). Viral molecular mechanisms involved in the progression from acute myocarditis to the clinical stage of DCM remain unknown.Deep sequencing analysis showed in 8 (33%) of 24 unexplained DCM patients the existence of major CVB3 populations with deletions of 19 to 50 nucleotides associated with a minority of complete viral forms. The proportion of deleted viral populations was negatively correlated with RNA+/RNA- ratio and the viral load levels. Immuno-histological and in situ hybridization assays of DCM cardiac tissues demonstrated that the cleavage of dystrophin was found only in cardiomyocytes infected with EV-B. To study the replication activities of persistent EV-B populations, a replicon (CVB3-emGFP) was generated from a cardiotropic strain (CV-B3/28). Transfection of synthesized complete and truncated (d50) viral RNAs in primary human cardiomyocytes cultures revealed mechanisms of recombination and / or trans-complementation between these two viral forms inducing low replication activities.In conclusions, our original results demonstrated the existence of new molecular mechanisms of cooperation between EV-B deleted and complete viral populations that could explain the development of a viral persistence mechanism observed during the clinical phase of DCM. These findings may contribute to the development of new therapeutic strategies to prevent and treat persistent heart EV-B infections.

<p dir="ltr">The presence of digital devices in various settings, from workplaces to personal spaces, necessitates reliable and secure data storage solutions. These devices store data on non-volatile media like Solid State Drives (SSDs) and Hard Disk Drives (HDDs), ensuring data preservation even after power loss. Files, fundamental units of data storage, are created, modified, and deleted through user activities like application installations or file management. File systems, acting as the backbone of the system, manage these files on storage devices.</p><p dir="ltr">This research explores how three key factors: (1) different operating systems running various file system types (ext4, NTFS, FAT, etc.), (2) different disk types (SSD and HDD), and (3) common user activities (system shutdowns, reboots, web browsing, downloads, etc.) influence the persistence of deleted files.</p><p dir="ltr">This research aims to fill a gap in the understanding by looking at how these factors influence how quickly new information overwrites deleted files. This is especially important for digital forensics, where investigators need to be sure they can find all the evidence on a device. The research will focus on how operating systems handle deleted files and how everyday activities affect the chances of getting them back. This can ultimately improve data security and make digital forensics more reliable.</p>

Made available in DSpace on 2016-04-29T14:27:27Z (GMT). No. of bitstreams: 1 Adinael Carlos Miguel.pdf: 459130 bytes, checksum: dfed08cf8da240ea0f0ca6e06d580daa (MD5) Previous issue date: 2015-08-12 / In this research intends to focus on the importance of the Kingdom of God on Christology in Latin America, that is, we want to do work that discusses the context of Christology in Latin American reality, and the operation and structure of the Kingdom of God in Jesus, and the proposal presented by him, compared to the reality of our continent, however, to develop this research we report a Christology that reveals a historical Jesus, as from its history, we have knowledge of their mission, and analyze the situation of the poor of Latin America. Is portrayed the proposal of the Kingdom of God by Jesus, who were the recipients, so their preference for the excluded of his time. Conclude commenting on the Kingdom of God in view of Christology in Latin America, recalling the reality faced by this continent, and comparing to the situation of disadvantaged poor in Jesus' time / Nessa pesquisa pretende enfocar a importância do Reino de Deus na Cristologia da América Latina, ou seja, queremos realizar um trabalho que relata o contexto da Cristologia na realidade Latino-americana, e o funcionamento e estrutura do Reino de Deus na época de Jesus, e a proposta apresentada por Ele, comparando com a realidade do nosso continente atual. Entretanto, para desenvolver esta pesquisa iremos relatar uma Cristologia que revela um Jesus histórico, pois a partir da sua história, teremos conhecimento da sua missão, e analisar a situação dos pobres da América Latina. Será retratada a proposta de Reino de Deus apresentada por Jesus, quem eram seus destinatários, e por isso a sua preferência pelos excluídos da sua época. Concluiremos comentando sobre o Reino de Deus na perspectiva da Cristologia da América Latina, relembrando a realidade vivenciada por este continente, e comparando com a situação dos pobres desfavorecidos na época de Jesus

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