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Dissertations / Theses on the topic 'Bioinformatics Molecular biology Bioinformatics'
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Pettersson, Fredrik. "A multivariate approach to computational molecular biology." Doctoral thesis, Umeå : Univ, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-609.
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Lopes, Pinto Fernando. "Development of Molecular Biology and Bioinformatics Tools : From Hydrogen Evolution to Cell Division in Cyanobacteria." Doctoral thesis, Uppsala universitet, Institutionen för fotokemi och molekylärvetenskap, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-110842.
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The use of fossil fuels presents a particularly interesting challenge - our society strongly depends on coal and oil, but we are aware that their use is damaging the environment. Currently, this awareness is gaining momentum, and pressure to evolve towards an energetically cleaner planet is very strong. Molecular hydrogen (H2) is an environmentally suitable energy carrier that could initially supplement or even substitute fossil fuels. Ideally, the primary energy source to produce hydrogen gas should be renewable, and the process of conversion back to energy without polluting emissions, making this cycle environmentally clean. Photoconversion of water to hydrogen can be achieved using the following strategies: 1) the use of photochemical fuel cells, 2) by applying photovoltaics, or 3) by promoting production of hydrogen by photosynthetic microorganisms, either phototrophic anoxygenic bacteria and cyanobacteria or eukaryotic green algae. For photobiological H2 production cyanobacteria are among the ideal candidates since they: a) are capable of H2 evolution, and b) have simple nutritional requirements - they can grow in air (N2 and CO2), water and mineral salts, with light as the only energy source. As this project started, a vision and a set of overall goals were established. These postulated that improved H2 production over a long period demanded: 1) selection of strains taking in consideration their specific hydrogen metabolism, 2) genetic modification in order to improve the H2 evolution, and 3) cultivation conditions in bioreactors should be exmined and improved. Within these goals, three main research objectives were set: 1) update and document the use of cyanobacteria for hydrogen production, 2) create tools to improve molecular biology work at the transcription analysis level, and 3) study cell division in cyanobacteria. This work resulted in: 1) the publication of a review on hydrogen evolution by cyanobacteria, 2) the development of tools to assist understanding of transcription, and 3) the start of a new fundamental research approach to ultimately improve the yield of H2 evolution by cyanobacteria.
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Sparring, Leonard. "Conquering Chemical Space : Optimization of Docking Libraries through Interconnected Molecular Features." Thesis, Uppsala universitet, Institutionen för biologisk grundutbildning, 2020. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-416977.
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Copied selected text to selection primary: The development of new pharmaceuticals is a long and ardous process that typically requires more than 10 years from target identification to approved drug. This process often relies on high throughput screening of molecular libraries. However, this is a costly and time-intensive approach and the selection of molecules to screen is not obvious, especially in relation to the size of chemical space, which has been estimated to consist of 10 60 compounds. To accelerate this exploration, molecules can be obtained from virtual chemical libraries and tested in-silico using molecular docking. Still, such methods are incapable of handling the increasingly colossal virtual libraries, currently reaching into the billions. As the libraries continue to expand, a pre-selection of compounds will be necessitated to allow accurate docking-predictions. This project aims to investigate whether the search for ligands in vast molecular libraries can be made more efficient with the aid of classifiers extended with the conformal prediction framework. This is also explored in conjunction with a fragment based approach, where information from smaller molecules are used to predict larger, lead-like molecules. The methods in this project are retrospectively tested with two clinically relevant G protein-coupled receptor targets, A 2A and D 2 . Both of these targets are involved in devastating disease, including Parkinson’s disease and cancer. The framework developed in this project has the capacity to reduce a chemical library of > 170 million tenfold, while retaining the 80 % of molecules scoring among the top 1 % of the entire library. Furthermore, it is also capable of finding known ligands. This will allow for reduction of ultra-large chemical libraries to manageable sizes, and will allow increased sampling of selected molecules. Moreover, the framework can be used as a modular extension on top of almost any classifier. The fragment-based approaches that were tested in this project performed unreliably and will be explored further.
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Yao, Xiaoquan. "Sequence features affecting translation initiation in eukaryotes: A bioinformatic approach." Thesis, University of Ottawa (Canada), 2008. http://hdl.handle.net/10393/27658.
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Sequence features play an important role in the regulation of translation initiation. This thesis focuses on the sequence features affecting eukaryotic initiation. The characteristics of 5' untranslated region in Saccharomyces cerevisiae were explored. It is found that the 40 nucleotides upstream of the start codon is the critical region for translation initiation in yeast. Moreover, this thesis attempted to solve some controversies related to the start codon context. Two key nucleotides in the start codon context are the third nucleotide upstream of the start codon (-3 site) and the nucleotide immediately following the start codon (+4 site). Two hypotheses regarding +4G (G at +4 site) in Kozak consensus, the translation initiation hypothesis and the amino acid constraint hypothesis, were tested. The relationship between the -3 and +4 sites in seven eukaryotic species does not support the translation initiation hypothesis. The amino acid usage at the position after the initiator (P1' position) compared to other positions in the coding sequences of seven eukaryotic species was examined. The result is consistent with the amino acid constraint hypothesis. In addition, this thesis explored the relationship between +4 nucleotide and translation efficiency in yeast. The result shows that +4 nucleotide is not important for translation efficiency, which does not support the translation initiation hypothesis. This work improves our current understanding of eukaryotic translation initiation process.
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Lesurf, Robert. "Molecular pathway analysis of mouse models for breast cancer." Thesis, McGill University, 2009. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=32499.
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Human breast cancer is an extremely heterogeneous disease, consisting of a number of different subtypes with varying levels of aggressiveness reflected by distinct, but largely undefined, molecular profiles. Here we have analyzed several novel mouse models for breast cancer in the context of the human subtypes, and have shown parallels between the mice and humans at numerous biologically relevant levels. In addition, we have developed a statistical framework to help elucidate the individual molecular components that are at play across a panel of human breast or murine mammary tumors. Our results indicate that, while no mouse model captures all aspects of the human disease, they each contain components that are shared by a subset of human breast tumors. Furthermore, our statistical framework provides numerous advantages over previous methodologies, in helping to reveal the individual molecular pathways that make up the biology of the tumors.<br>Le cancer du sein est connue pour être une maladie très hétérogène, composé d'un nombre de différents sous-types avec différents niveaux de l'agressivité et distinctes, mais indéfini, profils moléculaires. Ici, nous avons analysé plusieurs nouveaux modèles de souris pour le cancer du sein, dans le cadre des sous-types, et nous avons trouver des parallèles à un certain nombre de niveaux pertinents biologiques. En outre, nous avons développé une méthodologie statistique pour aider à élucider les différents composants moléculaires qui sont à jouer dans un groupe de tumours de sein d'humains ou mammaires murins. Nos résultats indiquent que, même si aucun modèle de souris capte tous les aspects de la maladie chez l'homme, chacun contiennent des composants qui sont partagées par un sous-ensemble de tumeurs mammaires humaines. En outre, notre outil statistique offre de nombreux avantages par rapport aux précédentes méthodes, pour aider à révéler les voies moléculaires qui composent la biologie des tumeurs.
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Lutimba, Stuart. "Determination of specificity and affinity of the Lactose permease (LacY) protein of Escherichia coli through application of molecular dynamics simulation." Thesis, Högskolan i Skövde, Institutionen för biovetenskap, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-15933.
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Proteins are essential in all living organisms. They are involved in various critical activities and are also structural components of cells and tissues. Lactose permease a membrane protein has become a prototype for the major facilitator super family and utilises an existing electrochemical proton gradient to shuttle galactoside sugars to the cell. Therefore it exists in two principle states exposing the internal binding site to either side of the membrane. From previous studies it has been suggested that protonation precedes substrate binding but it is still unclear why this has to occur in the event of substrate binding. Therefore this study aimed to bridge this gap and to determine the chemical characteristics of the transport pathway. Molecular dynamics simulation methods and specialised simulation hardware were employed to elucidate the dependency of substrate binding on the protonation nature of Lactose permease. Protein models that differed in their conformation as well as their protonation states were defined from their respective X-ray structures. Targeted molecular dynamics was implemented to drive the substrate to the binding site and umbrella sampling was used to define the free energy of the transport pathway. It was therefore suggested that protonation for sugar binding is due to the switch-like mechanism of Glu325 in the residue-residue interaction (His322 and Glu269) that leads to sugar binding only in the protonated state of LacY. Furthermore, the free energy profile of sugar transport path way was lower only in the protonated state which indicates stability of sugar binding in the protonated state.
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Chen, Huiling Zhou Huan Xiang Ferrone Frank A. "Prediction of protein structures and protein-protein interactions : a bioinformatics approach /." Philadelphia, Pa. : Drexel University, 2005. http://dspace.library.drexel.edu/handle/1860/481.
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Rho, Mina. "Probabilistic models in computational molecular biology applied to the identification of mobile genetic elements and gene finding." [Bloomington, Ind.] : Indiana University, 2009. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3386714.
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Thesis (Ph.D.)--Indiana University, School of Informatics and Computing, 2009.<br>Title from PDF t.p. (viewed on Jul 22, 2010). Source: Dissertation Abstracts International, Volume: 70-12, Section: B, page: 7299. Adviser: Haixu Tang.
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Ma, Siming. "Molecular Patterns and Signatures of Longevity." Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:33493444.
