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[ES] En las últimas décadas, los avances en la tecnología de secuenciación han producido cantidades significativas de datos genómicos, hecho que ha revolucionado nuestra comprensión de la biología. Sin embargo, la cantidad de datos generados ha superado con creces nuestra capacidad para interpretarlos. Descifrar el código de la vida es un gran reto. A pesar de los numerosos avances realizados, nuestra comprensión del mismo sigue siendo mínima, y apenas estamos empezando a descubrir todo su potencial, por ejemplo, en áreas como la medicina de precisión o la farmacogenómica. El objetivo principal de esta tesis es avanzar en nuestra comprensión de la vida proponiendo una aproximación holística mediante un enfoque basado en modelos que consta de tres artefactos: i) un esquema conceptual del genoma, ii) un método para su aplicación en el mundo real, y iii) el uso de ontologías fundacionales para representar el conocimiento del dominio de una forma más precisa y explícita. Las dos primeras contribuciones se han validado mediante la implementación de sistemas de información genómicos basados en modelos conceptuales. La tercera contribución se ha validado mediante experimentos empíricos que han evaluado si el uso de ontologías fundacionales conduce a una mejor comprensión del dominio genómico. Los artefactos generados ofrecen importantes beneficios. En primer lugar, se han generado procesos de gestión de datos más eficientes, lo que ha permitido mejorar los procesos de extracción de conocimientos. En segundo lugar, se ha logrado una mejor comprensión y comunicación del dominio., [CA] En les últimes dècades, els avanços en la tecnologia de seqüenciació han produït quantitats significatives de dades genòmiques, fet que ha revolucionat la nostra comprensió de la biologia. No obstant això, la quantitat de dades generades ha superat amb escreix la nostra capacitat per a interpretar-los. Desxifrar el codi de la vida és un gran repte. Malgrat els nombrosos avanços realitzats, la nostra comprensió del mateix continua sent mínima, i a penes estem començant a descobrir tot el seu potencial, per exemple, en àrees com la medicina de precisió o la farmacogenómica. L'objectiu principal d'aquesta tesi és avançar en la nostra comprensió de la vida proposant una aproximació holística mitjançant un enfocament basat en models que consta de tres artefactes: i) un esquema conceptual del genoma, ii) un mètode per a la seua aplicació en el món real, i iii) l'ús d'ontologies fundacionals per a representar el coneixement del domini d'una forma més precisa i explícita. Les dues primeres contribucions s'han validat mitjançant la implementació de sistemes d'informació genòmics basats en models conceptuals. La tercera contribució s'ha validat mitjançant experiments empírics que han avaluat si l'ús d'ontologies fundacionals condueix a una millor comprensió del domini genòmic. Els artefactes generats ofereixen importants beneficis. En primer lloc, s'han generat processos de gestió de dades més eficients, la qual cosa ha permés millorar els processos d'extracció de coneixements. En segon lloc, s'ha aconseguit una millor comprensió i comunicació del domini., [EN] Over the last few decades, advances in sequencing technology have produced significant amounts of genomic data, which has revolutionised our understanding of biology. However, the amount of data generated has far exceeded our ability to interpret it. Deciphering the code of life is a grand challenge. Despite our progress, our understanding of it remains minimal, and we are just beginning to uncover its full potential, for instance, in areas such as precision medicine or pharmacogenomics. The main objective of this thesis is to advance our understanding of life by proposing a holistic approach, using a model-based approach, consisting of three artifacts: i) a conceptual schema of the genome, ii) a method for its application in the real-world, and iii) the use of foundational ontologies to represent domain knowledge in a more unambiguous and explicit way. The first two contributions have been validated by implementing genome information systems based on conceptual models. The third contribution has been validated by empirical experiments assessing whether using foundational ontologies leads to a better understanding of the genomic domain. The artifacts generated offer significant benefits. First, more efficient data management processes were produced, leading to better knowledge extraction processes. Second, a better understanding and communication of the domain was achieved., Las fructíferas discusiones y los resultados derivados de los proyectos INNEST2021 /57, MICIN/AEI/10.13039/501100011033, PID2021-123824OB-I00, CIPROM/2021/023 y PDC2021- 121243-I00 han contribuido en gran medida a la calidad final de este tesis.

[ES] La domesticación de los cítricos es un proceso en gran parte desconocido. Las investigaciones llevadas a cabo hasta la fecha indican que las variedades de cítricos que se comercializan en la actualidad son, en general, el producto de introgresiones ancestrales intraespecíficas e interespecíficas. Las introgresiones de tipo intraespecífico tuvieron lugar entre las poblaciones de dos subespecies antiguas de mandarinas y aportaron el comportamiento apomíctico que la mayoría de las mandarinas actuales y otros cítricos relacionados conservan todavía hoy. Una vez dichas mezclas genéticas o admixtures quedaron establecidas, otras introgresiones interespecíficas de pummelo establecieron nuevas características en el genoma de mandarina. Durante el proceso de domesticación, la variabilidad genética de estos cítricos se adquirió fundamentalmente a través de mutación espontánea, y mediante la práctica del injerto los genotipos carentes de semillas se extendieron rápidamente, acentuándose el papel de las mutaciones en la selección de cítricos como las naranjas y las mandarinas modernas. En este trabajo, mostramos que las mutaciones somáticas en cítricos se propagan siguiendo un patrón iterativo determinado por un modelo de ramificación simpodial y, en consecuencia, se agrupan en sectores a lo largo del árbol, donde algunas mutaciones quedan permanentemente fijadas. En este escenario, un árbol puede considerarse un mosaico genéticamente compuesto de diferentes genomas, en el que las ramas más jóvenes acumulan un número mayor de mutaciones. En promedio, nuestro árbol experimental de Clementina de 36 años muestra una tasa de mutación de 4.4 × 10-10 bp-1 yr-1, pudiendo contener en total entre 1500 y 5000 variantes y producir 1 mutación somática en cada meristemo axilar. Este número relativamente alto de mutaciones está en línea con el gran número de variedades derivadas de mutaciones espontáneas que se comercializan en cítricos. Para identificar caracteres esenciales adquiridos durante la domesticación, se han analizado los transcriptomas de la pulpa de frutos en desarrollo de la variedad silvestre e incomestible papeda de Ichang, la mandarina ácida Sun Chu Sha Kat y tres segregantes comestibles derivados de un cruce entre mandarinas comerciales. Basándonos en los resultados obtenidos, se propone que durante la transición de las papedas incomestibles a las mandarinas ácidas, la domesticación se caracterizó por una primera fase de cambios radicales en la expresión de los genes que regulan rutas fundamentales tanto del metabolismo primario como del secundario. Esta fase estuvo determinada principalmente por la estimulación de los procesos de crecimiento y la reducción de las defensas químicas, constituidas por compuestos de sabores desagradables. También se sugiere que en una segunda fase los atributos asociados a la palatabilidad de las mandarinas, especialmente la acidez, se mejoraron progresivamente a través de modificaciones específicas. Otros cambios de relevancia se relacionaron con la regulación de genes involucrados tanto en la síntesis de sustancias básicas de agradable aroma y sabor, como en la modulación de transportadores de azúcares. Por lo tanto, la transición entre las papedas incomestibles y las mandarinas de tipo ácido se caracterizó esencialmente por una drástica reprogramación de la expresión génica de rutas metabólicas fundamentales, mientras que las mandarinas modernas evolucionaron posteriormente mediante un refinamiento