Trichomes are hair-like structures found on the surface of virtually all terrestrial plants (Yang et al., 2015). They are epidermal outgrowths that can occur on all of the aerial parts of a plant, varying markedly in size, shape, distribution, and in their ability to produce secondary metabolites. About 30% of all vascular plants carry the glandular trichomes capable of producing secondary metabolites (Glas et al., 2012). Trichomes are vitally important to plants as a defence mechanism, they are highly significant commercially, and they are of interest to plant biologists in that they serve as an excellent model system to study all aspects of plant differentiation at the single-cell level (Hulskamp, 2004). The simple, non-glandular trichomes found in Arabidopsis have been studied extensively. However the glandular trichomes of the kind found on the surface of Antirrhinum are much less well understood. The primary aim of the research reported here is to identify the gene(s) responsible for variation in epidermal hair (trichome) distribution between Antirrhinum species. Following an introduction which provides essential background on trichomes and on Antirrhinum, the thesis is presented in four parts. The first part describes a RAD-seq experiment used to produce linkage maps for the eight chromosomes making up the Antirrhinum genome and estimates the position of the Hairy gene on linkage group 8. The results are cross-validated against maps produced independently by the Xue group. It also describes novel methods developed to address a number of problems that arose during the course of the analysis, and explores the value of imputation methods in helping to overcome gaps and inconsistencies in the data. The second part presents the findings from a fine-mapping Pool-seq experiment designed to estimate the position of Hairy more precisely. The findings suggest that Hairy lies on one of a small number of scaffolds, with Scaffold 1097 being the most likely candidate. Also covered are the findings of another experiment to estimate the position of the gene that determines whether flowers are pale or dark. In this case the results indicate that the gene lies on one of a small number of scaffolds on linkage group 5. The third part presents the results of an RNA-seq experiment which, when combined with the results of the Pool-seq experiment provides evidence that Hairy might be a glutaredoxin gene on Scaffold 1097. Finally the interim results of three experiments designed to confirm that the gene identified as Hairy controls the distribution of trichomes in Antirrhinum are presented.

Thesis advisor: Jeffrey H. Chuang / The field of lipidomics has rapidly progressed since its inception only a decade ago. Technological revolutions in mass spectrometry, chromatography, and computational biology now enables high-throughput high-accuracy quantification of the cellular lipidome. One significant improvement of these technologies is that lipids can now be identified and quantified as individual molecular species. Lipidomics provides an additional layer of information to genomics and proteomics and opens a new opportunity for furthering our understanding of cellular signaling networks and physiology, which have broad therapeutic values. As with other 'omics sciences, these new technologies are producing vast amounts of lipidomic data, which require sophisticated statistical and computational approaches for analysis and interpretation. However, computational tools for utilizing such data are sparse. The complexity of lipid metabolic systems and the fact that lipid enzymes remain poorly understood also present challenges to computational lipidomics. The focus of my dissertation has been the development of novel computational methods for systematic study of lipid metabolism in cellular function and human diseases using lipidomic data. In this dissertation, I first present a mathematical model describing cardiolipin molecular species distribution in steady state and its relationship with fatty acid chain compositions. Knowledge of this relationship facilitates determination of isomeric species for complex lipids, providing more detailed information beyond current limits of mass spectrometry technology. I also correlate lipid species profiles with diseases and predict potential therapeutics. Second, I present statistical studies of mechanisms influencing phosphatidylcholine and phosphatidylethanolamine molecular architectures, respectively. I describe a statistical approach to examine dependence of sn1 and sn2 acyl chain regulatory mechanisms. Third, I describe a novel network inference approach and illustrate a dynamic model of ethanolamine glycerophospholipid acyl chain remodeling. The model is the first that accurately and robustly describes lipid species changes in pulse-chase experiments. A key outcome is that the deacylation and reacylation rates of individual acyl chains can be determined, and the resulting rates explain the well-known prevalence of sn1 saturated chains and sn2 unsaturated chains. Lastly, I summarize and remark on future studies for lipidomics. / Thesis (PhD) — Boston College, 2012. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Medicago sativa (Alfalfa) is an important agricultural plant for animal forage and nitrogen fixation, and has potential value in ligno-cellulosic energy production. In the quest to understand the plant, I generated a draft genome sequence of M. sativa via two reduced representation sequencing approaches: methylation-dependent filtration, and high CoT filtration. Libraries created from each approach were sequenced on an Illumina next-generation sequencing platform yielding approximately 2.5Gb of raw data. A combination of reference-based genome assembly approaches using the closely related species, Medicago truncatula as a reference, and de novo genome assembly approaches were performed to assemble the draft genome. The reference-based assembly generated 312,011 contigs with weighted median contig length (N50) of 247 bases, whereas de novo assembly produced 547,304 contigs with N50 of 275 bases. The creation of the M. sativa draft genome is vital for downstream functional analyses such as genome wide gene mining and gene expression profiling.