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Since their divergence from a common ancestor some 200 million years ago, mammals have undergone significant diversification in physiology, morphology, habitat, size, and longevity. The maximum lifespan of mammalian species ranges from under 3 to over 200 years, but the molecular basis of such variation is poorly understood. While many genes, pathways, dietary interventions, and pharmacological compounds have been shown to influence the lifespan of model organisms, it is not known whether the same mechanisms are responsible for the longevity variation across different species. This thesis presents the analyses of gene expression and the levels of metabolites, chemical elements, and/or proteins, across multiple organs and tissues of up to 42 species of mammals, as well as the analyses of 5 long-lived mouse models, 22 natural isolates of yeast, and 16 species of fruit flies, to identify the molecular patterns and signatures associated with species longevity. The results show that longer-lived mammals up-regulate ribosomal proteins and genes involved in DNA repair, and down-regulate ubiquitin-mediated proteolysis and apoptotic functions. Some of the metabolic changes in long-lived mammals, such as higher levels of sphingomyelins and glycerophospholipids but lower levels of polyunsaturated triacylglycerols, were also observed in long-lived mouse models. Yeast strains of varying replicative lifespan differed in their aerobic respiration capacity, attributable to different protein composition in mitochondria. Long-lived fruit flies overexpressed the genes involved in lipid metabolism but suppressed the genes involved in neuronal development. Many genes previously implicated in lifespan control in model organisms also showed the expected correlation with the longevity traits across species. This thesis presents the snapshots of the complex changes associated with species natural lifespan variation and offers new insights into the mechanisms of longevity control and potential lifespan extension strategies.<br>Medical Sciences
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Chaivorapol, Christina. "Systematic identification of regulatory pathways in human and mouse embryonic stem cells and other mammalian systems." Diss., Search in ProQuest Dissertations & Theses. UC Only, 2008. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3328095.
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Peterson, Mark Erik. "Evolutionary constraints on the structural similarity of proteins and applications to comparative protein structure modeling." Diss., Search in ProQuest Dissertations & Theses. UC Only, 2008. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3339202.
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Mendez, Gregory Scott. "Dinoflagellate genomic organization and phylogenetic marker discovery utilizing deep sequencing data." Thesis, University of Maryland, College Park, 2016. http://pqdtopen.proquest.com/#viewpdf?dispub=10159160.
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<p> Dinoflagellates possess large genomes in which most genes are present in many copies. This has made studies of their genomic organization and phylogenetics challenging. Recent advances in sequencing technology have made deep sequencing of dinoflagellate transcriptomes feasible. This dissertation investigates the genomic organization of dinoflagellates to better understand the challenges of assembling dinoflagellate transcriptomic and genomic data from short read sequencing methods, and develops new techniques that utilize deep sequencing data to identify orthologous genes across a diverse set of taxa. To better understand the genomic organization of dinoflagellates, a genomic cosmid clone of the tandemly repeated gene Alchohol Dehydrogenase (AHD) was sequenced and analyzed. The organization of this clone was found to be counter to prevailing hypotheses of genomic organization in dinoflagellates. Further, a new non-canonical splicing motif was described that could greatly improve the automated modeling and annotation of genomic data. A custom phylogenetic marker discovery pipeline, incorporating methods that leverage the statistical power of large data sets was written. A case study on Stramenopiles was undertaken to test the utility in resolving relationships between known groups as well as the phylogenetic affinity of seven unknown taxa. The pipeline generated a set of 373 genes useful as phylogenetic markers that successfully resolved relationships among the major groups of Stramenopiles, and placed all unknown taxa on the tree with strong bootstrap support. This pipeline was then used to discover 668 genes useful as phylogenetic markers in dinoflagellates. Phylogenetic analysis of 58 dinoflagellates, using this set of markers, produced a phylogeny with good support of all branches. The <i>Suessiales</i> were found to be sister to the <i>Peridinales.</i> The <i>Prorocentrales </i> formed a monophyletic group with the Dinophysiales that was sister to the <i>Gonyaulacales.</i> The <i>Gymnodinales</i> was found to be paraphyletic, forming three monophyletic groups. While this pipeline was used to find phylogenetic markers, it will likely also be useful for finding orthologs of interest for other purposes, for the discovery of horizontally transferred genes, and for the separation of sequences in metagenomic data sets.</p>
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Chew, Guo-Liang. "Non-Canonical Translation in Vertebrates." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17467487.
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Translation is a key process during gene expression: to produce proteins, ribosomes translate the coding sequences of mRNAs. However, vertebrate genomes contain more translation potential than these annotated coding sequences: translation has been detected in many non-coding RNAs and in the non-coding regions of mRNAs. To understand the role of such translation in vertebrates, I investigated: 1) the distribution of translation in vertebrate long non-coding RNAs, and 2) the effects of translation in the 5’ leaders of vertebrate mRNAs.
To quantify and localize translation in a genome-wide manner, we produced and analyzed ribosome profiling data in zebrafish, and analyzed ribosome profiling data produced by others. The nucleotide resolution afforded by ribosome profiling allows localization of translation to individual ORFs within a transcript, while its quantitative nature enables measurement of how much translation occurs within individual ORFs.
We combined ribosome profiling with a machine-learning approach to classify lncRNAs during zebrafish development and in mouse ES cells. We found that dozens of proposed lncRNAs are protein-coding contaminants and that many lncRNAs have ribosome profiles that resemble that of the 5’ leaders of coding mRNAs. These results clarify the annotation of lncRNAs and suggest a potential role for translation in lncRNA regulation.
Because much of the translation in non-coding regions of mRNAs occurs within uORFs, we further examined the effects of their translation on the cognate gene expression. While much is known about the repression of individual genes by their uORFs, how uORF repressiveness varies within a genome and what underlies this variation had not been characterized. To address these questions, we analyzed transcript sequences and ribosome profiling data from human, mouse and zebrafish.
Linear modeling revealed that sequence features at both uORFs and coding sequences contribute similarly and substantially toward modulating uORF repressiveness and coding sequence translational efficiency. Strikingly, uORF sequence features are conserved in mammals, and mediate the conservation of uORF repressiveness in vertebrates. uORFs are depleted near coding sequences and have initiation contexts that diminish their translation. These observations suggest that the prevalence of vertebrate uORFs may be explained by their functional conservation as weak repressors of coding sequence translation.<br>Biology, Molecular and Cellular
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Rahrig, Ryan Robert. "Automated Alignment of RNA 3D Structures." Bowling Green State University / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1276873588.
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Patel, Vishal N. "Colon Cancer and its Molecular Subsystems: Network Approaches to Dissecting Driver Gene Biology." Case Western Reserve University School of Graduate Studies / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=case1310087563.
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Pennington, Steven. "Pulsed induction, a method to identify genetic regulators of determination events." Thesis, Oklahoma State University, 2015. http://pqdtopen.proquest.com/#viewpdf?dispub=3727701.
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<p> Abstract: Determination is the process in which a stem cell commits to differentiation. The process of how a cell goes through determination is not well understood. Determination is important for proper regulation of cell turn-over in tissue and maintaining the adult stem cell population. Deregulation of determination or differentiation can lead to diseases such as several forms of cancer. In this study I will be using microarrays to identify candidate genes involved in determination by pulse induction of mouse erythroleukemia (MEL) cells with DMSO and looking at gene expression changes as the cells go through the early stages of erythropoiesis. The pulsed induction method I have developed to identify candidate genes is to induce cells for a short time (30 min, 2 hours, etc.) and allow them then to grow for the duration of their differentiation time (8 days). For reference, cells were also harvested at the time when the inducer is removed from the media. Results show high numbers of genes differentially expressed including erythropoiesis specific genes such as GATA1, globin genes and many novel candidate genes that have also been indicated as playing a role in the dynamic early signaling of erythropoiesis. In addition, several genes showed a pendulum effect when allowed to recover, making these interesting candidate genes for maintaining self-renewal of the adult stem cell population.</p>
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Hernandez-Garcia, Carlos M. "ISOLATION AND CHARACTERIZATION OF SOYBEAN PROMOTERS." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1366303238.
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Mudenda, Lwiindi. "Identification of Dermacentor andersoni saliva proteins that modulate mammalian phagocyte function." Thesis, Washington State University, 2015. http://pqdtopen.proquest.com/#viewpdf?dispub=3717421.
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<p> Ticks are obligate blood sucking parasites which transmit a wide range of pathogens worldwide including protozoa, bacteria and viruses. Additionally, tick feeding alone may result in anemia, dermatosis and toxin-induced paralysis. <i> Dermacentor andersoni</i> is a species of tick found in the western United States that transmits pathogens of public health importance including <i> Rickettsia rickettsii, Francisella tularensis,</i> and Colorado Tick Fever Virus, as well as <i>Anaplasma marginale</i>, a rickettsial pathogen that causes economic losses in both the dairy and beef industries worldwide. <i>D. andersoni</i> ticks are obligate blood sucking parasites that require a blood meal through all stages of their lifecycle. During feeding, ticks secrete factors that modulate both innate and acquired immune responses in the host which enables them to feed for several days without detection. The pathogens transmitted by ticks exploit these immunomodulatory properties to facilitate invasion of and replication in the host. Molecular characterization of these immunomodulatory proteins secreted in tick saliva offers an opportunity to develop novel anti-tick vaccines as well as anti-inflammatory drug targets. To this end we performed deep sequence analysis on unfed ticks and ticks fed for 2 or 5 days. The pooled data generated a database of 21,797 consensus sequences. Salivary gland gene expression levels of unfed ticks were compared to 2- and 5-day fed ticks to identify genes upregulated early during tick feeding. Next we performed mass spectrometry on saliva from 2- and 5-day fed ticks and used the database to identify 677 proteins. We cross referenced the protein data with the transcriptome data to identify 157 proteins of interest for immunomodulation and blood feeding. Both proteins of unknown function and known immunomodulators were identified. We expressed four of these proteins and tested them for inhibition of macrophage activation and/or cytokine expression in vitro. The results showed diverse effects of the various test proteins on the inflammatory response of mouse macrophage cell lines. The proteins upregulated some cytokines while downregulating others. However, all the proteins upregulated the regulatory cytokine IL-10.</p>
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Dybas, Joseph M. "The evolution of protein folds from the perspective of structure motifs." Thesis, Yeshiva University, 2015. http://pqdtopen.proquest.com/#viewpdf?dispub=3664550.