progresivo de las propiedades relacionadas con la palatabilidad. En su conjunto, estas observaciones sugieren que en cítricos, las tasas de mutación de los tejidos somáticos son consistentes con la idea de que las mutaciones desempeñaron un papel importante en las últimas fases de la domesticación., [CAT] La domesticació dels cítrics és un procés en gran part desconegut. Les investigacions poratades a terme fins al moment indiquen que les varietats de cítrics comercialitzades hui en dia són, en general, el producte de introgressions ancestrals intraespecífiques i interespecífiques. Les introgressions de tipus intraespecífic van tindre lloc entre les poblacions de dues subespècies antigues de mandarina tot aportant el comportament apomíctic que la majoria de les mandarines actuals i altres cítrics relacionats encara conserven. Una vegada que aquestes barreges genètiques o admixtures es consolidaren, diferents introgressions interespecífiques de pummelo establiren noves característiques en el genoma de mandarina. Durant el procés de domesticació, la variabilitat genètica dels cítrics va ser assolida mitjançant mutació espontània i, per mitjà de la pràctica de l'empelt, genotips sense llavors es van estendre ràpidamente, accentuant el paper de les mutacions en la selecció de cítrics com les taronges i les mandarines modernes. En aquest treball, mostrem com les mutacions somàtiques en cítrics es propaguen seguint un patró iteratiu determinat per un model de ramificació simpodial i, en conseqüència, queden agrupades al llarg de l'arbre, on algunes d'aquestes mutacions romanen fixades. En aquest escenari, un arbre pot ser considerat un mosaic genèticament format per diferents genomes, en el qual les branques més joves acumulen un nombre major de mutacions. De mitjana, el nostre arbre experimental de Clementina mostra una ràtio de mutació de 4.4 × 10−10 bp−1 yr−1, arribant a incloure en total entre 1500 i 5000 variants i produint-hi 1 mutació somática en cada meristem axil·lar. Aquest nombre relativament elevat de mutacions està en línia amb el gran nombre de varietats provinents de mutacions espontànies que es comercialitzen en cítrics. Per tal d'abordar la identificació de trets essencials adquirits durant la domesticació, s'ha analitzat el transcriptoma de fruits en desenvolupament de la varietat silvestre i incomestible papeda d'Ichang, la mandarina àcida Sun Chu Sha Kat i tres segregants comestibles derivats d'un creuament entre mandarines comercials. Basant-nos en els resultats obtinguts, s'ha proposat que durant la transició de les papedes incomestibles a les mandariens àcides, la domesticació es va caracteritzar per una primera fase de canvis radicals en l'expressió dels gens que regulen rutes fonamentals del metabolisme primari i secundari. Aquesta fase va estar determinada principalment per l'estimulació dels processos de creixement i la reducció de les defenses químiques de gust desplaent. També es suggereix que en una segona fase atributs associats a la palatabilitat de les mandarines, especialment l'acidesa, van ser millorats progressivament mitjançant modificacions específiques. Altres canvis de rellevància van estar relacionats amb l'estimulació de gens involucrats tant en la síntesi de substàncies bàsiques d'agradable aroma i sabor, com en la modulació de transportadors de sucres. Per tant, la transició entre les papedes incomestibles i les mandarines àcides va estar caracteritzada essencialment per una dràstica reprogramació de l'expressió gènica de rutes metabòliques fonamentals mentre que les mandarines modernes evolucionaren posteriorment mitjançant un refinament progressiu de propietats relacionades amb la palatabilitat. En conjunt, aquestes observacions suggereixen que, en els cítrics, les ràtios de mutació dels teixits somàtics són consistents amb la idea que les mutacions tingueren un paper important en les últimes fases de la domesticació., [EN] Citrus domestication is a largely unknown process. Research carried out to date indicates that current commercial citrus varieties are, in general, the product of ancestral intraspecific and interspecific introgressions. Intraspecific introgressions took place between the populations of two antique subspecies of mandarins and contributed to the apomictic behavior that most current mandarins and other related citrus still retain today. Once these genetic admixtures were established, interspecific pummelo introgression brought new traits to the admixtured mandarin genome. During domestication, genetic variability of these citrus occurred mainly through spontaneous mutations and with the practice of grafting, seedless genotypes expanded rapidly, emphasizing the role of mutations in the selection of citrus such as oranges and modern mandarins. In this work, we provide evidence that somatic mutations in citrus propagate following an iterative pattern determined by the sympodial branching model and consequently are grouped in sectors along the tree, where some of them remain fixed. In this scenario, a tree can be considered a mosaic genetically composed of different genomes, in which younger branches accumulate greater number of mutations. On average, our 36-yr-old experimental Clementine tree shows a mutation rate of 4.4 × 10-10 bp-1 yr-1, may carry a total of 1,500 to 5,000 variants and produces 1 somatic mutation per axillary meristem. This relatively high number of mutations is in line with the large number of varieties derived from spontaneous mutations that are commercialized in citrus. To identify key traits elicited by citrus domestication, we analyzed transcriptomes from developing fruit pulp of wild inedible Ichang papeda, Sun Chu Sha Kat sour mandarin and three palatable segregants derived from a cross between commercial mandarins. Based on these results, we propose that during the transition from inedible papedas to sour mandarins, domestication involved a first phase of major changes in the expression of genes regulating central pathways of primary and secondary metabolism. This stage was mainly characterized by both the stimulation of growth processes and the reduction of distasteful chemicals defenses. It is also suggested that in a second phase, edible attributes of mandarins, especially acidity, were progressively improved through specific modifications. Other relevant changes included upregulation of genes involved in the synthesis of key substances of pleasant aroma and flavor, and the modulation of sugar transporters. Thus, the transition from inedible papeda to sour mandarin was essentially defined by a drastic reprogramming of gene expression of fundamental metabolic pathways, while modern mandarins evolved later through progressive refinement of palatability properties. Taken together, these observations suggest that the rates of mutations occurring in citrus somatic tissues are consistent with the idea that they have played an important role during the later steps of citrus domestication., This research is co-funded by the Ministerio de Ciencia, Innovación y Universidades (Spain) through grant RTI2018-097790-R-100 and by the European Union through the European Regional Development Fund (ERDF) of the Generalitat Valenciana 2014- 2020, through grants IVIA 51915 and 52002

For successful regeneration, the identity of the missing tissue must be specified according to the pre-existing tissue. Planarians are ideal for the study of the mechanisms underlying this process; the same field of cells can regrow a head or a tail according to the missing body part. After amputation, the differential activation of the Wnt/β-catenin signal specifies anterior versus posterior identity. Initially, both wnt1 and notum (Wnt inhibitor) are expressed in all wounds, but 48 hours later they are restricted to posterior or anterior facing wounds, respectively, by an unknown mechanism. Here we show that 12 hours after amputation, the chromatin accessibility of cells in the wound region changes according to the polarity of the pre-existing tissue in a Wnt/β-catenin-dependent manner. Genomic analyses suggest that homeobox transcription factors and chromatin-remodeling proteins are direct Wnt/β-catenin targets, which trigger the expression of posterior effectors. Finally, we identify FoxG as a wnt1 up-stream regulator, probably via binding to its first intron enhancer region.