Medicago sativa (Alfalfa) is an important agricultural plant for animal forage and nitrogen fixation, and has potential value in ligno-cellulosic energy production. In the quest to understand the plant, I generated a draft genome sequence of M. sativa via two reduced representation sequencing approaches: methylation-dependent filtration, and high CoT filtration. Libraries created from each approach were sequenced on an Illumina next-generation sequencing platform yielding approximately 2.5Gb of raw data. A combination of reference-based genome assembly approaches using the closely related species, Medicago truncatula as a reference, and de novo genome assembly approaches were performed to assemble the draft genome. The reference-based assembly generated 312,011 contigs with weighted median contig length (N50) of 247 bases, whereas de novo assembly produced 547,304 contigs with N50 of 275 bases. The creation of the M. sativa draft genome is vital for downstream functional analyses such as genome wide gene mining and gene expression profiling.

Dentro da bioinformática uma das atividades mais realizadas é o alinhamento de seqüências biológicas [1]. Seus resultados são utilizados em várias atividades que desdobram-se em áreas de pesquisa interdisciplinares com geração de diversos subprodutos. Sendo uma das primeiras etapas de tais tarefas, o multialinhamento é então importante para garantir a qualidade dos resultados obtidos em vários estudos do material genético. Para este trabalho espera-se a reprodução dos resultados já publicados na área [2]; [3]; [4]; [5]; [6]). A implementação de um programa de multialinhamento global de seqüências biológicas utilizando uma abordagem iterativa estocástica por algoritmo genético, uma forma relativamente recente [7] de se atacar tal problema. Obtenção de um panorama sobre as soluções alternativas existentes / Multialignment of biological sequences is one of the most frequently used activities in bioinformatics. The results provided by sequence alignment are used in the solution of other bioinformatics problems. Since a multialignment procedure is one of the first steps of many bioinformatics problems, the condition of an alignment affects the quality of the results obtained for these problems. Multialignment of biological sequences is a complex problem (NP complete) and requires usually heuristics to obtain acceptable performance. Evolutionary algorithms have been used with relevant results. This work aims to find better solutions for the multialignment problem using evolutionary computation. In order to achieve that, this research investigates techniques using evolutionary computation applied to multialignment problem and searches to reproduce their results. Moreover, the development of an approach that performs global multialignment of biological sequences using evolutionary algorithms and an evaluation of the available multialignment techniques are also proposed

Les améliorations des technologies de séquençage offrent aujourd’hui la possibilité d’explorer la variabilité intraspécifique au sein d’une espèce à travers le séquençage complet du génome d’un grand nombre d’individus. Dans ce contexte, mes travaux de thèse se sont basés sur l’étude et la comparaison de la variabilité génomique à travers des études de génomique des populations au sein de plusieurs espèces de levures. Dans un premier temps, j’ai réalisé une étude systématique de la variabilité intraspécifique au sein de 6 espèces de levures, me donnant notamment la possibilité d’étudier la variabilité du contenu en gènes entre les espèces. Dans un second temps, je me suis focalisé sur l’utilisation des dernières technologies de séquençage dans l’objectif de produire une séquence de référence de Dekkera bruxellensis, dont l’absence pour un grand nombre d’espèces limite l’établissement d’étude de génomique des populations. Cette séquence a été utilisée dans un dernier temps afin d’étudier l’évolution de l’espèce. Dans l’ensemble, ces travaux apportent de solides fondations dans l’exploration de la diversité génétique au sein d’espèces non-modèles. / Advent of high throughput technologies as well as the reduction of their price open the way to the exploration of the intraspecific genetic variation at the species level by sequencing the complete genome of a wide range of individuals. Doing so, I first produced populations genomics studies of 6 yeast species based on the same framework, allowing the exploration and comparison of the genes repository of each species. I then used new sequencing technologies to produce a reference sequence for the yeast species Dekkera bruxellensis. Using this sequence, I was then able to produce for the first time a population genomic study at the genome wide scale for this species.