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<p> Understanding how protein structures evolve is essential for deciphering relationships between homologous proteins, which can inform structure classification and function annotation and aid in protein modeling and design methods. The observation that structure is more conserved than sequence, over the course of evolution, implies a model of evolution where sequences diverge within a discrete set of well-defined folds, which suggests that homology does not exist across fold definitions. However, as more structures have been experimentally solved and the coverage of the universe of folds has increased, the original view of a discrete fold space has been revised to include a more nuanced view of a continuous space defined by regions in which structural similarities can connect globally disparate topologies.</p><p> Structural, functional and evolutionary relationships are known to, in some cases, span fold definitions. A hallmark of relationships connecting disparate topologies is the conservation of local structure motifs within globally different folds. In order to systematically identify and analyze these relationships, a new approach to structure comparisons and structure classification is required. The goal of this work is to systematically identify evolutionary relationships between folds and to generate a classification of the fold universe that can accurately represent even the relationships that exist across disparate topologies. </p><p> An exhaustive library of supersecondary-structure motifs (Smotif), defined as two secondary structures connected by a loop, is established and characterized. A novel Smotif-based, superposition-independent structure comparison method (SmotifCOMP) is developed that quantitatively measures the Smotif-based similarity of compared structures in order to identify evolutionary relationships. SmotifCOMP is able to provide a quantitative and robust measure of similarity between disparate topologies since it does not rely on a global superposition. The comparison method is used to perform a systematic analysis of the SCOP Superfamilies and generate a non-hierarchical, network-based representation of the fold universe.</p><p> This thesis describes the development of a novel method of comparing structures and an improved representation of the relationships within the fold space. This work provides insight into the existence of evolutionary relationships between folds and strengthens the view of a connected and continuous fold universe.</p>
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Hart, James Clinton. "Genetic and Genomic Bases of Evolved Increases in Stickleback Dentition." Thesis, University of California, Berkeley, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10817049.
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<p> Evolution—the great tinkerer—has produced the astounding diversity of form within and between existing species. It is a fundamental goal of evolutionary biology to understand the origin of such diversity. What types of genes underlie evolved changes in morphology? Are certain types of mutations (notably changes within regulatory regions) more likely to be used to produce adaptive changes in form? When distinct populations evolve similar morphological changes, are the underlying genetic bases changes to the same genes, the same genetic pathways, or largely independent? Are changes in form modular, or are their concerted changes to multiple developmentally similar organs? The ever cheapening cost of sequencing, coupled the availability of high-quality reference genomes, allows high-throughput approaches to identifying the loci of evolution. The emergence of a robust genome engineering system, CRISPR/Cas9, allows for efficient and direct testing of a gene's phenotype. Combining both of these techniques with a model system with naturally evolved phenotypic variation, the threespine stickleback, allows for systems-level answers to the many evolutionary questions. </p><p> Chapter one outlines the field of evolutionary developmental biology. It proposes two alternative viewpoints for thinking about the evolution of form. The first is the view of the `Modern Synthesis', linking Mendelian inheritance with Darwinian natural selection, which explains evolution as the change in allele frequencies over time. The second views evolution through the lens of deep homology, focusing on changes to developmental programs over time, even across related organs within the same animal. It then introduces key concepts within evolutionary and developmental biology, including <i> cis</i>-regulation of gene expression, and gene regulatory networks. It then provides examples of evolution reusing similar gene regulatory networks, including <i>Hox</i> genes, <i>Pax6</i> dependent eye initiation, and ectodermal placode development. Teeth use highly conserved signaling pathways, during both their initiation and replacement. Threespine sticklebacks <i>Gasterosteus aculeatus</i> have repeatedly adapted following a shift from marine to freshwater environments, with many independently derived populations sharing common morphological traits, including a gain in tooth number. The following chapters investigate this gain in tooth number in multiple distinct populations of sticklebacks. </p><p> Chapter two describes the discovery and mapping of a spontaneous stickleback albino mutation, named <i>casper</i>. <i>casper</i> is a sex-linked recessive mutation that results in oculocutaneous albinism, defective swim bladders, and blood clotting defects. Bulked segregant mapping of <i> casper</i> mutants revealed a strong genetic signal on chromosome 19, the stickleback X chromosome, proximal to the gene <i>Hps5</i>. <i> casper</i> mutants had a unique insertion of a G in the 6<sup>th</sup> exon on <i>Hps5</i>. As mutants in the human orthologue of <i> Hps5</i> resulted in similar albino and blood clotting phenotypes, <i> Hps5</i> is a strong candidate underlying the <i>casper</i> phenotype. Further supporting this model, genome editing of <i>Hps5</i> phenocopied <i>casper</i>. Lastly, we show that <i>casper</i> is an excellent tool for visualizing the activity of fluorescent transgenes at late developmental stages due to the near-translucent nature of the mutant animals. </p><p> Chapter three details the fine mapping of a quantitative trail locus (QTL) on chromosome 21 controlling increases in tooth number in a Canadian freshwater stickleback population. Recombinant mapping reduced the QTL-containing region to an 884kb window. Repeated QTL mapping experiments showed the presence of this QTL on multiple, but not all, wild derived chromosomes from the Canadian population. Comparative genome sequencing revealed the perfect correlation with genetic data of ten variants, spanning 4.4kb, all within the 4<i> th</i> intron of the gene <i>Bmp6</i>. Transgenic analysis of this intronic region uncovered its role as a robust tooth enhancer. TALEN induced mutations in <i>Bmp6</i> revealed required roles for the gene in stickleback tooth development. Finally, comparative RNA-seq between <i> Bmp6</i> wild-type and mutant dental tissue showed a loss of mouse hair stem cell genes in <i>Bmp6</i> mutant fish teeth, suggesting deep homology of the regeneration of these two organs. </p><p> Chapter four investigates the evolved changes in gene expression that accompany evolved increases in tooth number in two distinct freshwater populations. Independently derived stickleback populations from California and Canada have both evolved increases in tooth number, and previous work suggested that these populations used distinct genetic changes during their shared morphological changes. RNA-seq analysis of dental tissue from both freshwater populations compared to marine revealed a gain in critical regulators of tooth development in both freshwater populations. These evolved changes in gene expression can be partitioned in <i>cis</i> changes (mutations within regulatory elements of a gene) and trans changes (changes to the overall regulatory environment) using phased RNA-seq data from marine-freshwater F1 hybrids. Many genes show evidence for stabilizing selection of expression levels, with <i>cis </i> and <i>trans</i> changes in opposing directions (Abstract shortened by ProQuest.). </p><p>
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Rhodes, Christopher. "Epigenetic Repression in the Context of Adult Neurogenesis." Thesis, The University of Texas at San Antonio, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10686193.
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<p> Neural stem progenitor cells (NSPCs) in the mammalian brain contribute to life-long neurogenesis and brain health. Adult mammalian neurogenesis primarily occurs in the subventricular zone (SVZ) and the subgranular zone (SGZ) of the dentate gyrus. Epigenetic repression is a crucial regulator of cell fate specification during adult neurogenesis. How epigenetic repression impacts adult neurogenesis and how epigenetic dysregulation may impact neoplasia or tumorigenesis remains poorly understood. Examination of epigenetic regulation in the adult mammalian brain is complicated by the heterogeneous nature of neurogenic niches and by the highly orchestrated fate specification processes within neural stem progenitor cells involving myriad intrinsic and extrinsic factors. To overcome these challenges, we utilized a cross-species approach. To model histone modifications as they exist <i>in vivo</i> for epigenetic profiling, we isolated neural stem progenitor cells from the adult SVZ and SGZ of non-human primate baboon brains. To determine cellular and molecular changes within the adult SVZ and SGZ following loss of epigenetic repression, we utilized multiple mouse models, including conditional <i> Ezh2</i> and <i>Suv4-20h1</i> knockouts. To model the non-cell type specific effects common to small molecule screening and brain chemotherapeutic agents, induction of conditional knockout utilized a recombinant Cre protein. Finally, to model epigenetic mechanisms during SVZ-associated glioblastoma (GBM) tumorigenesis, we conducted comparative analysis between healthy NSPCs and GBM specimens from humans. The convergence of baboon, mouse and human models of adult neurogenesis revealed that epigenetic repression is a critical mechanism regulating proper neural cell fate and that epigenetic dysregulation may be a driver of GBM.</p><p>
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Söderquist, Fredrik. "Proteus : A new predictor for protean segments." Thesis, Linköpings universitet, Teknisk biologi, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-121260.
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The discovery of intrinsically disordered proteins has led to a paradigm shift in protein science. Many disordered proteins have regions that can transform from a disordered state to an ordered. Those regions are called protean segments. Many intrinsically disordered proteins are involved in diseases, including Alzheimer's disease, Parkinson's disease and Down's syndrome, which makes them prime targets for medical research. As protean segments often are the functional part of the proteins, it is of great importance to identify those regions. This report presents Proteus, a new predictor for protean segments. The predictor uses Random Forest (a decision tree ensemble classifier) and is trained on features derived from amino acid sequence and conservation data. Proteus compares favourably to state of the art predictors and performs better than the competition on all four metrics: precision, recall, F1 and MCC. The report also looks at the differences between protean and non-protean regions and how they differ between the two datasets that were used to train the predictor.
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Schiller, Benjamin J. "Data Biology| A quantitative exploration of gene regulation and underlying mechanisms." Thesis, University of California, San Francisco, 2013. http://pqdtopen.proquest.com/#viewpdf?dispub=3587899.
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<p> Regulation of gene expression is a fundamental biological process required to adapt the full set of hereditary information (i.e., the genome) to the varied environments that any organism encounters. Here, we elucidate two distinct forms of gene regulation – of endogenous genes by binding of transcription factors to information-containing genomic sequences and of selfish genes (“transposons”) by targeting of small RNAs to repetitive genomic sequences – using a wide array of approaches. </p><p> To study regulation by transcription factors, we used glucocorticoid receptor (GR), a hormone-activated, DNA-binding protein that controls inflammation, metabolism, stress responses and other physiological processes. <i>In vitro</i>, GR binds as an inverted dimer to two imperfectly palindromic “half sites” separated by a “spacer”. Moreover, GR binds different sequences with distinct conformations, as demonstrated by nuclear magnetic resonance spectroscopy (NMR) and other biophysical methods. </p><p> <i>In vivo</i>, GR employs different functional surfaces when regulating different genes. We investigated whether sequences bound by GR <i> in vivo</i> might be a composite of several motifs, each biased toward utilization of a particular pattern of functional surfaces of GR. Using microarrays and deep sequencing, we characterized gene expression and genomic occupancy by GR, with and without glucocorticoid treatment, of cells expressing GR alleles bearing differences in three known functional surfaces. We found a “sub-motif”, the GR “half site”, that relates to utilization of the dimerization interface and directs genomic binding by GR in a distinct conformation. </p><p> To study repression of tranposons, we characterized the production and function of small RNAs in the yeast <i>Cryptococcus neoformans</i>. We found that target transcripts are distinguished by suboptimal introns and inefficient splicing. We identified a complex, SCANR, required for synthesis of small RNAs and demonstrate that it physically associates with the spliceosome. We propose that recognition of gene products by SCANR is in kinetic competition with splicing, thereby further promoting small RNA production from target transcripts. </p><p> To achieve these results, we developed new bioinformatics tools: twobitreader, a small Python package for efficient extraction of genomic sequences; scripter, a flexible back-end for easily creating scripts and pipeline; and seriesoftubes, a pipeline built upon scripter for the analysis of deep sequencing data. </p>
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Godec, Jernej. "Molecular Mechanisms of CD8+ T Cell Differentiation." Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:33493424.