[eng] One of the major and most challenging goals of Biomedicine during the last centuries has been the study of the human biological mechanisms, and its relation with traits and diseases. Particularly, in the case of complex diseases, such as Type 2 Diabetes (T2D), asthma or Alzheimer, special interest has been devoted to understanding the underlying molecular mechanisms that affect the development of complex diseases, and the biological processes involved in the preservation of these diseases across generations (genetic basis). In this direction, during the last decades, the advance of computing as well as the development of new DNA-related technologies has largely contributed to the faster development of methods, tools, and resources, which have enhanced the genetic study of traits and diseases. As a result of this revolution, new specialised fields such as Biomedicine, Bioinformatics, and Computational genomics have emerged to find the genomic basis of disease using computational tools. Hence, the identification of the genetic factors behind complex diseases has evolved into a multidisciplinary effort, which combines disciplines as diverse as Biology, Mathematics, Physics, Chemistry, and Information technology. The Computational genomics field, in the context of Biomedicine, focuses on the study of the relationship between genomic changes (variants) and the predisposition or the offset of disease with the final aim of understanding, predict and prevent diseases and, ultimately, to design better treatments. In this direction, numerous contributions have been made in this field to discover variants associated with the risk of developing a disease, and to interpret these associations in terms of function. Notably, some of these contributions, such as the assembly and annotation of the human reference genome, improvements on disease characterization, the better understanding of the effects of genomic variation in different populations, or the introduction of Genome Wide Association Studies (GWAS), have represented very relevant landmarks for the advance on the understanding of the genetic basis of diseases. Particularly, the broad use of GWAS, which mostly relies on the statistical comparison between the variants present in groups of diseased and non-diseased individuals, have led to the discovery of thousands of genomic variants associated with a great diversity of complex traits and diseases. Despite the great success of GWAS, the multiple limitations surrounding this type of approaches, has converted the study of complex diseases into a still challenging problem. Particularly, there are many elements, such as the need of analysing large cohorts of individuals, or the difficulties to generate a complete model to capture the whole complexity of common traits, which limit the discovery power of GWAS. Therefore, reducing the explanation of disease heritability, based on GWAS findings, to a small fraction. Moreover, the lack of biological and functional interpretation of the results obtained from GWAS has complicated its translation into something meaningful to be applied in the clinics. Consequently, many statistical and computational efforts have been devoted to improve GWAS discovery power, and to develop new analytical frameworks to find new disease- susceptibility variants. Additionally, other biological approaches, such as transcriptomics and epigenetics have emerged as a key to facilitate the interpretation of GWAS outcomes. Finally, the need for accessibility to this valuable genomic, transcriptomic and epigenetic information has led to the generation of a wide diversity of publicly available databases. This is the case of Type 2 diabetes (T2D), which is a complex metabolic disorder mainly known to be caused by islet beta-cell dysfunction usually surrounded by a background of insulin resistance. T2D is an example of a common disease that has been broadly studied from the perspective of different omic layers. Particularly, the genetic study of T2D has led to the discovery of more than 700 genomic variants significantly associated with the disease, thousands of genes with a putative effect on the disorder, and thousands of target genomic regions with potential regulatory effects. However, although the genomic explanation of its heritability is estimated around 70%, approximately only 20% has been already explained and, most importantly, the use of these markers to detect the predisposition of an individual to develop the disease is still far for the clinics. Additionally, most of these genomic signals lack functional explanation, thus representing a challenge for the understanding of disease pathophysiology. Consequently, the general objective of this thesis is to broaden the genetic understanding of complex diseases, focusing on the analysis of T2D, by finding new disease-susceptibility loci and improving the functional interpretation of genetic markers. In this direction, the objectives of this thesis can be summarised in: 1) Discover epistatic groups of variants associated with T2D, applying combined machine learning and statistical approaches, and analyse their underlying molecular mechanisms to enhance the early detection of the disease and a better comprehension of its pathophysiology. 2) Generate a comprehensive database of human pancreatic islets gene expression regulatory variation, which integrates genomic, transcriptomic and epigenetic data related to diseases, genes and variants to improve the functional study of T2D and other islets related traits (Alonso, Piron, Morán, & et al., 2021). Additionally, this thesis recapitulates the participation in two studies with the objectives: 3) Support the relevance of inversions and their effect in islets expression to improve the genetic knowledge about the shared-susceptibility of complex diseases (González et al., 2020). 4) Review current GWAS statistical frames to promote the development of new methods and tools that can enhance the study of complex diseases (Alonso, Morán, Salvoro, & Torrents, 2021). Therefore, I start this document with a detailed introduction that aims to facilitate the comprehension and motivation of this study, followed by the hypotheses related to milestones 1-2), and the corresponding list of objectives. This section is followed by a report made by Dr. David Torrents, the director of this thesis, summarising my trajectory during the PhD, and detailing my contributions to the studies related to milestones 1-4) during this period. This report is followed by a brief summary of the studies presented in this thesis. Then, for the study of milestone 1), an unpublished manuscript is provided summarising the preliminary results obtained from the analysis of variant-variant interactions and its association with T2D using machine learning and statistical approaches. Therefore, describing and discussing the last advances done, specifying the methods used, and discussing the outcomes and limitations of the preliminary analyses. Next, a publication is provided to support the results obtained from the study of milestone 2). Thus, detailing and discussing the human pancreatic islets gene expression variation results that constitute the core of the database. Additionally, two appendix sections have been provided in this document to include the publication and review related to milestones 3-4). Finally, the global results obtained from the study of milestones 1) and 2) are summarised and discussed, and a list of conclusions is provided to briefly recapitulate the main outcomes of this thesis.

