Thesis (Ph. D.)--West Virginia University, 2009. / Title from document title page. Document formatted into pages; contains vii, 104 p. : ill. Includes abstract. Includes bibliographical references.

The function and evolution of introns have been topics of great interest since introns were discovered in the 1970s. Introns that interrupt protein-coding regions have the most obvious potential to affect coding sequences; therefore, their evolution have been studied most intensively. Splicing of introns within untranslated regions does not contribute directly to the diversity of proteins, yet ~35% of human transcripts contain introns within the 5' untranslated region (UTR). The evolution and possible functions of 5'UTR introns (5UIs) remain largely unexplored. Here we undertook a genome-wide functional analysis of 5UIs. Our main results are as follows: First, the distribution of these introns in the human genome is nonrandom. While genes with regulatory roles are enriched in having 5UIs, genes encoding proteins that are targeted to the endoplasmic reticulum and mitochondria are surprisingly depleted of these introns. Second, we offered and supported a model whereby gene encoding secretory and nuclear-encoded mitochondrial proteins share a common regulatory mechanism at the level of mRNA export, which is dependent on the absence of 5'UTR introns. Specifically, the upstream element in a given transcript, be it an intron or RNA elements near the 5' end of coding sequences (CDS), dictates the mRNA export pathway used. Finally, we discovered a strong correlation between existence of 5'UTR introns and sequence features near the 5' end of CDS. We developed an integrated machine-learning framework that can predict absence of 5UIs using solely the sequence near the 5' end of CDS. Our model achieved >80% accuracy when validated against nuclear-encoded mitochondrial transcripts. Specific RNA elements predictive of 5UI absence are found in ~40% of human transcripts spanning a wide spectrum of functions. By analyzing hundreds of large-scale datasets, we functionally characterized the transcripts with these RNA elements; revealing their association with translational regulation. These RNA elements were bound by proteins interacting with the Exon Junction Complex in vivo suggesting a molecular mechanism that links these elements to their downstream effects in mRNA export and translational regulation. While some 5'UTR introns might be evolving neutrally, our results, taken together, suggest that complex evolutionary forces are acting on this distinct class of introns.

Human Immunodeficiency Virus type I genome consists of two identical RNA molecules that are non-covalently linked to form a dimer. HIV-1 immature and mature genomic RNA (gRNA) dimers were found in protease defective (PR -) and wild type virions, respectively, and the 5'untranslated region (5' UTR) was shown to play key roles during the genome dimerization process; but the dimerization mechanism still remains to be clarified My research project is to characterize the dimerization process and the role of 5' UTR in genome dimerization in virions produced by tissue culture cells. I'll firstly show the dimer maturation processes of HIV-1 gRNA isolated from newly released to grown-up (≥10h old) wild type, PR-, and SL1 defective (DeltaIDS) virions respectively. The results showed that HIV-1 gRNA dimer maturation process was protease-dependent and involved multiple steps: from low to high dimerization level and dimer thermostability, and from low dimer mobility to intermediate and high mobility. PR- virions did not freeze gRNA conformation in the primordial nascent state and gRNA changed from monomeric in newly released virions to half dimeric in grown-up virions, which showed that genome was packaged in the form of monomeric RNA or fragile dimers, more thermolabile than immature dimers in grown-up PR- virions. DeltaDIS inhibited gRNA dimerization by about 50% in newly released virions, though grown-up DeltaDIS gRNA was fully dimeric, which indicated that the DIS played the initiation role in gRNA dimerization in HIV-1 virions. The gRNA dimerization rate in PR- or DeltaDIS virions was much slower than that in wild type virions. These results show for the first time the whole process of dimer maturation after virion release, the gRNA conformation rearrangement in PR- virions, and the initiation role of the DIS in HIV-1 virions. Next, I'll provide a rather systematic search for the contribution of different regions in 5' UTR to HIV-1 gRNA dimerization by studying selected mutations singly or together with defective SL1. The results showed that the 5'trans-activation response element (5'TAR) was directly involved in gRNA dimerization, and a long distance base-pairing interaction between a sequence in U5 region (nts105-1l5) and another around the initiation codon of the gag gene (nts334-344) was structurally contributive to gRNA dimerization. Deletions of sequences around the 3'end of Primer Binding Site (PBS) stem-loop moderately decreased gRNA dimerization level. Other sequences in 5' UTR except DIS/SL1, which was previously known to play important roles, didn't show any systematic role. Here the results suggested that the absence of inhibition on gRNA dimerization level with defective DIS might be the compensation of the direct role of 5'TAR; and wild type-like dimerization level of DeltaTAR must be the direct contribution of the DIS.

Thesis (Ph.D. in Biochemistry & Molecular Genetics) -- University of Colorado at Denver and Health Sciences Center, 2006. / Typescript. Includes bibliographical references (leaves 119-131). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;

Thesis (Ph. D.)--University of Washington, 1999. / Vita. Includes bibliographical references (leaves 126-146).