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CD8+ T cells are a crucial component of the adaptive immune system and are required for optimal protection from most pathogens and cancer. They function by secreting pro-inflammatory cytokines and by directly eliminating infected and malignant cells. In order to be effective, CD8+ T cells must be activated through a complex sequence of steps involving engagement of the antigen-specific T cell receptor (TCR) and other receptors, which orchestrate transcriptional, epigenetic, and metabolic changes that direct the differentiation of the responding cells. Following optimal activation, naive CD8+ T cells rapidly undergo clonal expansion and effector differentiation that enables prompt resolution of infection. Following pathogen clearance, a fraction of effector CD8+ T cells differentiate into long-lived memory CD8+ T cells that provide robust protection from re-challenge with the same microbe. However, in the context of persistent abundance of antigen and inflammation, such as in chronic infections and in cancer, the T cells instead become gradually more dysfunctional – a state known as T cell exhaustion.
The overarching goal of this thesis is to identify the cardinal features and molecular mechanisms associated with three main states in which CD8+ T cells exist: T cell memory, T cell exhaustion, and T cell effector differentiation. I used two complementary approaches to examine CD8+ T cells at the different states in vivo. First, I used classical immunology techniques including knockout mice and cellular phenotypic analyses to examine the role of cell surface molecules PD-1 and CD39 on CD8+ T cells in the context of memory and exhaustion, respectively. Secondly, I developed a novel experimental platform that enables gene perturbation in naive CD8+ T cells in vivo during their differentiation. I used this approach to systematically interrogate the transcriptional programming of activated CD8+ T cells and to identify novel regulators of effector differentiation. In a proof of concept study, I used this system to further define how the transcription factor BATF regulates CD8+ T cell activation. Additionally, I used this experimental platform to systematically interrogate the functional role of a set of ~80 transcription factors in CD8+ T cell differentiation, and identified TGIF1 as a novel regulator of this process.
The role of the co-inhibitory receptor PD-1 on CD8+ T was examined in mice using an acute respiratory infection model. PD-1 is a co-inhibitory receptor that is up-regulated on T cells following activation and recruits SHP1/2 phosphatases to directly antagonize signals through the TCR and this way inhibit the activation of T cells. It is down-regulated following the resolution of an acute infection but remains persistently expressed on CD8+ T cells in chronic infections and cancer. As such, PD-1 has been exhaustively studied for its contribution to the functional exhaustion of T cells. However, its role in acute infections remains less defined. We found that this receptor prevents over-activation and over-expansion of CD8+ T cells following initial differentiation, and is crucial for optimal differentiation of effector CD8+ T cells into functional memory cells.
Exhausted CD8+ T cells express several markers distinctive of the state. Some, like PD-1, Tim-3, and Lag-3 are well known. However, genome-wide transcriptional studies identified numerous additional genes that are differentially expressed in the exhausted state. Thus, we hypothesized that additional markers may provide characteristic features of the exhausted cell state and may function in chronic infections. We investigated one such gene – ENTPD1 – that encodes for CD39. This cell surface molecule is an ectonucleotidase that hydrolyzes extracellular ATP into ADP and AMP, which can be further broken down to immunosuppressive adenosine by CD73. In the context of the immune system, CD39 has largely been studied on CD4+ regulatory T cells, which use CD39 as a mechanism to suppress immune responses. However, surprisingly, we found that CD8+ T cells can also express CD39, but its expression is largely restricted to terminally exhausted CD8+ T cells. These cells are most dysfunctional as measured by the most limited proliferative capacity and ability to produce pro-inflammatory cytokines. We have observed this biology in both human and mouse chronic viral infections. Additional studies further demonstrated the importance of CD39 and the purinergic pathway in regulating lethal immunopathology associated with chronic LCMV infection in mice.
In addition to interrogating memory and exhaustion fates of CD8+ T cells, we also examined the initial regulatory programs involved in CD8+ T cell differentiation in vivo through gene silencing. Gene perturbation in naive T cells without prior cellular stimulation has been a continuous challenge in the field. To circumvent this limitation, we engineered a novel experimental platform that enables inducible gene knock-down in any immune cell in mice in vivo without prior manipulation of these cells. Initially, I validated this system by knocking down BATF and confirmed its essential role in CD8+ T cell responses to acute LCMV infection. Additionally, leveraging the inducible nature of the platform, I showed that BATF functions in the early stages of T cell activation but becomes dispensable once its transcriptional program is initiated.
Several other transcription factors such as T-bet, Eomes, Bcl6, and Blimp-1 have been described to regulate CD8+ T cell differentiation. However, numerous additional transcription factors may function in this process based on their rapid up-regulation following T cell activation. I used the novel platform to systematically test the functional relevance of ~80 additional transcription factors in a pooled setting. These experiments identified several novel candidate regulators of this process. We validated one such gene – Tgif1 – to confirm its role in the effector CD8+ T cell differentiation following acute LCMV infection and provide clues to the potential mechanism in which it may function.
The above projects have benefited significantly from genome-wide transcriptional datasets of cells at various states or of different genotypes that we generated or that originate from published studies. One particularly powerful approach to examine differences between different groups is gene set enrichment analysis (GSEA) that examines coordinate up- or down-regulation of sets of genes rather than assessing differential expression of specific genes. This is particularly important because changes in biological processes are often guided by relative small changes of groups of genes that act in concert rather than by a robust expression change of a single gene. This approach, however, is only informative if a relevant gene-set collection is used to analyze the data. Existing collections are largely centered around cancer biology and general biological processes but no extensive gene-set collection existed that contained information describing immune processes. Thus, we created ImmuneSigDB – the largest collection of immunology-focused gene sets to date by identifying, annotating, and reanalyzing ~400 published immunology studies. To show its broad use, we used it to examine the cross-species conservation of transcriptional responses in the immune system. We focused on analyzing transcriptional data from systemic responses to sepsis using GSEA and a novel approach, called leading edge metagene analysis. Using these approaches, we uncovered shared and species-specific biology in mouse and human transcriptional responses to sepsis.
Deciphering CD8+ T cell biology is key for conceptualizing new medical interventions that may boost their activation, memory development, and rejuvenation from functional exhaustion. We have determined that PD-1 is essential for optimal CD8+ T cell memory responses, and that BATF is a key transcription factor initiating effector T cell transcriptional programming. We also identified CD39 as a new marker of terminally exhausted CD8+ T cells and uncovered a key role for purinergic signaling in regulating lethal immunopathology in LCMV Clone 13 infection in mice. Furthermore, we developed a new experimental platform that enables systematic interrogation of gene function in any hematopoietic cell type by inducible knock-down of genes and identified TGIF1 as a novel negative regulator of CD8+ T cell responses. We have also developed a new computational resource to improve analyses of transcriptional profiles in the immune system. Together, the body of work presented in this thesis advances our knowledge of major states of CD8+ T cell biology, uncovering both novel mechanisms underlying CD8+ T cell function, as well as highlighting potential novel therapeutic targets that may be transformative in creating better vaccines, treating infections, or fighting cancer.<br>Medical Sciences
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Rhodes, Christopher. "Molecular targets of chromatin marks H3K4me3, H3K9me3 and H3K27me3 in an adult germinal niche." Thesis, The University of Texas at San Antonio, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=1556578.
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<p> Neural stem cells (NSCs) participate in a delicate balance between maintaining cellular identity through self-renewal and differentiating into myriad neural cell types. Understanding exactly how epigenetic mechanisms regulate this balance and the subsequent differentiation process in adult mammalian brain is an ongoing effort. We conducted a genome wide association study to elucidate the roles of genes in neural progenitors regulated by chromatin modifications. Neural progenitors of baboon SVZ were examined using ChIP-Seq (chromatin immuneprecipitation followed by deep sequencing) to determine genome wide gene targets of three histone modifications: H3K4me3, H3K9me3 and H3K27me3. Our data suggest these chromatin marks are associated with genes responsible for cellular organization and morphology, proliferation and survival, neuron development. Taken together these processes suggest histone modifications, predominantly H3K27me3, are responsible for maintenance of NSC identity. Our findings also highlight the importance of using in vivo models to study the SVZ neurogenic niche and compel examination of the H3K27me3 catalyzing enzyme EZH2. In the future, the role of EZH2 will be determined by EZH2 conditional knockout and overexpression models, using stereotaxic injections of novel Cre protein and lentiviral delivery of EZH2, respectively.</p>
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Pasalic, Zlatana. "Evaluation of search models for Molecular Replacement using MolRep." Thesis, University of Skövde, Department of Computer Science, 2002. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-709.
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<p>he aim of this study is to use several homology models of different completeness and accuracy and to evaluate them as search models for Molecular Replacement (MR).Three structural groups are evaluated: α-, β- and α/β- group. From every group one template structure and a couple of search models are selected. The search models are manipulated and evaluated. B-factor manipulation, side chain removal and homology modelling are the ways the search models are manipulated. This work shows that B-factor manipulation do not improve the search models. The work also shows that removing the side chains is not improving the search models. Finally the work shows that homology modelling did not model better search models.</p>
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Mueller, William F. "Pre-mRNA Architecture and Sequence Element Regulation of Alternative Splicing." Thesis, University of California, Irvine, 2013. http://pqdtopen.proquest.com/#viewpdf?dispub=3557196.