Ophirinina is a recently described suborder of jakobid protists (Excavata) with only one described species to date, Ophirina amphinema. Despite the acquisition and analysis of massive transcriptomic and mitogenomic sequence data from O. amphinema, its phylogenetic position among excavates remained inconclusive, branching as sister group either to all Jakobida or to all Discoba. From a morphological perspective, it has not only several typical jakobid features but also unusual traits for this group, including the morphology of mitochondrial cristae (sac-shaped to flattened-curved cristae) and the presence of two flagellar vanes. In this study, we have isolated, morphologically characterized, and sequenced genome and transcriptome data of two new Ophirinina species: Ophirina chinija sp. nov. and Agogonia voluta gen. et sp. nov. Ophirina chinija differs from O. amphinema in having rounded cell ends, subapically emerging flagella and a posterior cell protrusion. The much more distantly related A. voluta has several unique ultrastructural characteristics, including sac-shaped mitochondrial cristae and a complex “B” fiber. Phylogenomic analyses with a large conserved-marker dataset supported the monophyly of Ophirina and Agogonia within the Ophirinina and, more importantly, resolved the conflicting position of ophirinids as the sister clade to all other jakobids. The characterization of the mitochondrial genomes showed that Agogonia differs from all known gene-rich jakobid mitogenomes by the presence of two group II introns and their corresponding maturase protein genes. A phylogenetic analysis of the diversity of known maturases confirmed that the Agogonia proteins are highly divergent from each other and define distant families among the prokaryotic and eukaryotic maturases. This opens the intriguing possibility that, compared to other jakobids, Ophirinina may have retained additional mitochondrial elements that may help to understand the early diversification of eukaryotes and the evolution of mitochondria. We thank Dr. Sergey A. Karpov (Zoological Institute RAS) and Dr. Denis V. Tikhonenkov (IBIW RAS) for their helpful discussions on the ultrastructural observations, and Dr. Hwan Su Yoon (Sungkyunkwan University) for the mitochondrial marker alignments. This work was supported by the European Research Council (ERC) Advanced Grants “Protistworld” and “Plast-Evol” (322669 and 787904, respectively) and the Horizon 2020 research and innovation program under the Marie Skłodowska-Curie ITN project SINGEK H2020-MSCA-ITN-2015-675752 (http://www.singek.eu/). G.T. was supported by the 2019 BP 00208 Beatriu de Pinos-3 Postdoctoral Program (BP3; 801370).

IntroductionMansonella species are filarial parasites that infect humans worldwide. Although these infections are common, knowledge of the pathology and diversity of the causative species is limited. Furthermore, the lack of sequencing data for Mansonella species, shows that their research is neglected. Apart from Mansonella perstans, a potential new species called Mansonella sp “DEUX” has been identified in Gabon, which is prevalent at high frequencies. We aimed to further determine if Mansonella sp “DEUX” is a genotype of M. perstans, or if these are two sympatric species.MethodsWe screened individuals in the area of Fougamou, Gabon for Mansonella mono-infections and generated de novo assemblies from the respective samples. For evolutionary analysis, a phylogenetic tree was reconstructed, and the differences and divergence times are presented. In addition, mitogenomes were generated and phylogenies based on 12S rDNA and cox1 were created.ResultsWe successfully generated whole genomes for M. perstans and Mansonella sp “DEUX”. Phylogenetic analysis based on annotated protein sequences, support the hypothesis of two distinct species. The inferred evolutionary analysis suggested, that M. perstans and Mansonella sp “DEUX” separated around 778,000 years ago. Analysis based on mitochondrial marker genes support our hypothesis of two sympatric human Mansonella species.DiscussionThe results presented indicate that Mansonella sp “DEUX” is a new Mansonella species. These findings reflect the neglect of this research topic. And the availability of whole genome data will allow further investigations of these species We acknowledge funding by DZIF (DZIF sequencing project number: TTU 03.820_00) and BMBF (ELIMONCHO-Project: 01DG20021). TG group acknowledges support from the Spanish Ministry of Science and Innovation for grant PID2021-126067NB-I00, cofounded by European Regional Development Fund (ERDF), from the Catalan Research Agency (AGAUR) SGR423; from the European Union’s Horizon 2020 research and innovation program (ERC-2016-724173); from the Gordon and Betty Moore Foundation (Grant GBMF9742); from the “La Caixa” foundation (Grant LCF/PR/HR21/00737), and from the Instituto de Salud Carlos III (IMPACT Grant IMP/0019 and CIBERINFEC CB21/13/00061-ISCIII-SGEFI/ERDF). NGS sequencing methods were performed with the support of the DFG-funded NGS Competence Center Tübingen (INST 37/1049-1). We acknowledge support by Open Access Publishing Fund of University of Tübingen. Peer Reviewed "Article signat per 14 autors/es: Miriam Rodi, Caspar Gross, Thaisa Lucas Sandri, Lilith Berner, Marina Marcet-Houben, Ersoy Kocak, Michaela Pogoda, Nicolas Casadei, Carsten Köhler, Andrea Kreidenweiss, Selidji Todagbe Agnandji, Toni Gabaldón, Stephan Ossowski and Jana Held"

Hybridization is widespread and constitutes an important source of genetic variability and evolution. In animals, its role in generating novel and independent lineages (hybrid speciation) has been strongly debated, with only a few cases supported by genomic data. The South American fur seal (SAfs) Arctocephalus australis is a marine apex predator of Pacific and Atlantic waters, with a disjunct set of populations in Peru and Northern Chile [Peruvian fur seal (Pfs)] with controversial taxonomic status. We demonstrate, using complete genome and reduced representation sequencing, that the Pfs is a genetically distinct species with an admixed genome that originated from hybridization between the SAfs and the Galapagos fur seal ( Arctocephalus galapagoensis ) ~400,000 years ago. Our results strongly support the origin of Pfs by homoploid hybrid speciation over alternative introgression scenarios. This study highlights the role of hybridization in promoting species-level biodiversity in large vertebrates.

