Modeling the evolution of regulatory elements by simultaneous detection and alignment with phylogenetic pair HMMs.

The computational detection of regulatory elements in DNA is a difficult but important problem impacting our progress in understanding the complex nature of eukaryotic gene regulation. Attempts to utilize cross-species conservation for this task have been hampered both by evolutionary changes of functional sites and poor performance of general-purpose alignment programs when applied to non-coding sequence. We describe a new and flexible framework for modeling binding site evolution in multiple related genomes, based on phylogenetic pair hidden Markov models which explicitly model the gain and loss of binding sites along a phylogeny. We demonstrate the value of this framework for both the alignment of regulatory regions and the inference of precise binding-site locations within those regions. As the underlying formalism is a stochastic, generative model, it can also be used to simulate the evolution of regulatory elements. Our implementation is scalable in terms of numbers of species and sequence lengths and can produce alignments and binding-site predictions with accuracy rivaling or exceeding current systems that specialize in only alignment or only binding-site prediction. We demonstrate the validity and power of various model components on extensive simulations of realistic sequence data and apply a specific model to study Drosophila enhancers in as many as ten related genomes and in the presence of gain and loss of binding sites. Different models and modeling assumptions can be easily specified, thus providing an invaluable tool for the exploration of biological hypotheses that can drive improvements in our understanding of the mechanisms and evolution of gene regulation.

The prevalence and regulation of antisense transcripts in Schizosaccharomyces pombe.

A strand-specific transcriptome sequencing strategy, directional ligation sequencing or DeLi-seq, was employed to profile antisense transcriptome of Schizosaccharomyces pombe. Under both normal and heat shock conditions, we found that polyadenylated antisense transcripts are broadly expressed while distinct expression patterns were observed for protein-coding and non-coding loci. Dominant antisense expression is enriched in protein-coding genes involved in meiosis or stress response pathways. Detailed analyses further suggest that antisense transcripts are independently regulated with respect to their sense transcripts, and diverse mechanisms might be potentially involved in the biogenesis and degradation of antisense RNAs. Taken together, antisense transcription may have profound impacts on global gene regulation in S. pombe.

An Exploration into Fern Genome Space.

Ferns are one of the few remaining major clades of land plants for which a complete genome sequence is lacking. Knowledge of genome space in ferns will enable broad-scale comparative analyses of land plant genes and genomes, provide insights into genome evolution across green plants, and shed light on genetic and genomic features that characterize ferns, such as their high chromosome numbers and large genome sizes. As part of an initial exploration into fern genome space, we used a whole genome shotgun sequencing approach to obtain low-density coverage (∼0.4X to 2X) for six fern species from the Polypodiales (Ceratopteris, Pteridium, Polypodium, Cystopteris), Cyatheales (Plagiogyria), and Gleicheniales (Dipteris). We explore these data to characterize the proportion of the nuclear genome represented by repetitive sequences (including DNA transposons, retrotransposons, ribosomal DNA, and simple repeats) and protein-coding genes, and to extract chloroplast and mitochondrial genome sequences. Such initial sweeps of fern genomes can provide information useful for selecting a promising candidate fern species for whole genome sequencing. We also describe variation of genomic traits across our sample and highlight some differences and similarities in repeat structure between ferns and seed plants.

Sequence variation in the CHAT locus shows no association with late-onset Alzheimer's disease

There is substantial evidence for a susceptibility gene for late-onset Alzheimer's disease (AD) on chromosome 10. One of the characteristic features of AD is the degeneration and dysfunction of the cholinergic system. The genes encoding choline acetyltransferase (ChAT) and its vesicular transporter (VAChT), CHAT and SLC18A3 respectively, map to the linked region of chromosome 10 and are therefore both positional and obvious functional candidate genes for late-onset AD. We have screened both genes for sequence variants and investigated each for association with late-onset AD in up to 500 late-onset AD cases and 500 control DNAs collected in the UK. We detected a total of 17 sequence variants. Of these, 14 were in CHAT, comprising three non-synonymous variants (D7N in the S exon, A120T in exon 5 and L243F in exon 8), one synonymous change (H547H), nine single-nucleotide polymorphisms in intronic, untranslated or promoter regions, and a variable number of tandem repeats in intron 7. Three non-coding SNPs were detected in SLC18A3. None demonstrated any reproducible association with late-onset AD in our samples. Levels of linkage disequilibrium were generally low across the CHAT locus but two of the coding variants, D7N and A120T, proved to be in complete linkage disequilibrium.