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<p> Human genes are split into regions that code for protein, exons, and regions that don't, introns. Upon transcription, the removal of these intervening introns is necessary if a usable mRNA molecule is to be translated. The process of intron removal and subsequent ligation of exons is called splicing and is carried out by a large complex called the spliceosome. This process is driven by sequence elements within the pre-mRNA itself and is the major contributor of diversity to the human transcriptome. Due to the ubiquitous nature of alternative splicing in almost every multi-exon gene, the regulation pathways of exon inclusion are a subject of wide study. </p><p> The different lengths of introns and exons as well as location of splice sites in a pre-mRNA molecule have been shown to have differing affects on the spliceosomes ability to recognize them. Using <i>in vitro</i> splicing and complex formation assays in parallel with cell transfection experiments, we determined that the distance between two splice sites across the intron or across the exon are strong predictors of splice site usage. Additionally, we found that two splice sites interact differently when placed at different lengths apart. Our findings suggest a mechanism for observed selection of specific intron/exon architectures. </p><p> Splice site recognition is also influenced by the presence of protein binding sequence elements in the pre-mRNA that alter spliceosomal recruitment. Previously, these proteins and sequence elements had been rigidly classified into splice enhancing or inhibiting categories. We show that this rigid classification is incorrect. We found that the location of these elements relative to the splice site determines their enhancing or silencing effect. That is, an enhancing element found upstream of a splice site imposes a silencing effect when relocated downstream of the splice site (and vice versa). </p><p> Spliceosomal proteins are conserved from yeast to humans. The sequence elements used in pre-mRNA sequences have been evolving over time but under pressure from multiple cellular processes, including splicing. To observe the effect of splicing on evolution, we took advantage of the synonymous mutation positions that are under the least amount of selective pressure from the genetic code. We mutated these positions and found that some caused a large decrease in exon inclusion. When we analyzed the comparative alignment data, we found that these specific nucleotide mutations were selected against across species in order to maintain exon inclusion. SNP analysis showed that this pattern of selection was broadly observable at synonymous positions throughout the human genome.</p>
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Armoskus, Christopher. "Identification and characterization of sexually dimorphic genes in the developing mouse cortex and hippocampus." Thesis, California State University, Long Beach, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=1527357.
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<p> In both mice and humans, males and females exhibit differences in behavior and response to neurological and psychological diseases that are linked to the cortex and hippocampus. The perinatal exposure of males to testosterone secreted by the testes creates alterations in neural structures and behaviors that can persist throughout their lives; however, the molecular mechanisms that underlie the actions of sex steroids to produce these lasting changes are still unclear. Given that regulation of gene expression is a primary mechanism whereby sex steroids exert changes to an organism, I sought to identify genes expressed at different levels between the sexes in the cortex and hippocampus and to determine the effect of testosterone on expression of these genes. Using gene expression microarrays and RT-qPCR, I identified genes that are differentially expressed between the sexes in the neonatal mouse cortex and hippocampus; however, whether perinatal testosterone is regulating these differences remains unclear.</p>
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Cocca, Stephanie M. "Phylogenetic analysis, modeling and experimental studies of the Saccharomyces cerevisiae palmitoylated protein kinase gene, ENV7." Thesis, California State University, Long Beach, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=1527907.
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<p> Env7 is a vacuole membrane-localized protein kinase that is orthologous to the human serine/threonine protein kinase, STK16. It is evolutionarily well-conserved throughout Eukarya, and it has one ortholog in Bacteria. Phylogenetic analyses of sequences homologous to Env7 revealed clades that are inconsistent with established eukaryotic phylogeny, suggesting that both horizontal and vertical gene transmission are responsible for their conservation. Conserved amino acid residues and motifs that are potentially important to Env7's catalytic activity, localization, and interactions with other proteins were also identified and assessed. Additionally, one such conserved motif—the glycine-rich loop—was mutated in an effort to affect ATP binding in Env7. The phenotype resulting from this mutation was a slightly increased number of mutant cells exhibiting multi-lobed vacuoles under normal conditions.</p>
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Venkatasubrahmanyam, Shivkumar. "Regulation of eukaryotic transcription by modification of chromatin structure." Diss., Search in ProQuest Dissertations & Theses. UC Only, 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3261249.
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Heimiller, Joseph Karl. "Genome-wide analysis of splicing requirements and function through mRNA profiling." Thesis, University of Colorado at Boulder, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=3607314.
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<p> The RNA-binding proteins U2AF and PTB play important roles in gene expression in many eukaryotic species. Although U2AF and PTB have been well-studied, their functional requirements have not been investigated on a genome-wide scale. In this thesis, I analyze RNA expression data to determine the requirement of the general splicing factor U2AF in <i>S. pombe</i> and to identify genes misregulated in Drosophila PTB mutants. I find that many introns are insensitive to U2AF inactivation in a <i>Schizosaccharomyces pombe</i> U2AF59 mutant, <i>prp2.1.</i> Bioinformatics analysis indicates that U2AF-insensitive introns have stronger 5' splice sites and higher A/U composition. The importance of intronic nucleotide composition was further investigated using wild type RNA expression data sets. I show that nucleotide composition is a relatively important factor for regulated intron retention in a variety of species. I also analyzed the RNA-binding protein PTB using RNA Seq data to reveal genes misregulated in PTB mutants in <i>D. melanogaster.</i> I identify misregulation of alternative splicing in PTB mutants and putative PTB binding sites. In the PTB embryonic lethal mutant, which shows dorsoventral patterning defects, I show that dorsal fate genes are significantly up-regulated. I present a model to link PTB to dorsal closure defects. This thesis provides the first genome-wide analysis of U2AF in <i>S. pombe</i> and PTB in <i>Drosophila melanogaster. </i></p>
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Lee, Ryan. "Mechanisms of Oncogenesis Driven by SNF5-Loss in Pediatric Rhabdoid Tumors." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17467199.
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The characterization of inactivating mutations affecting SNF5 in pediatric rhabdoid tumors constituted the first clear connection between an epigenetic regulator and tumor formation. SNF5 is a core subunit of the evolutionarily conserved SWI/SNF chromatin remodeling complex, a complex that has recently emerged as being frequently mutated across a wide spectrum of cancers. However, the exact mechanism by which SNF5 loss causes rhabdoid tumor, and SWI/SNF mutations are involved cancers generally, remains undetermined. Since SWI/SNF is a chromatin remodeling complex, it is unclear whether the perturbation of chromatin structure and epigenetic dysregulation caused by SNF5 loss alone may drive cancer formation or if SNF5 mutations are dependent on additional cooperative somatic mutations.
In order to determine what, if any, additional pathways cooperate with SNF5 loss, we have sequenced the exomes of 35 primary rhabdoid tumors. Despite the lethal nature of these cancers, we identified remarkably few coding mutations, with SNF5 loss being the only significantly recurring event. The mutation rate of rhabdoid tumors is among the lowest of all cancer genomes sequenced to date. Our results demonstrate that loss of SNF5 alone appears to a genetic event sufficient to drive rhabdoid tumor and that genomic instability and high mutation rates are not required for oncogenesis.
Because SNF5 loss is the only known genetic driver of these cancers, RT represents a good model system in which to study the effects of SWI/SNF subunit mutations on driving oncogenesis through disruption of transcriptional regulation. As one means of evaluating such disruptions, we performed chromatin immunoprecipitation-sequencing of histone modifications in primary RT tissues and RT cell lines. Despite being genetically indistinguishable, RT group according to tissue of origin when characterized by H3K27ac at active enhancers. Additionally, we found that SNF5 loss impairs SWI/SNF binding to enhancers, but its loss has minimal effect on the targeting of SWI/SNF to super-enhancers. Our data suggest a model whereby SNF5 loss blocks acquisition of enhancers required for differentiation but leaves intact super-enhancer structures that underlie the proliferative fate of the progenitor cells of origin. These studies collectively characterize an epigenetic mechanism underlying tumor formation upon SNF5 loss.<br>Medical Sciences
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Guo, Xiangxue. "Biochemical and Bioinformatics Analysis of CVAB C-Terminal Domain." Digital Archive @ GSU, 2006. http://digitalarchive.gsu.edu/biology_diss/3.
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Cytoplasmic membrane proteins CvaB and CvaA and the outer membrane protein TolC form the bacteriocin colicin V (ColV) secretion system in Escherichia coli. CvaB functions as an ATP-binding cassette transporter with nucleotide-binding motifs in the C-terminal domain (CTD). To study the role of CvaB-CTD in the ColV secretion, a truncated construct of this domain was made and over-expressed. Different forms of CvaB-CTD were obtained during purification, and were identified as monomer, dimer, and oligomer on gel filtration. Nucleotide binding was shown critical for the CvaB-CTD dimerization: oligomers could be converted into dimers by nucleotide bindings; the removal of nucleotide from dimers resulted in transient monomers followed by CTD oligomerization and aggregation; no dimer form could be cross-linked from the nucleotide-binding deficient mutant D654H. The spatial proximity of the Walker A site and ABC signature motif in CTD dimer was identified through disulfide cross-linking of mixed CvaB-CTD with mutants A530C and L630C, while mutations did not dimerize individually. Those results indicated that the CvaB-CTD formed a nucleotide-dependent head-to-tail dimer. Molecular basis of differential nucleotide bindings was also studied through bioinformatics prediction and biochemical verification. Through sequence alignment and homology modeling with bound ATP or GTP, it was found that the Ser503 and Gln504 on aromatic stacking region (Y501DSQ-loop) of CvaB-CTD provided two additional hydrogen-bonds to GTP, but not to ATP. Site-directed mutations of the S503A and/or Q504L were designed based on the model. While site-directed mutagenesis studies of Walker A&B sites or the ABC signature motif affected little on the GTP-binding preference, the double mutation (S503A/Q504L) on the Y501DSQ-loop increased both ATP-binding and ATPase activity at low temperatures. The double mutant showed slight decrease of GTP-binding and about 10-fold increase of the ATP/GTP-binding ratio. Similar temperature sensitivity in nucleotide-binding and activity assays were identified in the double mutant at the same time. Mutations on the Y501DSQ-loop did not affect the ColV secretion level in vivo. Together, the Y501DSQ-loop is structurally involved in the differential binding of GTP over ATP.
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Savelkoul, Elizabeth Jennings. "Molecular evolution of meiosis genes in fungi." Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/6635.