Long noncoding RNAs (lncRNAs) are emerging players in cancer and they entail potential as prognostic biomarkers or therapeutic targets. Earlier studies have identified somatic mutations in lncRNAs that are associated with tumor relapse after therapy, but the underlying mechanisms behind these associations remain unknown. Given the relevance of secondary structure for the function of some lncRNAs, some of these mutations may have a functional impact through structural disturbance. Here, we examined the potential structural and functional impact of a novel A > G point mutation in NEAT1 that has been recurrently observed in tumors of colorectal cancer patients experiencing relapse after treatment. Here, we used the nextPARS structural probing approach to provide first empirical evidence that this mutation alters NEAT1 structure. We further evaluated the potential effects of this structural alteration using computational tools and found that this mutation likely alters the binding propensities of several NEAT1-interacting miRNAs. Differential expression analysis on these miRNA networks shows upregulation of Vimentin, consistent with previous findings. We propose a hybrid pipeline that can be used to explore the potential functional effects of lncRNA somatic mutations. Funding information H2020 European Research Council, Grant/Award Number: 724173

La globalización ha generado redes de distribución más complejas y fragmentadas, que conllevan una mayor probabilidad de introducción y reemergencia de microorganismos alterantes y patógenos en alimentos. Tradicionalmente, los agentes microbiológicos patógenos y alterantes en alimentos se han identificado mediante métodos de aislamiento, y consecuente caracterización morfológica, fisiológica y bioquímica. Estos métodos son tediosos y precisan varios días para ser completados, por lo que no responden a las nuevas necesidades de una industria alimentaria global. Los nuevos desafíos en el control de la inocuidad alimentaria requieren de métodos de detección y caracterización precoz, no solo para reducir la morbilidad y la mortalidad relativas a las ETA, sino también para evitar pérdidas económicas tanto al productor como al distribuidor de alimentos. Por tanto, el principal objetivo de esta Tesis Doctoral ha sido desarrollar y optimizar protocolos de identificación y caracterización de bacterias alterantes y patógenos en alimentos, usando tecnologías de última generación (ómicas). Específicamente, el trabajo se ha centrado en el género Enterococcus debido a su naturaleza dual, y a un patógeno emergente como Raoultella ornithinolytica. Para ello se desarrollaron metodologías analíticas sensibles y precisas, basadas en la genómica y en la proteómica.

Esta tesis ha sido financiada por el Ministerio de Ciencia e Innovación a través de la ayuda “DI-17-09134” para contratos para la formación de doctores en empresas (Doctorados Industriales). Con el incremento de datos generados mediante el uso de las tecnologías de secuenciación, es necesario el diseño de protocolos y herramientas que permitan el análisis y la integración de los mismos con el objetivo de entender y comprender los sistemas biológicos que forman parte de cada estudio en particular. Estas herramientas, además, es preferible que sean intuitivas y de fácil manejo para los usuarios, de manera que puedan ser utilizadas por cualquier investigador y no solamente por aquellos que sean expertos o tengan un conocimiento más avanzado en el campo de la bioinformática. En esta tesis se presentan una serie de herramientas destinadas al análisis de datos procedentes de secuenciación para dar soporte a los estudios llevados a cabo en colaboración con distintas instituciones como son el Instituto de Acuicultura Torre de la Sal (IATS-CSIC), el Instituto de Recursos Naturales y Agrobiología de Salamanca (IRNASA), la Fundación Jiménez Díaz, el Museo Nacional de Ciencias Naturales de CSIC y el Hospital General Universitario de Valencia. En primer lugar, se desarrolló e implementó un flujo de trabajo para el ensamblaje de novo y anotación de genomas eucariotas ricos en duplicaciones, el cual se incluyó en la herramienta DeNovoSeq. Este protocolo fue testado ensamblando de novo y anotando el genoma de Sparus aurata (dorada) a partir de datos de secuenciación aportados por el IATS-CSIC. Como resultado, este protocolo no solo permitió la obtención de un borrador de alta calidad del genoma de la dorada y con un tamaño (1,24 Gb) más próximo al esperado según el análisis de k-mer realizado, sino que permitió establecer una hipótesis sobre el origen de la mayor parte de las expansiones sufridas por el genoma de esta especie, sugiriendo que estas derivan de las actividades de los elementos genéticos móviles y de la respuesta inmunitaria como procesos para la adaptabilidad de la especie. En segundo lugar, se rediseñó y adaptó un pipeline, llamado VQS-haplotyper, a partir de un pipeline creado por Mercedes Guerrero-Murillo y Josep Gregori i Font, y basado en el paquete de R llamada QSutils, para la identificación y cuantificación de cuasiespecies en muestras procedentes de pacientes infectados por un virus concreto. En este caso, se utilizaron muestras de pacientes infectados por el virus SARS-CoV-2 proporcionadas por la Fundación Jiménez Díaz. La modificación del pipeline original nos permitió obtener los cambios de nucleótidos y deleciones que caracterizaban los haplotipos presentes en las muestras para una abundancia relativa mínima de 0,5% y 0,1%, obteniendo un total de 105 y 1.154 mutaciones y/o deleciones, respectivamente. De esta manera, VQS-haplotyper es capaz de detectar pequeños cambios en la secuencia de un virus que pueden influir en las características del mismo. En tercer lugar, se desarrollaron e implementaron dos protocolos para el análisis de datos RNA-seq, incluyendo el análisis de enriquecimiento de términos GO y rutas metabólicas, uno a partir de datos procedentes de secuenciación de novo, es decir, sin genoma de referencia disponible, y otro a partir de datos procedentes de resecuenciación, es decir, con genoma de referencia disponible. Estos dos protocolos se implementaron para dar soporte a diversos estudios utilizando muestras de las siguientes especies: Ornithodoros erraticus y Ornithorodos moubata, proporcionadas por el IRNASA; Anisakis pegreffii, Anisakis simplex s.s. y sus híbridos, proporcionadas por el Museo Nacional de Ciencias Naturales de CSIC; Homo sapiens, proporcionadas por el Hospital General Universitario de Valencia. Complementariamente al protocolo para el análisis RNA-seq sin genoma de referencia disponible, ha sido necesario desarrollar un protocolo para el ensamblaje de novo de transcriptomas consenso contra el que, posteriormente, se mapeen las lecturas. Ambas implementaciones han permitido conocer las diferencias entre distintas condiciones de estudio o entre distintas especies. Con ello, es posible establecer potenciales antígenos que puedan ser diana para posibles terapias o vacunas, dilucidar posibles relaciones y diferencias entre distintas especies o descubrir biomarcadores de, por ejemplo, cáncer. Por último, en colaboración con el IATS-CSIC, desarrollamos SAMBA (Structure-Learning of Aquaculture Microbiomes Using a Bayesian-Network Approach), una implementación informática de un modelo de red bayesiano para investigar cómo se relacionan entre sí los pan-microbiomas de los peces y todas las demás variables de un sistema acuícola concreto. SAMBA se basa en un modelo entrenable de red bayesiana que aprende la estructura de red de un sistema de acuicultura utilizando información de distintas variables bióticas y abióticas de importancia en la piscicultura, con especial atención a los datos microbianos