The purpose of this project was to identify and evaluate nucleotide sequences enhancing translation of proteins in Chinese hamster ovary (CHO) cells. Candidate sequences were placed in the 5´-untranslated region (UTR) or 3´ UTR respectively and evaluated in a CHO-based expression system with a fluorescent Fc-fusion protein as a model protein.Five plasmid vectors were constructed, two of which designed to have a randomized nucleotide library in their 5´ and 3´ UTR respectively, and three of which designed to hold varying repeats of a known enhancing translation (ET) sequence in their 5´ or 3´ UTR. The plasmid constructs were transfected into CHO cells and the protein expression was analyzed both by fluorescence intensity in single cells using flow cytometry and in bulk by monoclonal antibody titer analysis based on Protein A affinity.The main result is that both flow cytometry and titer analysis indicate that insertion of five repeats of the ET in the 5´UTR has a negative effect on protein expression as compared to the control which had no ET repeats. Results related to the insertion of three ETs in the 5´ UTR were ambiguous. The titer analysis indicated that it had a negative effect on the protein expression compared to the control which had no ET repeats, whereas the flow cytometry results suggest that the effect is negligible. Transfection of library plasmids was unsuccessful; hence no library expression analysis results were achieved. Due to the time constraints of the project, the reason for the unsuccessful transfection of library plasmids was not investigated, but the LTX transfection method is stated as a highly plausible cause.Based on the outcome of this study, two recommendations for future work are suggested. The first one is to continue the focus on UTR sequences in terms of library screening, and to improve the method of transfecting library plasmid constructs into CHO cells using lipofection. The second suggestion for further studies is to test different UTR sequence lengths without involving potential ETs, to rule out the effect and positions of the ETs and investigate the expressional effect of UTR length solely.

abstract: Plants are a promising upcoming platform for production of vaccine components and other desirable pharmaceutical proteins that can only, at present, be made in living systems. The unique soil microbe Agrobacterium tumefaciens can transfer DNA to plants very efficiently, essentially turning plants into factories capable of producing virtually any gene. While genetically modified bacteria have historically been used for producing useful biopharmaceuticals like human insulin, plants can assemble much more complicated proteins, like human antibodies, that bacterial systems cannot. As plants do not harbor human pathogens, they are also safer alternatives than animal cell cultures. Additionally, plants can be grown very cheaply, in massive quantities. In my research, I have studied the genetic mechanisms that underlie gene expression, in order to improve plant-based biopharmaceutical production. To do this, inspiration was drawn from naturally-occurring gene regulatory mechanisms, especially those from plant viruses, which have evolved mechanisms to co-opt the plant cellular machinery to produce high levels of viral proteins. By testing, modifying, and combining genetic elements from diverse sources, an optimized expression system has been developed that allows very rapid production of vaccine components, monoclonal antibodies, and other biopharmaceuticals. To improve target gene expression while maintaining the health and function of the plants, I identified, studied, and modified 5’ untranslated regions, combined gene terminators, and a nuclear matrix attachment region. The replication mechanisms of a plant geminivirus were also studied, which lead to additional strategies to produce more toxic biopharmaceutical proteins. Finally, the mechanisms employed by a geminivirus to spread between cells were investigated. It was demonstrated that these movement mechanisms can be functionally transplanted into a separate genus of geminivirus, allowing modified virus-based gene expression vectors to be spread between neighboring plant cells. Additionally, my work helps shed light on the basic genetic mechanisms employed by all living organisms to control gene expression. / Dissertation/Thesis / Doctoral Dissertation Microbiology 2017