Integrating field sports, hare population management and conservation

Conflicts between field sports, animal welfare and species conservation are frequently contentious. In Ireland, the Irish Coursing Club (ICC) competitively tests the speed and agility of two greyhounds by using a live hare as a lure. Each coursing club is associated with a number of discrete localities, known as preserves, which are managed favourably for hares including predator control, prohibition of other forms of hunting such as shooting and poaching and the maintenance and enhancement of suitable hare habitat. We indirectly tested the efficacy of such management by comparing hare abundance within preserves to that in the wider countryside. In real terms, mean hare density was 18 times higher, and after controlling for variance in habitat remained 3 times higher, within ICC preserves than the wider countryside. Whilst we cannot rule out the role of habitat, our results suggest that hare numbers are maintained at high levels in ICC preserves either because clubs select areas of high hare density and subsequently have a negligible effect on numbers or that active population management positively increases hare abundance. The Irish hare Lepus timidus hibernicus Bell, 1837 is one of the highest priority species for conservation action in Ireland and without concessions for its role in conservation, any change in the legal status Of hare coursing under animal welfare grounds, may necessitate an increase in Government subsidies for conservation on private land together with a strengthened capacity for legislation enforcement.

miR-7 and miR-214 are specifically expressed during neuroblastoma differentiation, cortical development and embryonic stem cells differentiation, and control neurite outgrowth in vitro

The mammalian nervous system exerts essential control on many physiological processes in the organism and is itself controlled extensively by a variety of genetic regulatory mechanisms. microRNA (miR), an abundant class of small non-coding RNA, are emerging as important post-transcriptional regulators of gene expression in the brain. Increasing evidence indicates that miR regulate both the development and function of the nervous system. Moreover, deficiency in miR function has also been implicated in a number of neurological disorders. Expression profile analysis of miR is necessary to understand their complex role in the regulation of gene expression during the development and differentiation of cells. Here we present a comparative study of miR expression profiles in neuroblastoma, in cortical development, and in neuronal differentiation of embryonic stem (ES) cells. By microarray profiling in combination with real time PCR we show that miR-7 and miR-214 are modulated in neuronal differentiation (as compared to miR-1, -16 and -133a), and control neurite outgrowth in vitro. These findings provide an important step toward further elucidation of miR function and miR-related gene regulatory networks in the mammalian central nervous system. (C) 2010 Elsevier Inc. All rights reserved.

Cold-induced silencing by long antisense transcripts of an Arabidopsis Polycomb target.

Transcription in eukaryotic genomes generates an extensive array of non-protein-coding RNA, the functional significance of which is mostly unknown. We are investigating the link between non-coding RNA and chromatin regulation through analysis of FLC - a regulator of flowering time in Arabidopsis and a target of several chromatin pathways. Here we use an unbiased strategy to characterize non-coding transcripts of FLC and show that sense/antisense transcript levels correlate in a range of mutants and treatments, but change independently in cold-treated plants. Prolonged cold epigenetically silences FLC in a Polycomb-mediated process called vernalization. Our data indicate that upregulation of long non-coding antisense transcripts covering the entire FLC locus may be part of the cold-sensing mechanism. Induction of these antisense transcripts occurs earlier than, and is independent of, other vernalization markers and coincides with a reduction in sense transcription. We show that addition of the FLC antisense promoter sequences to a reporter gene is sufficient to confer cold-induced silencing of the reporter. Our data indicate that cold-induced FLC antisense transcripts have an early role in the epigenetic silencing of FLC, acting to silence FLC transcription transiently. Recruitment of the Polycomb machinery then confers the epigenetic memory. Antisense transcription events originating from 3' ends of genes might be a general mechanism to regulate the corresponding sense transcription in a condition/stage-dependent manner.