proporcionados por la secuenciación de amplicones 16S. SAMBA acepta variables tanto cualitativas como cuantitativas y trata de forma convincente las diferencias en la composición microbiana derivadas de la variación técnica o biológica entre microbiomas de distintos especímenes. Para ello, SAMBA contiene una variedad de herramientas para preanalizar los datos y elegir una distribución para construir y entrenar el modelo de red bayesiana. Una vez creado y validado el modelo, el usuario puede interrogarlo y obtener información sobre el sistema modelizado en dos modos diferentes: “Report” y “Prediction”. En el modo “Report”, SAMBA informa de cómo el pan-microbioma y todas las demás variables que intervienen en el sistema de acuicultura modelizado se influyen mutuamente y cuáles son las probabilidades de cada relación. En el modo “Prediction”, la aplicación predice cómo cambiarían probablemente la diversidad y el perfil funcional del pan-microbioma en función de cualquier cambio realizado en otras variables. Finalmente, SAMBA implementa un completo editor gráfico de redes que permite navegar, editar y exportar los resultados. El funcionamiento de SAMBA ha sido testado y validado utilizando estándares de comunidades microbianas y comunidades de microbiota intestinal de doradas de piscifactoría (Sparus aurata) procedentes de diferentes ensayos de alimentación, arrojando un valor de precisión en todos los casos superior al 0,62. En definitiva, esta tesis ha contribuido no solamente con el desarrollo de protocolos y herramientas que permiten y facilitan el análisis y la integración de diferentes datos NGS, sino que, además, ha contribuido con nuevo conocimiento biológico en diversos campos de estudio. With the increase in data generated through the use of sequencing technologies, there is a need to design protocols and tools that allow the analysis and integration of these data in order to understand and comprehend the biological systems that are part of each particular study. These tools, moreover, should preferably be intuitive and user-friendly, so that they can be employed by any researcher and not only by those that are experts or have a more advanced knowledge in the field of bioinformatics. This thesis presents a series of tools for the analysis of sequencing data so as to provide support for the studies carried out in collaboration with different institutions such as Instituto de Acuicultura Torre de la Sal (IATS-CSIC), Instituto de Recursos Naturales y Agrobiología de Salamanca (IRNASA), Fundación Jiménez Díaz, Museo Nacional de Ciencias Naturales de CSIC and Hospital General Universitario de Valencia. Firstly, a workflow for de novo assembly and annotation of duplication-rich eukaryotic genomes was developed and implemented, and was included in the DeNovoSeq tool. This protocol was tested by de novo assembly and annotation of the Sparus aurata (gilthead sea bream) genome from sequencing data provided by the IATS-CSIC. As a result, this protocol not only allowed us to obtain a high quality draft of the gilthead sea bream genome with a size (1.24 Gb) closer to the expected size according to the k-mer analysis performed, but also allowed us to establish a hypothesis about the origin of most of the expansions suffered by the genome of this species. This suggests that they derive from the activities of the mobile genetic elements and the immune response as processes for the adaptability of the species. Secondly, a pipeline, called VQS-haplotyper, was adapted and redesigned from a pipeline created by Mercedes Guerrero-Murillo and Josep Gregori i Font, and based on the R package called QSutils, to identify and quantify quasispecies in samples from patients infected by a specific virus. In this case, samples from patients infected by the SARS-CoV-2 virus provided by the Fundación Jiménez Díaz were used. The modification of the original pipeline allowed us to obtain nucleotide changes and deletions that characterized the haplotypes present in the samples for a minimum relative abundance of 0.5% and 0.1%, obtaining a total of 105 and 1,154 mutations and/or deletions, respectively. In this way, VQS-haplotyper is able to detect small changes in the sequence of a virus that can influence its characteristics. Thirdly, two protocols were developed and implemented to analyze RNA-seq data, including the enrichment analysis of GO terms and metabolic pathways, one from de novo sequencing data, i.e. with no reference genome available, and the other from resequencing data, i.e. with reference genome available. These two protocols were implemented to provide support for several studies using samples from the following species: Ornithodoros erraticus and Ornithodors moubata, provided by the IRNASA; Anisakis pegreffii, Anisakis simplex s.s. and their hybrids, provided by the Museo Nacional de Ciencias Naturales de CSIC and Homo sapiens, provided by the Hospital General Universitario de Valencia. Complementary to the protocol for RNA-seq analysis with no reference genome available, it has been necessary to develop a protocol for the de novo assembly of consensus transcriptomes against which, subsequently, the reads are mapped. Both implementations have provided insight into the differences between distinct study conditions or between distinct species. With this, it is viable to establish potential antigens that could be targets for possible therapies or vaccines, to elucidate possible relationships and differences between different species or to discover biomarkers of, for example, cancer. Finally, in collaboration with the IATS-CSIC, we developed SAMBA (Structure-Learning of Aquaculture Microbiomes Using a Bayesian-Network Approach), a computer implementation of a Bayesian network model to investigate how fish pan-microbiomes and all other variables in a given aquaculture system are related to each other. SAMBA is powered by a Bayesian network trainable model that learns the network structure of an aquaculture system using information from distinct biotic and abiotic variables of importance in fish farming, with special focus on microbial data provided from 16S amplicon sequencing. SAMBA accepts both qualitative and quantitative variables and convincingly deals with the differences in microbial composition derived by the technical or biological variation among microbiomes of distinct specimens. To this end, SAMBA is implemented with a variety of tools to pre-analyze the data and choose a distribution to build and train the Bayesian network model. Once the model has been created and validated, the user can interrogate the model and obtain information about the modelled system in two different modes: Report and Prediction. Using the Report mode SAMBA reports how the pan-microbiome and all other variables involved in the modelled aquaculture system influence each other and what the conditional probabilities of each relation are. Under the Prediction mode, the application predicts how the diversity and functional profile of the pan-microbiome would likely change depending on any alteration made on other variables. Finally, SAMBA implements a comprehensive graphical network editor allowing the user to navigate, edit and export outcomes. The performance of SAMBA has been tested and validated using microbial community standards and gut microbiota communities of farmed gilthead sea bream (Sparus aurata) from different feeding trials, giving an accuracy value in all cases higher than 0.62. In conclusion, this thesis has not only contributed to the development of protocols and tools that allow us to facilitate the analysis and integration of different NGS data, but has also contributed with new biological knowledge in various fields of study.