La traduction est une affaire culturelle. De prime abord, elle se présente comme la recherche d’équivalents lors du passage d’une langue à l’autre. Mais si on se penche sur le travail qu’effectuent les traducteurs, on se rend compte rapidement que traduire exige des opérations bien plus complexes. Ainsi, un texte littéraire rédigé en une langue ne peut pas être traduit vers une autre langue sans que le traducteur ou la traductrice fasse attention au contenu culturel que véhicule la langue. Un même message est communiqué de deux manières différentes par deux peuples issus de cultures distinctes. Ainsi opère un système très complexe qui déborde le champ de la linguistique et s’enracine dans la culture qu’exprime chaque langue. Toutes les actions humaines, la communication, les sentiments, les réactions, la compréhension, l’interprétation (pour en nommer quelques-unes) ont la culture pour fondement.Intitulée « L’épreuve de l’étranger : traductions françaises d’écrivains sri lankais contemporains de langue anglaise », notre thèse a pour mots-clés : « culture », « sri lankais » et « traduction ». Elle a pour point de départ une question formulée en quelques mots simples : comment transmettre en français la culture sri lankaise ? Ces deux cultures sont distantes sur le plan géographique mais aussi pour ce qui concerne leurs pratiques et leurs valeurs. C’est donc une rencontre entre l’Orient et l’Occident que permet la traduction par le truchement de la langue anglaise.Les deux œuvres que nous avons choisies pour notre corpus, Funny Boy de Shyam Selvadurai (1994) et Running in the Family de Michael Ondaatje (1982), sont imprégnées de culture sri lankaise. Nous avons tenté de mettre en évidence systématiquement les stratégies utilisées par les traducteurs pour préserver l’identité de la culture source. La tâche du traducteur n’est pas de dissimuler ou de minimiser les éléments culturels sri lankais mais de les rendre visibles dans ses textes. Par les stratégies qu’il utilise, le traducteur parvient à préserver la culture sri lankaise dans le texte d’arrivée, du moins dans des limites qu’il nous faudra également définir. Lawrence Venuti (2004 : 20) propose un système binaire, la traduction ethnocentrique ou domesticating (naturalisation) et la traduction éthique qui laisse place à l’étrangeté qu’il qualifie de foreignizing (dépaysement). Conserver les traces de l’œuvre originale est considéré comme la chose la plus importante. Nous nous plaçons ainsi entre les stratégies de naturalisation et de dépaysement. Traduire, c’est effectuer un travail qui « est ouverture, dialogue, métissage et décentrement » comme l’écrit Berman (1984 : 16), c’est aussi négocier un autre type de produit final.Dans un premier temps, nous verrons comment opère le dépaysement en tant que stratégie de traduction dans Drôle de garçon (Frédéric Limare et Susan Fox-Limare, 1998) de Shyam Selvadurai et Un air de famille (Marie-Odile Fortier-Masek, 1991) de Michael Ondaatje. Dans un deuxième temps, nous prêterons attention à la stratégie de la naturalisation qui rend la lecture plus fluide, en atténuant les différences trop importantes entre cultures. Dans un troisième temps, nous verrons de quelle manière certaines faits culturels restent intraduits dans les traductions pour diverses raisons que nous identifierons au cours de l’analyse, tout en repérant également les ellipses qui modifient le message d’origine. Notre analyse tentera de démontrer que la traduction est une rencontre entre les cultures : une rencontre qui se fait de manière fructueuse pour enrichir la littérature d’une culture nouvelle en permettant au lecteur un voyage vers une destination lointaine.Mots clés : culture, dépaysement, Funny Boy, Michael Ondaatje, naturalisation, Running in the Family, Shyam Selvadurai, Sri Lanka, traduction / Translation is a cultural matter. At first sight, it may appear as a search for equivalents in the transfer from one language to another. However, in depth translation analysis reveals much more than meets the eye. A literary text written in one language cannot be translated into another language without paying attention to its associated cultural background. It has become a truism today to say that individuals belonging to different cultures do not communicate in the same way; while the linguistic dimension is important, so is the cultural one, since cultural habits are at the root of all human actions.The title of this thesis, “The Trials of the Foreign: French Translations of Contemporary Sri Lankan Writers in English”, combines three key words: “culture”, “Sri Lanka” and “translation”. The broad research question we started out with is: how is it possible to convey Sri Lankan culture in French literary translation? The two countries are distant not only geographically but also in terms of practices and values. Therefore, a true encounter between East and West is at stake here, mediated by the English language, which the authors of the two Sri Lankan novels we study here chose as a medium of expression.In his or her attempt to identify viable equivalents of different cultural realities, the translator is confronted with decisions about whether differences should be mitigated or, on the contrary, preserved, in order to maintain the local colour. When cultural differences are smoothed over in translation and the target text contains very few traces, if any, of the source culture, the reader may have the impression of reading an original. On the other hand, when the source culture is given prominence, the translation has the potential to make the reader travel abroad, and gain new experience.The two literary works which make the object of our research, Michael Ondaatje’s Running in the Family (1982) and Shyam Selvadurai’s Funny Boy (1994), are imbued with Sri Lankan culture and pose significant challenges to translation. We draw on Lawrence Venuti’s (1995/2004) distinction between ethnocentric or domesticating translation (naturalisation) and foreignizing translation (dépaysement), while at the same time recognizing the importance of not taking this dichotomy for granted. And we assume, as Antoine Berman did, that translation is “openness, dialogue, blending and decentring” (1984: 16).We start by outlining a number of theoretical considerations about translation strategy, culture, and translating culture. We then carry out fine-grained analyses of the texts and endeavour to show how foreignization operates in Drôle de garçon (1998), the French translation by Frédéric Limare and Susan Fox-Limare of Selvadurai’s novel Funny Boy, and in Un air de famille (1991), the translation of Ondaatje’s Running in the Family by Marie-Odile Fortier-Masek. In the second part of our analysis, we focus on the strategy of domestication, which makes reading more fluent due to the mitigation of differences between cultures. Finally, we discuss some of the ways in which certain cultural facts remain untranslated, with implications for the integrity of the message, and the target readers’ experience of the text. We conclude that translation is indeed an encounter between cultures: a meeting that is fruitful and has the potential to enrich the literature of a new country, by allowing the reader to embark on a journey to a distant destination.Key words: culture, domestication, foreignization, Funny Boy, Michael Ondaatje, Running in the Family, Shyam Selvadurai, Sri Lanka, translation

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