Defining the Impact of Non-Native Species

Non-native species cause changes in the ecosystems to which they are introduced. These changes, or some of them, are usually termed impacts; they can be manifold and potentially damaging to ecosystems and biodiversity. However, the impacts of most non-native species are poorly understood, and a synthesis of available information is being hindered because authors often do not clearly define impact. We argue that explicitly defining the impact of non-native species will promote progress toward a better understanding of the implications of changes to biodiversity and ecosystems caused by non-native species; help disentangle which aspects of scientific debates about non-native species are due to disparate definitions and which represent true scientific discord; and improve communication between scientists from different research disciplines and between scientists, managers, and policy makers. For these reasons and based on examples from the literature, we devised seven key questions that fall into 4 categories: directionality, classification and measurement, ecological or socio-economic changes, and scale. These questions should help in formulating clear and practical definitions of impact to suit specific scientific, stakeholder, or legislative contexts.

Molecular Medicine and Molecular Pathology for Haematologists:I. Gene Speak made Easy: An Overview of Genes and Gene Expression

Molecular Medicine and Molecular Pathology are integral parts of Haematology as we enter the new millennium. Their origins can be linked to fundamental developments in the basic sciences, particularly genetics, chemistry and biochemistry. The structure of DNA and the genetic code that it encrypts are the critical starting points to our understanding of these new disciplines. The genetic alphabet is a simple one, consisting of just 4 letters, buts its influence is crucial to human development and differentiation. The concept of a gene is not a new one but the Human Genome Project (a joint world-wide effort to characterise our entire genetic make-up) is providing an invaluable understanding of how genes function in normal cellular processes and pinpointing how disruption of these processes can lead to disease. Transcription and translation are the key events by which our genotype is converted to our phenotype (via a messenger RNA intermediate), producing the myriad proteins and enzymes which populate the cellular factory of our body. Unlike the bacterial or prokaryotic genome, the human genome contains a large amount of non coding DNA (less than 1% of our genome codes for proteins), and our genes are interrupted, with the coding regions or exons separated by non coding introns. Precise removal of the intronic material after transcription (though a process called splicing) is critical for efficient translation to occur. Incorrect splicing can lead to the generation of mutant proteins, which can have a dilaterious effect on the phenotype of the individual. Thus the 100,000-200,000 genes which are present in each cell in our body have a defined control mechanism permitting efficient and appropriate expression of proteins and enzymes and yet a single base change in just one of those genes can lead to diseases such as haemophilia or fanconis anaemia.

A new look towards BAC-based array CGH through a comprehensive comparison with oligo-based array CGH

BACKGROUND: Currently, two main technologies are used for screening of DNA copy number; the BAC (Bacterial Artificial Chromosome) and the recently developed oligonucleotide-based CGH (Chromosomal Comparative Genomic Hybridization) arrays which are capable of detecting small genomic regions with amplification or deletion. The correlation as well as the discriminative power of these platforms has never been compared statistically on a significant set of human patient samples.

RESULTS: In this paper, we present an exhaustive comparison between the two CGH platforms, undertaken at two independent sites using the same batch of DNA from 19 advanced prostate cancers. The comparison was performed directly on the raw data and a significant correlation was found between the two platforms. The correlation was greatly improved when the data were averaged over large chromosomic regions using a segmentation algorithm. In addition, this analysis has enabled the development of a statistical model to discriminate BAC outliers that might indicate microevents. These microevents were validated by the oligo platform results.

CONCLUSION: This article presents a genome-wide statistical validation of the oligo array platform on a large set of patient samples and demonstrates statistically its superiority over the BAC platform for the Identification of chromosomic events. Taking advantage of a large set of human samples treated by the two technologies, a statistical model has been developed to show that the BAC platform could also detect microevents.

Aplicação de métodos de customer relationship management ao tratamento de dados para o desenvolvimento de negócios

Tese de Doutoramento, Gestão, na especialidade de Marketing, Faculdade de Economia, Universidade do Algarve, 2007

Ecological genetics in the North Atlantic: environmental gradients and adaptation at specific loci