Single-nucleotide variants (SNVs) in segmental duplications (SDs) have not been systematically assessed because of the limitations of mapping short-read sequencing data1,2. Here we constructed 1:1 unambiguous alignments spanning high-identity SDs across 102 human haplotypes and compared the pattern of SNVs between unique and duplicated regions3,4. We find that human SNVs are elevated 60% in SDs compared to unique regions and estimate that at least 23% of this increase is due to interlocus gene conversion (IGC) with up to 4.3 megabase pairs of SD sequence converted on average per human haplotype. We develop a genome-wide map of IGC donors and acceptors, including 498 acceptor and 454 donor hotspots affecting the exons of about 800 protein-coding genes. These include 171 genes that have ‘relocated’ on average 1.61 megabase pairs in a subset of human haplotypes. Using a coalescent framework, we show that SD regions are slightly evolutionarily older when compared to unique sequences, probably owing to IGC. SNVs in SDs, however, show a distinct mutational spectrum: a 27.1% increase in transversions that convert cytosine to guanine or the reverse across all triplet contexts and a 7.6% reduction in the frequency of CpG-associated mutations when compared to unique DNA. We reason that these distinct mutational properties help to maintain an overall higher GC content of SD DNA compared to that of unique DNA, probably driven by GC-biased conversion between paralogous sequences5,6. We thank T. Brown for help in editing this manuscript, P. Green for valuable suggestions, and R. Seroussi and his staff for their generous donation of time and resources. This work was supported in part by grants from the US National Institutes of Health (NIH 5R01HG002385, 5U01HG010971 and 1U01HG010973 to E.E.E.; K99HG011041 to P.H.; and F31AI150163 to W.S.D.). W.S.D. was supported in part by a Fellowship in Understanding Dynamic and Multi-scale Systems from the James S. McDonnell Foundation. E.E.E. is an investigator of the Howard Hughes Medical Institute (HHMI). This article is subject to HHMI’s Open Access to Publications policy. HHMI laboratory heads have previously granted a nonexclusive CC BY 4.0 licence to the public and a sublicensable licence to HHMI in their research articles. Pursuant to those licences, the author-accepted manuscript of this article can be made freely available under a CC BY 4.0 licence immediately on publication. Peer Reviewed "Article signat per 19 autors/es: Mitchell R. Vollger, Philip C. Dishuck, William T. Harvey, William S. DeWitt, Xavi Guitart, Michael E. Goldberg, Allison N. Rozanski, Julian Lucas, Mobin Asri, Human Pangenome Reference Consortium, Katherine M. Munson, Alexandra P. Lewis, Kendra Hoekzema, Glennis A. Logsdon, David Porubsky, Benedict Paten, Kelley Harris, PingHsun Hsieh & Evan E. Eichler"

Genetics; Minimal change disease; Paediatric kidney disease Genética; Enfermedad de cambios mínimos; Enfermedad renal pediátrica Genètica; Malaltia de canvis mínims; Malaltia renal pediàtrica Pediatric steroid-sensitive nephrotic syndrome (pSSNS) is the most common childhood glomerular disease. Previous genome-wide association studies (GWAS) identified a risk locus in the HLA Class II region and three additional independent risk loci. But the genetic architecture of pSSNS, and its genetically driven pathobiology, is largely unknown. Here, we conduct a multi-population GWAS meta-analysis in 38,463 participants (2440 cases). We then conduct conditional analyses and population specific GWAS. We discover twelve significant associations-eight from the multi-population meta-analysis (four novel), two from the multi-population conditional analysis (one novel), and two additional novel loci from the European meta-analysis. Fine-mapping implicates specific amino acid haplotypes in HLA-DQA1 and HLA-DQB1 driving the HLA Class II risk locus. Non-HLA loci colocalize with eQTLs of monocytes and numerous T-cell subsets in independent datasets. Colocalization with kidney eQTLs is lacking but overlap with kidney cell open chromatin suggests an uncharacterized disease mechanism in kidney cells. A polygenic risk score (PRS) associates with earlier disease onset. Altogether, these discoveries expand our knowledge of pSSNS genetic architecture across populations and provide cell-specific insights into its molecular drivers. Evaluating these associations in additional cohorts will refine our understanding of population specificity, heterogeneity, and clinical and molecular associations. K.I., K.N., and K.T. were supported by the Japan Agency for Medical Research and Development (AMED) under grant number JP17km0405108h0005. K.T. was supported by the Japan Agency for Medical Research and Development (AMED) under grants JP17km0405205h0002 and 18km0405205h0003. K.I., T.H., C.N., and K.N. were supported by the Japan Society for the Promotion of Science (JSPS) under Grant-in-Aid for Scientific Research fostering Joint International Research (B) 18KK0244. K.I., X.J., T.H., C.N., and K.N. were supported by the Japan Society for the Promotion of Science (JSPS) under Grant-in-Aid for Scientific Research fostering Joint International Research (B) 21KK0147. This work is supported by the Department of Defense (PR190746, PR212415) to S.S-C., by the National Center for Advancing Translational Sciences, National Institutes of Health (Grant Number UL1TR001873) to S.S-C., and by the National Institute of Health Grant RC2DK122397, M.Sam, S.S-C., M.R.P., and F.H. A.M. received support from the American Society of Nephrology KidneyCure Ben J. Lipps Research Fellowship. Y.G. received support from the NEPTUNE Career Development Award. P.R. and H.D. were funded by European Research Council grant ERC-2012- ADG_20120314 (grant agreement 322947) and Agence Nationale pour la Recherche “Genetransnephrose” grant ANR-16-CE17-004-01. M.Sam. was supported by NIH grants R01DK119380, 2U54DK083912, and a gift from The Pura Vida Kidney Foundation. The Nephrotic Syndrome Study Network (NEPTUNE) is part of the Rare Diseases Clinical Research Network (RDCRN), which is funded by the National Institutes of Health (NIH) and led by the National Center for Advancing Translational Sciences (NCATS) through its Division of Rare Diseases Research Innovation (DRDRI). NEPTUNE is funded under grant number U54DK083912 as a collaboration between NCATS and the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK). Additional funding and/or programmatic support is provided by the University of Michigan, NephCure Kidney International, and the Halpin Foundation. RDCRN consortia are supported by the RDCRN Data Management and Coordinating Center (DMCC), funded by NCATS and the National Institute of Neurological Disorders and Stroke (NINDS) under U2CTR002818. The authors wish to thank Seong Kyu Han, Ph.D. (Boston Children’s Hospital and Harvard Medical School) for his assistance in creating figures.