Malaria is a challenging infection with increasing and wide-spread treatment failure risk due to resistance. With a estimated death toll of 1-3 Million per year, most cases of Malaria affect children under the age of five years in Sub-Saharan Africa. In this thesis, I analyse the current status of malaria control (focussing on diagnosis and therapy) in Burkina Faso to show how this disease burdens public health in endemic countries and to identify possible approaches to improvement. MB is discussed as a therapeutic option under these circumstances. Burkina Faso is used as a representative example for a country in Sub-Saharan Africa with high endemicity for malaria and is here portrayed, its health system characterised and discussed under socioeconomic aspects. More than half of this country’s population live in absolute poverty. The burden that malaria, especially treatment cost, poses on these people cannot be under-estimated. A retrospective study of case files from the university pediatric hospital in Burkina Faso’s capital, Ouagadougou, shows that the case load is huge, and especially the specific diagnosis of severe malaria is difficult to apply in the hospital’s daily routine. Treatment policy as proposed by WHO is not satisfactorily implemented neither in home treatment nor in health services, as data for pretreatment clearly show. In the face of growing resistance in malaria parasites, pharmacological combination therapies are important. Artemisinins currently are the last resort of malaria therapy. As I show with homology models, even this golden bullet is not beyond resistance development. Inconsidered mass use has rendered other drugs virtually useless before. Artemisinins should thus be protected similar to reserve antibiotics against multi-resistant bacteria. There is accumulating evidence that MB is an effective drug against malaria. Here the biological effects of both MB alone and in combination therapy is explored via modeling and experimental data. Several different lines of MB attack on Plasmodium redox defense were identified by analysis of the network effects. Next, CQ resistance based on Pfmdr1 and PfCRT transporters as well as SP resistance were modeled in silico. Further modeling shows that MB has a favorable synergism on antimalarial network effects with these commonly used antimalarial drugs, given their correct application. Also from the economic point of view MB shows great potential: in terms of production price, it can be compared to CQ, which could help to diminuish the costs of malaria treatment to affordable ranges for those most affected and struk by poverty. Malaria control is feasible, but suboptimal diagnosis and treatment are often hindering the achievment of this goal. In order to achieve malaria control, more effort has to be made to implement better adjusted and available primary treatment strategies for uncomplicated malaria that are highly standardised. Unfortunately, campaigns against malaria are chronically underfinanced. In order to maximize the effect of available funds, a cheap treatment option is most important, especially as pharmaceuticals represent the biggest single matter of expense in the fight against malaria. / Malaria ist eine Krankheit, die uns vor große Herausforderungen stellt. Insbesondere die weltweit verbreiteten Resistenzen, die viele Therapieoptionen nutzlos werden lassen, haben den Kampf gegen die Malaria in den letzten Jahrzehnten deutlich verkompliziert. Schätzungen gehen davon aus, dass Malaria jährlich 1 bis 3 Millionen Todesopfer fordert. Mortalität und Morbidität der Erkrankung konzentrieren sich dabei