The North Atlantic intertidal community provides a rich set of organismal and environmental material for the study of ecological genetics. Clearly defined environmental gradients exist at multiple spatial scales: there are broad latitudinal trends in temperature, meso-scale changes in salinity along estuaries, and smaller scale gradients in desiccation and temperature spanning the intertidal range. The geology and geography of the American and European coasts provide natural replication of these gradients, allowing for population genetic analyses of parallel adaptation to environmental stress and heterogeneity. Statistical methods have been developed that provide genomic neutrality tests of population differentiation and aid in the process of candidate gene identification. In this paper, we review studies of marine organisms that illustrate associations between an environmental gradient and specific genetic markers. Such highly differentiated markers become candidate genes for adaptation to the environmental factors in question, but the functional significance of genetic variants must be comprehensively evaluated. We present a set of predictions about locus-specific selection across latitudinal, estuarine, and intertidal gradients that are likely to exist in the North Atlantic. We further present new data and analyses that support and contradict these simple selection models. Some taxa show pronounced clinal variation at certain loci against a background of mild clinal variation at many loci. These cases illustrate the procedures necessary for distinguishing selection driven by internal genomic vs. external environmental factors. We suggest that the North Atlantic intertidal community provides a model system for identifying genes that matter in ecology due to the clarity of the environmental stresses and an extensive experimental literature on ecological function. While these organisms are typically poor genetic and genomic models, advances in comparative genomics have provided access to molecular tools that can now be applied to taxa with well-defined ecologies. As many of the organisms we discuss have tight physiological limits driven by climatic factors, this synthesis of molecular population genetics with marine ecology could provide a sensitive means of assessing evolutionary responses to climate change.

Human factors required for mycobacterium tuberculosis macrophage infection

Tese de doutoramento, Farmácia (Biologia Celular e Molecular), Universidade de Lisboa, Faculdade de Farmácia, 2014

Évaluation et contrôle de l'irrégularité de la prise médicamenteuse : proposition et développement de stratégies rationnelles fondées sur une démarche de modélisations pharmacocinétiques et pharmacodynamiques

L'hétérogénéité de réponses dans un groupe de patients soumis à un même régime thérapeutique doit être réduite au cours d'un traitement ou d'un essai clinique. Deux approches sont habituellement utilisées pour atteindre cet objectif. L'une vise essentiellement à construire une observance active. Cette approche se veut interactive et fondée sur l'échange ``médecin-patient '', ``pharmacien-patient'' ou ``vétérinaire-éleveurs''. L'autre plutôt passive et basée sur les caractéristiques du médicament, vise à contrôler en amont cette irrégularité. L'objectif principal de cette thèse était de développer de nouvelles stratégies d'évaluation et de contrôle de l'impact de l'irrégularité de la prise du médicament sur l'issue thérapeutique. Plus spécifiquement, le premier volet de cette recherche consistait à proposer des algorithmes mathématiques permettant d'estimer efficacement l'effet des médicaments dans un contexte de variabilité interindividuelle de profils pharmacocinétiques (PK). Cette nouvelle méthode est fondée sur l'utilisation concommitante de données \textit{in vitro} et \textit{in vivo}. Il s'agit de quantifier l'efficience ( c-à-dire efficacité plus fluctuation de concentrations \textit{in vivo}) de chaque profil PK en incorporant dans les modèles actuels d'estimation de l'efficacité \textit{in vivo}, la fonction qui relie la concentration du médicament de façon \textit{in vitro} à l'effet pharmacodynamique. Comparativement aux approches traditionnelles, cette combinaison de fonction capte de manière explicite la fluctuation des concentrations plasmatiques \textit{in vivo} due à la fonction dynamique de prise médicamenteuse. De plus, elle soulève, à travers quelques exemples, des questions sur la pertinence de l'utilisation des indices statiques traditionnels ($C_{max}$, $AUC$, etc.) d'efficacité comme outil de contrôle de l'antibiorésistance. Le deuxième volet de ce travail de doctorat était d'estimer les meilleurs temps d'échantillonnage sanguin dans une thérapie collective initiée chez les porcs. Pour ce faire, nous avons développé un modèle du comportement alimentaire collectif qui a été par la suite couplé à un modèle classique PK. À l'aide de ce modèle combiné, il a été possible de générer un profil PK typique à chaque stratégie alimentaire particulière. Les données ainsi générées, ont été utilisées pour estimer les temps d'échantillonnage appropriés afin de réduire les incertitudes dues à l'irrégularité de la prise médicamenteuse dans l'estimation des paramètres PK et PD . Parmi les algorithmes proposés à cet effet, la méthode des médianes semble donner des temps d'échantillonnage convenables à la fois pour l'employé et pour les animaux. Enfin, le dernier volet du projet de recherche a consisté à proposer une approche rationnelle de caractérisation et de classification des médicaments selon leur capacité à tolérer des oublis sporadiques. Méthodologiquement, nous avons, à travers une analyse globale de sensibilité, quantifié la corrélation entre les paramètres PK/PD d'un médicament et l'effet d'irrégularité de la prise médicamenteuse. Cette approche a consisté à évaluer de façon concomitante l'influence de tous les paramètres PK/PD et à prendre en compte, par la même occasion, les relations complexes pouvant exister entre ces différents paramètres. Cette étude a été réalisée pour les inhibiteurs calciques qui sont des antihypertenseurs agissant selon un modèle indirect d'effet. En prenant en compte les valeurs des corrélations ainsi calculées, nous avons estimé et proposé un indice comparatif propre à chaque médicament. Cet indice est apte à caractériser et à classer les médicaments agissant par un même mécanisme pharmacodynamique en terme d'indulgence à des oublis de prises médicamenteuses. Il a été appliqué à quatre inhibiteurs calciques. Les résultats obtenus étaient en accord avec les données expérimentales, traduisant ainsi la pertinence et la robustesse de cette nouvelle approche. Les stratégies développées dans ce projet de doctorat sont essentiellement fondées sur l'analyse des relations complexes entre l'histoire de la prise médicamenteuse, la pharmacocinétique et la pharmacodynamique. De cette analyse, elles sont capables d'évaluer et de contrôler l'impact de l'irrégularité de la prise médicamenteuse avec une précision acceptable. De façon générale, les algorithmes qui sous-tendent ces démarches constitueront sans aucun doute, des outils efficients dans le suivi et le traitement des patients. En outre, ils contribueront à contrôler les effets néfastes de la non-observance au traitement par la mise au point de médicaments indulgents aux oublis