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Meiosis as a general process is prevalent across the eukaryotes, as are the orthologs of many genes encoding proteins known to function in meiosis. However, many organisms have experienced derived losses of otherwise well-conserved meiosis genes without losing meiosis and sexual reproduction. Although this general conservation of meiosis genes and precedent for derived meiosis gene losses has been previously established, questions remain about the frequency of and evolutionary forces contributing to these trends. This work sought (i) to characterize the phylogenetic distribution of 15 meiosis genes (most of which are known to function only in meiosis) in the exemplar eukaryotic kingdom Fungi and (ii) to use this dataset to investigate evolutionary processes contributing to the loss and retention of these genes.
Orthologs of 15 meiosis genes (Rad51, Rad21, Spo11, Rec8, Dmc1, Hop2, Mnd1, Sae3/Swi5, Mei5/Sfr1, Pch2, Hop1, Msh4, Msh5, Mer3, Zip3) were identified by BLAST-based techniques and phylogenetically validated in most of the 109 publicly available sequenced fungal genomes investigated, but numerous putative derived losses were also detected. Rad51, Rad21, Rec8, and Spo11 were nearly universally conserved; the remaining genes were each undetectable or independently pseudogenized multiple times within fungi, particularly often for Pch2. Genes with previously known functional interactions tended to show parallel presence, absence, or pseudogenization patterns. Although this work primarily established the conserved presence of meiosis gene orthologs at the DNA level, examination of expressed sequence tags (ESTs) showed that many species--including some not previously known to undergo sexual reproduction--were competent to transcribe (and often splice) mRNA from the identified meiosis genes.
Factors potentially influencing derived meiosis gene losses were investigated in two ways. First, degenerate PCR was used to amplify loci expected to contain orthologs of Msh4, Msh5, Pch2, and Zip3 in various Aspergillus species closely related to Aspergillus nidulans (a species with undetected or pseudogenized orthologs of these four genes.) The loss of Pch2 substantially predated the pseudogenization of Msh4, Msh5, and Zip3. Evolutionary rate analyses using the Ka/Ks ratio found no change in nonsynonymous substitution patterns in Msh4 and Msh5 in species that had lost Pch2 compared to those retaining Pch2. Elevated Zip3 Ka/Ks values were found in species with pseudogenized Msh4 and Msh5, suggesting possible obligate functional interactions of Zip3 with Msh4 and Msh5. Second, phylogenetically independent contrasts (PIC) analyses were performed on species from the 109-taxon inventory with published chromosome number and chromosome size estimates to investigate whether changes in either parameter were consistently associated with changes in the presence or absence of meiosis genes. Many analyses had low statistical power, neither detecting nor being able to exclude an association between gene loss and the tested variables. However, several comparisons did detect significant or nearly significant trends: for example, fungi that had lost genes related to crossover interference (Msh4, Msh5, or Pch2) tended to have fewer and/or larger chromosomes than their closest relatives without gene loss.
A final objective was to determine the distribution of meiosis genes in lichenized fungi and green algae to see whether this form of symbiosis was associated with differences in the presence or molecular evolution of meiosis genes. Rad51, Dmc1, and Mnd1 were each amplified by degenerate PCR from multiple lichenized fungi that lacked sequenced genomes, and no systematic difference in evolutionary rate was found between examined lichenized fungi compared to other examined classes in phylum Ascomycota. Bioinformatic analyses of meiosis gene distribution in green algae revealed not only no obvious increased tendency for derived gene losses in examined lichenized green algae but also very few derived meiosis gene losses in green algae in general. This suggests that lichenization may not be associated with consistent differences in the evolution of meiosis genes in either fungal or green algal symbionts. The green algal results also illustrate the need to investigate the extent to which eukaryotes as a whole exhibit the same trends of meiosis gene evolution described here for fungi: frequent derived losses of meiosis genes, genes encoding proteins with function interactions showing similar distributions, likely roles for post-transcriptional regulation of meiosis gene transcripts, and loss of crossover distribution-related genes potentially being associated with constraints on chromosome size and/or haploid chromosome number.
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Mitchell, Carter Alexander. "Structural, functional, and computational insights into the ANL superfamily of enzymes." Thesis, State University of New York at Buffalo, 2013. http://pqdtopen.proquest.com/#viewpdf?dispub=3598714.
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<p> Members of the ANL superfamily of enzymes are involved in primary and secondary metabolism throughout all domains of life and identify key pathways that contribute to essential physiological reactions as well as defense mechanisms to evade competition. Specifically, acetyl-CoA synthetases are directly involved in energy metabolism, while NonRibosoaml Peptide Synthetases and some Aryl-CoA Ligases produce secondary natural products that confer virulence for the producing organism. Due to the ANL superfamily's ubiquitous involvement in primary and secondary metabolism, gaining an understanding of how these enzymes work and identifying ways to regulate them could provide an alternative route for antibiotic targets. It is well documented that domain alternation is paramount for the ANL superfamily of enzymes including the adenylation and thioester-forming reactions of NRPS adenylation domains. This thesis utilizes structural and functional analysis in conjunction with computational methods to further our understanding of these unique enzymes. </p><p> In chapter 2 we present the structure of an adenylation:Peptidyl Carrier Protein di-omain NRPS from the cryptic PA1221 biosynthetic operon from <i> Pseudomonas aeruginosa.</i> The PA1221 structure is the second example of an adenylation:PCP in the PDB and validates the chimeric fusion interactions of EntE-B. The similar interacting regions are between the 2<sup>nd</sup> PCP helix and a helix in the N-terminal subdomain of the adenylation domain as well as the loop connecting the longest β-strands of the C-terminal subdomains interacting with loop 1 of the PCP. </p><p> Chapter 3 presents the structure of an acetoacetatyl-CoA Synthetase that is a confirmed substrate for a protein acetyltransferase, PatA, for inactivation through acetylation of the catalytic A10 lysine. This <i>Streptomyces lividans</i> acetoacetyl-CoA synthetase is the first structure to fully resolve the loop connecting C-terminal extension helix to the C-terminal subdomain. The C-terminal extension is only present in ACS proteins revealing an interaction where the C-terminal extension stabilizes the dynamic P-loop in the adenylate forming conformation. </p><p> In chapter 4 we further explore the PA1221 operon by functionally identifying the substrate preference of PA1215, the hypothetical fatty-acyl-CoA Ligase, that is proposed to acylate the charge PCP of PA1221. We computationally validate the substrate preference with a homology model and AutoDock to gain insight into the proteins slow kinetics. We also provide further insight into the biochemistry of a subset of ANL superfamily members, the phenylacetic acid CoA ligases, involved in the utilization of aryl-carboxylic acids as a carbon source as well as the derivatization of penicillin. We analyze their unique dimeric structures identifying structural motifs that are contributed through the dimeric interface, but are otherwise located to different sides of the enzyme in a monomeric form. </p><p> Finally, to help identify how the protein moves between the two productive conformations we subject members of the superfamily to computational dynamic simulations including Anisotropic Network Modeling, Interpolative Elastic Network Modeling, all-atom molecular dynamics, and analyze the output from these methods with Principal Component and Normal Mode Analysis. We developed a method to visualize a dynamic reaction coordinate through measuring the Conformation Determining Angle (defined by structural motifs that are present in superfamily members) and use this metric to interrogate all ANL superfamily member PDB entries for domain organization. Finally, we test our hypothesis that domain alternation proceeds through an extended, open conformation with structural comparisons and MD. Here we report functional and structural analysis of ANL superfamily members that are related through bacterial cell metabolism and natural product biosynthesis.</p>
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Bachman, John. "Identifying Mechanisms of Apoptotic Pore Formation With Programmatic Ensemble Modeling." Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:26718762.
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Mitochondrial outer membrane permeabilization is a key step in the apoptotic cell death program, regulating life-death decisions in response to cytotoxic drugs and other forms of cell stress. In this thesis I use mathematical modeling of a reconstituted biochemical system to identify and integrate mechanisms of apoptotic pore formation. A key bottleneck in using mathematical models to characterize mechanisms has been the difficulty of efficiently creating and revising alternative models and evaluating them against data. This problem is addressed through the use of a software framework, PySB, that allows ensembles of models to be transparently described using tools and approaches from computer programming. These alternative hypotheses can then be evaluated against data using methods from Bayesian statistics for discrimination of models with varying numbers of (possibly non-identifiable) parameters. Using this framework, calibration of a set of models to in vitro kinetic measurements of the membrane insertion of Bax identifies a conformational intermediate associated with BH3-only:Bax complex formation and membrane association but not pore formation. Functional measurements of Bax point mutants from tumors show that the pore formation process can be blocked at the transition into or out of this intermediate, preventing mitochondrial permeabilization. In another study, model-based analysis of Bax insertion and permeabilization kinetics across a range of BH3-only, Bax, and liposome concentrations reveals the context-dependence of the mechanisms regulating pore formation. Bax recruitment is shown to depend on liposome concentration kinetically but not stoichiometrically, whereas cBid recruitment is shown to be limited at high cBid:liposome concentrations. I show that Bax distribution among liposomes is dependent on the presence of pre-existing pores, and that pores grow to include large numbers of Bax monomers but have a minimum size of four subunits. More generally, these studies serve as examples of how ensemble modeling can be used to integrate information about complex mechanisms from disparate sets of experimental observations.<br>Systems Biology
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Bliven, Spencer Edward. "Structure-Preserving Rearrangements| Algorithms for Structural Comparison and Protein Analysis." Thesis, University of California, San Diego, 2015. http://pqdtopen.proquest.com/#viewpdf?dispub=3716489.