in besonderer Weise auf Kinder unter fünf Jahren in Afrika südlich der Sahara. In der hier vorgestellten Doktorarbeit analysiere ich den aktuellen Stand der Malaria-Kontrolle in Burkina Faso und zeige beispielhaft auf, warum diese Krankheit eine derart große Bürde für die Volksgesundheit darstellt und wo Ansatzpunkte zur Verbesserung der Kontrollmaßnahmen zu sehen sind, mit einem besonderen Fokus auf Diagnostik und Therapieoptionen. Dabei wird MB als Therapieoption genauer beleuchtet. Um die besonderen Gegebenheiten eines Landes wie Burkina Faso - welches hier als repräsentatives Beispiel für einen Staat mit hoher Endemizität für Malaria herangezogen wird - aufzuzeigen, wird ein Porträt des Landes und seines Gesundheitssystems insbesondere unter Sozio- Ökonomischen Gesichtspunkten gezeichnet. Burkina Faso ist ein sehr armes Land, über die Hälfte seiner Bevölkerung lebt unterhalb der Armutsgrenze. Die Kosten von Malaria sind für diese Menschen gigantisch, und insbesondere die Kosten von Medikamenten wiegen schwer. Eine retrospektive Studie aus Fallakten des Universitäts-Kinderkrankenhauses in Burkina Fasos Hauptstadt Ouagadougou zeigt vor allem, dass allein die Fallzahlen überwältigend sind, und vor allem die spezifische Diagnose der schweren Verlaufsform der Malaria ist unter den vorherrschenden Bedingungen eine Mammutaufgabe. Die Behandlungsvorschriften wie von der WHO vorgegeben werden weder vom Gesundheitssystem noch von der Therapie zu Hause erfüllt, wie in den präsentierten Daten für die Vorbehandlung zeigen. Die zur Verfügung stehenden Malaria-wirksamen Therapeutika sind leider dank Resistenzentwicklung - oft durch unbedachten Masseneinsatz verursacht - sehr begrenzt. Artemisinine sind momentan das einzige Mittel gegen welches noch keine Resistenzen im Feld nachgewiesen wurden. Mittels Homologie-Modellierung zeige ich auf wie einfach eine solche Resistenzentwicklung jedoch denkbar wäre. Artemisinine sollten daher durch sehr gezielten Einsatz als ”letzter Trumpf” möglichst lange vor Resistenzentwicklung geschützt werden, ähnlich wie Reserveantibiotika gegen Multi-resistente Keime. MB ist ein hervorragender Kandidat für eine Kombinationsbehandlung gegen Malaria und eventuell eine Option, Artemisinine länger zu ”schonen”. Hier wird dieses Medikament mit bioinformatischen Mitteln genauer in seinen Wirkmechanismen beleuchtet und in Kombination mit anderen Medikamenten getestet mittels einer experimentell gestützten bioinformatischen Pathway-Modellierung. Durch diese Netzwerk-Analyse wurden verschiedene Angriffspunkte von MB auf das Redox-Netzwerk der Malariaerreger identifiziert. Daraufhin wurden CQ und SP-Resistenzen in silico simuliert. Weitere Analysen zeigten dabei, dass MB synergisitische Wirkungen mit anderen Therapeutika gegen Malaria aufzeigt, wenn sie zielgerichtet eingesetzt werden. Finanziell gesehen hat MB Potenzial, ein zweites CQ zu werden, und somit endlich wieder die Kosten der Behandlung für Menschen die in Armut leben erschwinglich zu machen. Malaria Kontrolle ist erreichbar, aber suboptimale Diagnosestellung und Behandlung behindern das Erreichen dieses Zieles. Hierfür muss eine angepasste, dezentrale und hochgradig standardisierte Primärbehandlung unkomplizierter Malaria implementiert werden und für eine bessere Verfügbarkeit dieser gesorgt werden. Leider leidet die Finanzierung der Kampagnen gegen Malaria an chronischer Unterversorgung. Um den maximalen Nutzen aus den vorhandenen Mitteln ziehen zu können ist eine günstigere medikamentöse Therapie ein entscheidender Beitrag, zumal Medikamente den größten Einzelbetrag im Kampf gegen Malaria verbrauchen.