A comparative study of the resilience of coal logistics chains in Australia, South Africa and Canada

Au cours des dernières années l'industrie du charbon a connu un essor important. L'importance du charbon dans l'économie mondiale provient d'une demande mondiale soutenue et de niveaux de production en hausse constante. De ce fait, le nombre élevé d'importateurs et d'exportateurs est à l'origine d'un système d'échange complexe où la compétition est féroce. En effet, un nombre grandissant de pays importateurs se partagent les sources d'approvisionnement tandis qu'un nombre limité de pays exportateurs s'efforcent de répondre à la demande tout en essayant de s'accaparer le plus de parts du marché mondial. L'objectif de cette recherche s'inscrit dans ce contexte en démontrant les bénéfices associés aux chaînes logistiques résilientes pour tout acteur de l'industrie soucieux de devancer la compétition. Une analyse de la logistique de l'industrie du charbon permet entre autres de se pencher sur les questions suivantes: Comment les infrastructures influencent-elles la résilience d'une chaîne logistique? Quels risques est-ce que les catastrophes naturelles présentent pour une chaîne logistique? Comment la gouvernance influence-t-elle la résilience d'une chaîne logistique? Une chaîne logistique représente le trajet effectué par un bien ou produit au cours de son cycle de vie, du point d'origine au point de consommation. Ceci étant dit, le meilleur moyen de régler les problèmes inhérents aux chaînes logistiques est de maintenir de hauts niveaux de résilience. Cette recherche évaluera donc la résilience de chaînes logistiques du charbon des industries australienne, sud-africaine et canadienne. Pour ce faire, trois variables seront étudiées: les infrastructures, les catastrophes naturelles et la gouvernance. La comparaison des trois cas à l'étude se fera par un nombre défini d'indicateurs (12 au total) pour chacune des variables étudiées. Les résultats de cette recherche démontrent que la résilience des trois cas à l'étude se ressemble. Cependant, certaines chaînes logistiques détiennent des avantages comparatifs qui améliorent grandement leur résilience et leur compétitivité. Plusieurs sujets de recherche pourraient être utilisés pour compléter cette recherche. L'analyse comparative pourrait être appliquée à d'autres chaînes logistiques pour vérifier la viabilité des résultats. Une analyse semblable pourrait également être entreprise pour le secteur en aval de la chaîne logistique. Finalement, une méthodologie basée sur des interviews pourrait ajouter un regard différent sur les questions abordées.