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<p> Protein structure is fundamental to a deep understanding of how proteins function. Since structure is highly conserved, structural comparison can provide deep information about the evolution and function of protein families. The Protein Data Bank (PDB) continues to grow rapidly, providing copious opportunities for advancing our understanding of proteins through large-scale searches and structural comparisons. In this work I present several novel structural comparison methods for specific applications, as well as apply structure comparison tools systematically to better understand global properties of protein fold space. </p><p> Circular permutation describes a relationship between two proteins where the N-terminal portion of one protein is related to the C-terminal portion of the other. Proteins that are related by a circular permutation generally share the same structure despite the rearrangement of their primary sequence. This non-sequential relationship makes them difficult for many structure alignment tools to detect. Combinatorial Extension for Circular Permutations (CE-CP) was developed to align proteins that may be related by a circular permutation. It is widely available due to its incorporation into the RCSB PDB website. </p><p> Symmetry and structural repeats are common in protein structures at many levels. The CE-Symm tool was developed in order to detect internal pseudosymmetry within individual polypeptide chains. Such internal symmetry can arise from duplication events, so aligning the individual symmetry units provides insights about conservation and evolution. In many cases, internal symmetry can be shown to be important for a number of functions, including ligand binding, allostery, folding, stability, and evolution. </p><p> Structural comparison tools were applied comprehensively across all PDB structures for systematic analysis. Pairwise structural comparisons of all proteins in the PDB have been computed using the Open Science Grid computing infrastructure, and are kept continually up-to-date with the release of new structures. These provide a network-based view of protein fold space. CE-Symm was also applied to systematically survey the PDB for internally symmetric proteins. It is able to detect symmetry in ~20% of all protein families. Such PDB-wide analyses give insights into the complex evolution of protein folds. </p>
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Hixson, Kim Kathleen. "Network and Multi-Omics Analyses of Arabidopsis Arogenate Dehydratase Knock-Out and Over-Expression Mutants." Thesis, Washington State University, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10785490.
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<p> Arogenate dehydratases (ADTs) are enzymes found within the aromatic amino acid pathway. They are responsible for catalyzing the final step in phenylalanine (Phe) biosynthesis in vascular plants. While being essential for protein production in all living systems, Phe additionally is the starting precursor to a multitude of secondary metabolites produced in the phenylpropanoid pathway. Our group discovered that by knocking out ADT isoenzymes in <i>Arabidopsis thaliana </i>, measurable reductions in lignin levels can be achieved in stem tissue. This finding provides the opportunity to study potential mechanisms related to lignin biosynthesis and could have implications in bioengineering applications where alterations in lignin level might be desired. </p><p> Any alteration to a gene family, as important as that of the ADTs, imparts plant-wide biomolecular changes and because of this, it is not only important to know that lignin is reduced but that optimal plant function is maintained or to understand how it has been changed in order to mediate any undesirable effects. Here we utilized a multitude of analytical platforms and data analysis techniques on both ADT knock-outs (KOs) and over-expression (OE) lines. By using both KO and OE lines we could provide validation to our findings, as KO and OE mutants of the same enzyme/s typically show converse biomolecular abundance changes. As a systems level understanding was desired, we utilized a multi-omics strategy (metabolomics, transcriptomics and proteomics). </p><p> Identified metabolites showed which metabolite and metabolite classes were most affected. Major KEGG defined pathway changes were identified at the transcript and protein enzyme family level. Integration of all omics data revealed which enzymatic reactions were most correlated to observed metabolite abundance changes. Network and clustering algorithms identified patterns of molecular change between metabolites, transcripts and proteins and these patterns were further correlated to reveal possible post-transcriptional regulatory processes involved in lignin biosynthesis. </p><p> Taken altogether, these data informed us of how ADT alterations affect the entire biomolecular system of <i>Arabidopsis</i> and also revealed targets for future studies aimed at elucidating further how lignin biosynthesis is regulated at the post-transcriptional and translational levels.</p><p>
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Hu, Jinnan. "Exploring Genome Structure and Gene Regulation Related to Virulence in Fungal Phytopathogens Using Next Generation Sequencing Techniques." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1366213390.
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Westbrook, Anthony. "The Paladin Suite| Multifaceted Characterization of Whole Metagenome Shotgun Sequences." Thesis, University of New Hampshire, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10685940.
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<p> Whole metagenome shotgun sequencing is a powerful approach for assaying many aspects of microbial communities, including the functional and symbiotic potential of each contributing community member. The research community currently lacks tools that efficiently align DNA reads against protein references, the technique necessary for constructing functional profiles. This thesis details the creation of PALADIN—a novel modification of the Burrows-Wheeler Aligner that provides orders-of-magnitude improved efficiency by directly mapping in protein space. In addition to performance considerations, utilizing PALADIN and associated tools as the foundation of metagenomic pipelines also allows for novel characterization and downstream analysis. </p><p> The accuracy and efficiency of PALADIN were compared against existing applications that employ nucleotide or protein alignment algorithms. Using both simulated and empirically obtained reads, PALADIN consistently outperformed all compared alignment tools across a variety of metrics, mapping reads nearly 8,000 times faster than the widely utilized protein aligner, BLAST. A variety of analysis techniques were demonstrated using this data, including detecting horizontal gene transfer, performing taxonomic grouping, and generating declustered references.</p><p>
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Wiredja, Danica. "Phosphoproteomic Characterization of Systems-Wide Differential Signaling Induced by Small Molecule PP2A Activation." Case Western Reserve University School of Graduate Studies / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=case1511288772792226.
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Chen, Chenxi. "Analysis of the molecular basis of virulence in pathogenic fungi." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1374006951.
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Santos, Tiago Filipe Melo. "Bioinformatics analysis of the Pedobacter sp. NL19 genome." Master's thesis, Universidade de Aveiro, 2015. http://hdl.handle.net/10773/16041.
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Mestrado em Biologia Molecular e Celular<br>The last decades of the 20th century defined the genetic engineering advent, climaxing in the development of techniques, such as PCR and Sanger sequencing. This, permitted the appearance of new techniques to sequencing whole genomes, identified as next-generation sequencing. One of the many applications of these techniques is the in silico search for new secondary metabolites, synthesized by microorganisms exhibiting antimicrobial properties. The peptide antibiotics compounds can be classified in two classes, according to their biosynthesis, in ribosomal or nonribosomal peptides. Lanthipeptides are the most studied ribosomal peptides and are characterized by the presence of lanthionine and methylanthionine that result from posttranslational modifications. Lanthipeptides are divided in four classes, depending on their biosynthetic machinery. In class I, a LanB enzyme dehydrate serine and threonine residues in the C-terminus precursor peptide. Then, these residues undergo a cyclization step performed by a LanC enzyme, forming the lanthionine rings. The cleavage and the transport of the peptide is achieved by the LanP and LanT enzymes, respectively. Although, in class II only one enzyme, LanM, is responsible for the dehydration and cyclization steps and also only one enzyme performs the cleavage and transport, LanT. Pedobacter sp. NL19 is a Gram-negative bacterium, isolated from sludge of an abandon uranium mine, in Viseu (Portugal). Antibacterial activity in vitro was detected against several Gram-positive and Gram-negative bacteria. Sequencing and in silico analysis of NL19 genome revealed the presence of 21 biosynthetic clusters for secondary metabolites, including nonribosomal and ribosomal peptides biosynthetic clusters. Four lanthipeptides clusters were predicted, comprising the precursor peptides, the modifying enzymes (LanB and LanC), and also a bifunctional LanT. This result revealed the hybrid nature of the clusters, comprising characteristics from two distinct classes, which are poorly described in literature. The phylogenetic analysis of their enzymes showed that they clustered within the bacteroidetes clade. Furthermore, hybrid gene clusters were also found in other species of this phylum, revealing that it is a common characteristic in this group. Finally, the analysis of NL19 colonies by MALDI-TOF MS allowed the identification of a 3180 Da mass that corresponds to the predicted mass of a lanthipeptide encoded in one of the clusters. However, this result is not fully conclusive and further experiments are needed to understand the full potential of the compounds encoded in this type of clusters. In conclusion, it was determined that NL19 strain has the potential to produce diverse secondary metabolites, including lanthipeptides that were not functionally characterized so far.<br>O final do século XX marcou o advento da engenharia genética, que culminou com o desenvolvimento de diversas técnicas, como o PCR ou a sequenciação de Sanger. Isto permitiu o aparecimento de novas técnicas de sequenciação de genomas, conhecidas como next-generation sequencing. Uma das suas aplicações é a procura in silico de novos metabolitos secundários, sintetizados por microrganismos e com ação antimicrobiana. Os péptidos antimicrobianos podem ser classificados em péptidos ribossomais e péptidos não-ribossomais, de acordo com a sua biossíntese. Os lantipéptidos são os péptidos ribossomais mais estudados, sendo caracterizados pela presença de lantioninas e metillantioninas na sua estrutura, que resultam de modificações pós-traducionais. Estes podem ser classificados em quatro classes consoante a sua maquinaria de biossíntese. Na classe I, resíduos de serina e treonina são desidratados no terminal C do péptido precursor por uma enzima LanB. Em seguida, estes resíduos sofrem ciclização por ação de uma enzima LanC, formando ligações de lantionina. A clivagem e transporte são posteriormente realizadas por duas enzimas LanP e LanT, respectivamente. Na classe II uma enzima bifuncional LanM é responsável pela desidratação e ciclização, e uma enzima LanT, pela clivagem e transporte. Pedobacter sp. NL19 é uma bactéria de Gram-negativo, isolada a partir de lamas de uma mina de urânio abandonada, em Viseu (Portugal). Possui atividade antimicrobiana in vitro contra várias bactérias de Gram-positivo e de Gram-negativo. A sequenciação e análise do genoma desta bactéria permitiu identificar a presença de 21 clusters biossintéticos para metabolitos secundários, incluindo clusters que codificam para péptidos ribossomais e nãoribossomais. Foram identificados quatro clusters de lantipéptidos contendo péptidos precursores, enzimas de modificação (LanB e LanC) de classe I, e a enzima bifuncional LanT, de classe II. Este resultado revela a existência de clusters de genes híbridos, pouco descritos na literatura, possuindo características de duas classes distintas. A análise filogenética efectuada revelou que as enzimas destes clusters agrupam dentro da clade de bacteroidetes. Assim, verificou-se que outras espécies deste filo também possuem os clusters de gene híbridos de lantipéptidos, mostrando que esta não é uma característica rara neste grupo de organismos. Por fim, a análise de colónias da NL19 por MALDI-TOF MS permitiu detectar uma massa com 3180 Da, correspondente à massa prevista para um lantipéptido codificado por um dos clusters híbridos. Contudo, este resultado não é totalmente conclusivo e mais procedimentos experimentais terão que ser realizados para caracterizar totalmente o potencial destes péptidos. Assim, a análise realizada revelou que a bactéria NL19 possui potencial para produzir diversos metabolitos secundários, incluindo lantipéptidos que não se encontram ainda funcionalmente caracterizados.