Orientador: Maria Isabel Nogueira Cano / Resumo: Os parasitos do gênero Leishmania são protozoários primitivos entre os quais estão os causadores de um espectro de doenças conhecidas como leishmaniose, as quais afetam milhões de pessoas no mundo inteiro, sendo o Brasil um dos países que apresenta maior número de casos por ano. Os tratamentos e vacinas disponíveis para a leishmaniose apresentam problemas como toxicidade, alto custo e baixa eficiência, tornando importante a busca por novos alvos terapêuticos. Dada sua importância para a estabilidade genômica e proliferação celular, os telômeros têm sido considerados como alvos terapêuticos potenciais no tratamento da leishmaniose. Os telômeros são as extremidades físicas dos cromossomos lineares compostos por complexos ribonucleoproteicos que envolvem a interação entre proteínas, DNA e RNA. A maioria dos eucariotos mantêm o tamanho dos telômeros pela ação do complexo telomerase, que é minimamente composto por uma proteína (TERT, Telomerase Reverse Transcriptase) e um RNA longo não codificador (TER, Telomerase RNA). Os dois componentes do complexo telomerase em Leishmania sp. foram identificados, porém não há informação disponível sobre suas estruturas. O objetivo principal deste estudo foi a caracterização estrutural do componente TERT de Leishmania major. Utilizando alinhamentos múltiplos de sequências, foi possível verificar que os quatro domínios estruturais das telomerases canônicas estão presentes no componente TERT em Leishmania sp. Três destes domínios foram estudados ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Parasites of the Leishmania genus are primitive protozoa and among them are the causative agents of a spectrum of diseases called leishmaniasis, which affect millions of people in tropical countries worldwide, being Brazil one of the countries with higher number of cases each year. The drugs or vaccines available to treat leishmaniasis present problems such as toxicity, excessive cost a low efficiency, so it is important to search for new potential therapeutic targets. Due to their importance in genome stability and cell proliferation, telomeres have been considered a potential therapeutic target against leishmaniasis. Telomeres are the ends of linear chromosomes composed by ribonucleoproteic complexes that involve the interaction between proteins, DNA and RNA. Most eukaryotes maintain their telomere length by the action of the telomerase complex, minimally composed by a protein (TERT, Telomerase Reverse Transcriptase) and a long non-coding RNA (TER, Telomerase RNA). Both components of the Leishmania sp. telomerase complex have already been identified, however, little is known about their structure. This study had the aim to characterize the structure of the Leishmania major TERT component. Using multiple sequence alignments, we were able to verify that the four structural domains of canonical telomerases are present in Leishmania sp. Using different bioinformatic approaches three of these domains were independently studied. The TEN (Telomerase essential N-terminal) domain is... (Complete abstract click electronic access below) / Doutor

Dentro da bioinformática uma das atividades mais realizadas é o alinhamento de seqüências biológicas [1]. Seus resultados são utilizados em várias atividades que desdobram-se em áreas de pesquisa interdisciplinares com geração de diversos subprodutos. Sendo uma das primeiras etapas de tais tarefas, o multialinhamento é então importante para garantir a qualidade dos resultados obtidos em vários estudos do material genético. Para este trabalho espera-se a reprodução dos resultados já publicados na área [2]; [3]; [4]; [5]; [6]). A implementação de um programa de multialinhamento global de seqüências biológicas utilizando uma abordagem iterativa estocástica por algoritmo genético, uma forma relativamente recente [7] de se atacar tal problema. Obtenção de um panorama sobre as soluções alternativas existentes / Multialignment of biological sequences is one of the most frequently used activities in bioinformatics. The results provided by sequence alignment are used in the solution of other bioinformatics problems. Since a multialignment procedure is one of the first steps of many bioinformatics problems, the condition of an alignment affects the quality of the results obtained for these problems. Multialignment of biological sequences is a complex problem (NP complete) and requires usually heuristics to obtain acceptable performance. Evolutionary algorithms have been used with relevant results. This work aims to find better solutions for the multialignment problem using evolutionary computation. In order to achieve that, this research investigates techniques using evolutionary computation applied to multialignment problem and searches to reproduce their results. Moreover, the development of an approach that performs global multialignment of biological sequences using evolutionary algorithms and an evaluation of the available multialignment techniques are also proposed

Durant cette thèse nous nous sommes intéressés aux réseaux métaboliques et notamment leur modélisation sous forme d'un graphe bipartite dirigé pondéré. Ce dernier permet d'étudier la production d'éléments cibles métaboliques regroupés dans une biomasse à partir de composants provenant du milieu de croissance de l'organisme. Nous nous sommes plus particulièrement penchés sur le rôle des métabolites internes au réseau et la notion d'essentialité de ces derniers pour la production d'une biomasse dont nous avons raffiné la définition dans le cas d'une étude de flux (métabolite essentiel du point de vue de la productibilité du réseau et métabolite essentiel du point de vue de l'efficacité du réseau) puis étendu cette dernière dans le cas d'une étude topologique (métabolite essentiel du point du vue de la persistance du réseau). Nous nous sommes pour cela reposés sur le formalisme d'un part de Flux Balance Analysis et ses dérivés, et d'autre part d'expansion de réseau, afin de définir un métabolite essentiel (ou carrefour), nous permettant de mettre au point un package python (Conquests) cherchant les carrefours dans un réseau métabolite. Nous avons appliqué ce dernier à six réseaux métaboliques dont quatre provenant d'espèces modèles (iJO1360, iAF1260et iJR904 d'E. coli et Synecchocystis) et les deux autres d'espèces plus spécifiques (A. ferrooxidans et T. lutea). Nous avons aussi défini le concept de cluster de métabolites essentiels du point du vue de la persistance du réseau lié aux composants de la biomasse auxquels ils sont nécessaires et que nous avons appliqué sur les six réseaux métaboliques précédents et sur 3600 réseaux dégradés du réseau iJR904de E. coli puis reconstruits selon trois méthodes de gapfilling (Gapfill, Fastgapfill et Meneco) afin de comparer ces dernières. Ces études nous ont permis de mette en avant l'importance de métabolites internes dans la production de composés cibles. / In this thesis we are interested in metabolic networks and, in particular, their modelling with a weighted directed bipartite graph. This representation makes it possible to study the production of target metabolic elements, constituting a biomass, from components coming from the growth medium of the organism. We focused on the role of metabolites inside the network and the notion of essentiality for this elements for the production of a biomass whose definition we have refined in the case of a flow study (metabolite essential for biomass producibility and metabolite essential for biomass efficiency) and extended this notion in the case of a topological study (metabolite essential for biomass sustainability). We rely on the formalism of Flux Balance Analysis and its derivatives, and of network expansion, in order to define an essential metabolite (ME or crossroad), allowing us to develop a python package (Conquests) looking for crossroads in a metabolic network. We applied our concept to six metabolic networks, four of which came from model species (iJO1360, iAF1260 and iJR904 of E. coli and Synecchocystis) and the other two from more specific species (A. ferrooxidans and T. lutea). We have also defined the concept of cluster of ME-sustainability, related to the biomass components to which they are required and which we have applied over the six previous metabolic networks and over 3600 degraded networks of iJR904 of E. coli and reconstructed according to three methods of gapfilling (Gapfill, Fastgapfill and Meneco) to compare the results. These studies have allowed us to highlight the importance of internal metabolites in the production of target compounds.