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Assis, Luciana Moura de. "Predição de epitopos de célula B em proteínas de Leishmania infantum: uma análise in silico." Programa de Pós-Graduação em Medicina e Saúde, 2013. http://www.repositorio.ufba.br/ri/handle/ri/13148.
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Previous issue date: 2013<br>A Leishmaniose visceral (LV) é uma doença crônica, endêmica em 62 países e representa um sério problema de saúde pública no Brasil. Os testes sorodiagnósticos convencionais empregam antígenos inteiros ou extratos solúveis que limitam a padronização do antígeno, e podem gerar reações cruzadas com outras doenças. Um método alternativo é o uso de peptídeos a partir de epitopos de célula B identificados através de ferramentas de bioinformática. Objetivou-se identificar epitopos lineares e conformacionais de célula B das proteínas de Leishmania infantum cisteína peptidase calpaina-like, redutase thiol dependente 1 (TDR1) e HSP70, bem como identificar sua estrutura secundária através de metodologia in silico; em seguida, buscou-se selecionar os epitopos lineares comuns aos diferentes métodos de predição para verificar a composição dos resíduos de aminoácidos dos mesmos. Metodologia: As ferramentas de bioinformática IEDB, BepiPred e BcePred foram usadas para predição de epitopos lineares de célula B e o programa CBtope para predição de epitopos conformacionais. A estrutura secundária das proteínas foi predita pelo servidor PHD. Resultados: As análises de predição produziram um total de 148 epitopos lineares e 164 epitopos conformacionais a partir das três proteínas, a maioria desses epitopos está localizada na mesma região. A estrutura secundária das proteínas é composta por -hélice, fita estendida e randômica. Nas proteínas TDR1 e HSP70, os epitopos preditos estão localizados principalmente em regiões de -hélice e randômica. Conclusões: Epitopos lineares e conformacionais de célula B de proteínas de L. infantum foram identificados in silico e poderão contribuir como novos antígenos com potencial aplicação no diagnóstico e controle da leishmaniose visceral. Sugere-se que vários métodos de predição de epitopos lineares sejam combinados a fim de se obter resultados mais confiáveis.<br>Salvador
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Raj, Kumar Praveen Kumar. "APPLICATION OF TRANSCRIPTOMICS TO ADDRESS QUESTIONS IN MOLECULAR BIOLOGY AND EVOLUTION." Miami University / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=miami1410427259.
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Stahl, Morgan A. "The Perilipin Family of Proteins: Structural and Bioinformatic Analysis." Otterbein University Honors Theses / OhioLINK, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=otbnhonors1620460421392971.
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Tyler, Scott Robert. "Graph theory analysis of single cell transcriptomes define islet signaling networks and cell identity." Diss., University of Iowa, 2016. https://ir.uiowa.edu/etd/2287.
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Several challenges face bioinformaticians on a regular basis. One of these is unsupervised clustering. In RNA sequencing (RNAseq), this may come in the form of blindly sequencing single cells without a priori knowledge of the cell types being sequenced. Here we create new methods to address this problem that show increased accuracy and speed compared to competing methods. We also have developed a methodology for discovering non-parametric networks which represent relationships between the variables that have been measured across samples. In the context of RNAseq, this is the expression relationships between genes (for example a positive or negative Spearman correlation). We have packaged these techniques into a software tool called PyMINEr. We show the implementation of PyMINEr here in the analysis of single cell RNAseq (scRNAseq), and integrate this dataset with others to yield novel insights to the signaling networks among within and between pancreatic islet cell types. Additionally we used this data to predict the cell type specific importance of Type 2 Diabetes (T2D) single nucleotide polymorphisms (SNPs). Lastly we have demonstrated the use of PyMINEr’s analytic techniques in discovering genetic circuitry underlying the transcriptional networks of two transcription factors (NeuroD1 and Pdx1) in beta cells. We utilized a RNA interference to modulate the expression of these transcription factors in a beta cell line (MIN6), and observe the changes in the transcriptome over time. We used this data to generate graph network models of transcription and integrated them with ChIP-seq of these transcription factors; this enabled annotation of the functional binding sites of these transcription factors. Furthermore, this approach has enabled the discovery of regulators of beta and alpha cell identity. Overall, we have developed novel informatics methods which can be applied to complex datasets to guide bench experiments towards to discovery of molecular signaling networks.
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Almeida, André Atanasio Maranhão 1981. "Comparação algebrica de genomas : o caso da distancia de reversão." [s.n.], 2007. http://repositorio.unicamp.br/jspui/handle/REPOSIP/276265.
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Previous issue date: 2007<br>Resumo: Nas últimas décadas presenciamos grandes avanços na biologia molecular que levaram ao acúmulo de um grande volume de dados acerca de moléculas, tais como DNAs e proteínas, essenciais para a vida e para seu entendimento.O estágio atual é de busca por ferramentas que permitam extrair informações com relevância biológica destes dados. Neste contexto, a comparação de genomas surge como uma das ferramentas e nesta categoria incluímos rearranjo de genomas. Em rearranjo, o genoma é representado por uma seqüência de blocos conservados e, dados dois genomas e um conjunto de operações, busca-se pela que transformem um genoma no outro. Em 1995, Hannenhallie Pevzner apresentaram o primeiro algoritmo polinomial para o problema da ordenação por reversões orientadas. Tal algoritmo executa em tempo O(n4) e foi o primeiro algoritmo polinomial para um modelo realístico de rearranjo de genomas. Desde então, surgiram algoritmos que apresentam desempenho assintoticamente melhor. O melhor deles, apresentado por Tannier e Sagot em 2004, é capaz de executar em tempo O (n(n log n)1/2). Há um algoritmo linear, desenvolvido por Bader e colegas[2], mas este capaz de determinar a seqüência de reversões, apenas calcula a distância. Motivado pela carência de uma derivação algébrica mais formal da teoria desenvolvida em rearranjo de genomas, desenvolvemos uma solução formal para o problema da distância de reversão com sinal. Utilizamos, em tal solução, um formalismo algébrico para rearranjo de genomas que relaciona a recente teoria de rearranjo de genomas ?basicamente fundamentada no trabalho de Hannenhalli e Pevzner ? e a teoria de grupos de permutação de uma nova forma. Pretendemos criar a base para grandes avanços na área através de um formalismo algébrico forte<br>Abstract: In the last decades we have seen a great progress in molecular biology. That lead to a large volume of data on molecules, DNA and proteins, essential for life.The current stage of research lies in the pursuit of tools to extract information with biological relevance from this data. In this context, comparison of genomes is an important tool and genome rearrangements is a way of doing that comparison. In rearrangement analysis the genome is represented by a sequence of conserved blocks. The aim is to ?nd a minimum sequence of operations that transform a genome into another given as input two genomes and a set of allowed operations. In 1995, Hannenhalli and Pevzner presented the ?rst polinomial algorithm for sorting signed permutations by reversals. This algorithm has complexity O(n4) in time and was the ?rst polinomial algorithm for a realistic model of genome rearrangement. Since then, new algorithms with better asintotic performance had appeared. The fastest algorithm, with complexity O(n?n logn), was developed byTannier and Sagot in 2004. Motivated by a lack of a more formal derivation in the genome rearrangement developed theory, we developed a formal solution for the signed reversal distance problem. We use an algebraic formalism that relates the recent genome rearrangement theory ? basically based on a work of Hannenhalli and Pevzner ? to permutation group theory in a new form. We intend to build a solid theoretical base for further advances in the area through strong algebraic formalism<br>Mestrado<br>Teoria da Computação<br>Mestre em Ciência da Computação
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Herdy, Joseph R. III. "SMALL RNA EXPRESSION DURING PROGRAMMED REARRAGEMENT OF A VERTEBRATE GENOME." UKnowledge, 2014. http://uknowledge.uky.edu/biology_etds/25.
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The sea lamprey (Petromyzon marinus) undergoes programmed genome rearrangements (PGRs) during embryogenesis that results in the deletion of ~0.5 Gb of germline DNA from the somatic lineage. The underlying mechanism of these rearrangements remains largely unknown. miRNAs (microRNAs) and piRNAs (PIWI interacting RNAs) are two classes of small noncoding RNAs that play important roles in early vertebrate development, including differentiation of cell lineages, modulation of signaling pathways, and clearing of maternal transcripts. Here, I utilized next generation sequencing to determine the temporal expression of miRNAs, piRNAs, and other small noncoding RNAs during the first five days of lamprey embryogenesis, a time series that spans the 24-32 cell stage to the formation of the neural crest. I obtained expression patterns for thousands of miRNA and piRNA species. These studies identified several thousand small RNAs that are expressed immediately before, during, and immediately after PGR. Significant sequence variation was observed at the 3’ end of miRNAs, representing template-independent covalent modifications. Patterns observed in lamprey are consistent with expectations that the addition of adenosine and uracil residues plays a role in regulation of miRNA stability during the maternal-zygotic transition. We also identified a conserved motif present in sequences without any known annotation that is expressed exclusively during PGR. This motif is similar to binding motifs of known DNA binding and nuclear export factors, and our data could represent a novel class of small noncoding RNAs operating in lamprey.
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Simon, Philippe. "Undersökning av Nosema hos bin : utbredning och förekomst i Sverige." Thesis, Umeå universitet, Institutionen för ekologi, miljö och geovetenskap, 2019. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-163500.
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This report investigates the presence of Nosema in Sweden. Nosema is an intracellular parasite within the order Microsporidia and is considered a global threat. The most recent academic study on the presence of Nosema in Sweden was published in 2013, however in that study no presence of Nosema in the north of Sweden was reported. The aim of this study was to explore the presence of Nosema in Sweden, particularly in the north of Sweden. Samples were gathered from different locations in Sweden, and thereafter analysed (n=74), 54 samples were analysed under microscope. A geographical map was created to establish the range of Nosema on a global perspective. PCR primers and FISH gene probes for molecular identification were evaluated and tested both in SILVA and in an own database created in the bioinformatics software package ‘ARB’. Main findings of the study were that Nosema were detected in two samples in Sweden and that further studies with more sophisticated methods and better sequencing needs to be developed in order to fully investigate the presence of Nosema in the north of Sweden.