[eng] During the last decade, advances in high-throughput sequencing (HTS) technologies have increased exponentially the generation of genomic data, especially of non-model species. In this thesis, we generated and used available HTS data to shed light on relevant evolutionary processes that shape genome structure and organization, such as the origin and evolution of multigene families, and the genomic determinants of adaptive radiations. First, we contribute to the genome assembly of the spider Dysdera silvatica, the first published assembly of a member of the Dysderidae family. For that we used a hybrid assembly strategy based on short (Illumina) and long (low coverage PacBio and Nanopore) reads to generate an assembly of moderate to low continuity (N50 of 38 kb). Then, we upgraded the assembly to a high quality, chromosome-level, using Chicago and Hi-C libraries, which raised the continuity of the previous draft considerably (N50 of 174.19 Mb). This newly generated genomic resource is not only a significant contribution to the field of spider genomics, but also the backbone for many of the analyses presented in this thesis. We identified 33,275 putative protein-coding genes in the genome of D. silvatica (87% of them were functionally annotated) and estimated that 52.87% of the genome are repetitive elements. We used this high-quality assembly to perform a comprehensive study of the chromosomal location and evolutionary divergence of chemoreceptor gene families. We identified 545 chemoreceptor genes, which are highly clustered in the genome of D. silvatica and show a clear correlation between physical and evolutionary distances. Second, we used available genomics data, and newly generated transcriptomes, to perform a phylogenetically deep study of the evolution of the chemosensory gene families across Panarthropoda. We characterized the expression of chemoreceptor genes in the antenna of the velvet worm Euperipatoides rowelli and performed a comparative genomic analysis of these gene families across representatives of the phyla composing the Panarthropoda clade. Noticeably, we did not find the expression of the Ionotropic co-receptor IR25a in onychophorans, which questions the chemosensory role of this family in this lineage. Surprisingly, eutardigrades lack genes encoding the DEG-ENaC and CD36/SNMPs gene families, a feature that could be related to their extraordinary resistance to desiccation. Finally, we propose the NPC2 gene family as the candidate chemosensory soluble protein in the Panarthropoda ancestor. We also used the chromosome-level assembly of D. silvatica to study some aspects of the adaptive radiation of the spider genus Dysdera in the Canary Islands. We performed a population genomics study of a natural population of D. silvatica from La Gomera using a whole-genome re-sequencing strategy. We found unexpected high levels of nucleotide polymorphism and a lower X/autosomes polymorphism ratio than the theoretically anticipated from their corresponding effective number of chromosomes. Demographic inference based on the coalescence predicted that in the past this species likely underwent a long period characterized by a large effective population size (about 100 times greater than the estimate for the present). We hypothesize that the high levels of polymorphism currently observed in the population of D. silvatica is the molecular hallmark of a long period of ancestral population structure that dominated the history of this species for almost 200 Kya. Globally, our results provide very valuable new knowledge about the genomic bases of adaptation at different evolutionary timescales, ranging from the deep evolutionary dynamics of chemosensory repertories across Panarthropoda phyla, to the very recent origin of new copies and the early steps of their evolution within a genome, including the genomic signals of positive selection and demographic history at the scale of an adaptive radiations. This thesis also demonstrates the great advantage of having highly continuous genome assemblies to obtain reliable data when working gene families, and other repetitive elements, or to perform comprehensive chromosome-level population genomics analyses., [cat] Durant la darrera dècada, els avenços en les tecnologies de High- Throughput Sequencing (HTS) van augmentar exponencialment la generació de dades massives, proporcionant una gran quantitat d'informació genòmica d'espècies no models. L'objectiu principal d'aquesta tesi és investigar processos evolutius com l'origen i l'evolució de les famílies de gens i els mecanismes genòmics subjacents a l'adaptació en la diversificació d'espècies. Per això, en primer lloc, hem contribuït a la construcció de l’assemblatge genòmic de l’aràcnid Dysdera silvatica, el primer genoma de la família Dysderidae amb lectures curtes, amb una petita fracció de lectures llargues i un valor N50 de 38 kb. A continuació, el vam millorar a un genoma a nivell de cromosoma d'alta qualitat, utilitzant llibreries de Chicago i Hi-C, a més d'augmentar el valor de N50 a 174,19 Mb. Vam identificar 33.275 gens (el 87% amb anotacions funcionals), i determinar que el 52,87% del genoma estava representat per elements repetitius. Vàrem identificar 545 gens quimioreceptors, el 54% d'ells distribuïts en 83 clústers genòmics que es trobaven majoritàriament en els scaffolds més petits. D'altra banda, hem estudiat i caracteritzat les famílies multigèniques quimiosensorials a diverses espècies dels grup dels Panarthropoda, centrant-nos en Onychophora i Tardigrada. Hem caracteritzat les principals famílies quimiosensorials de l'onicòfor Euperipatoides rowelli i hem trobat que no tenen el gen que codifica el receptors ionotròpic IR25a, present en la majoria d’espècies de Protòstoms. A més, les famílies de gens DEG-ENaC i CD36/SNMPs són absents a les espècies de la classe Eutardigrada, mentre que la família de gens NPC2 seria l'única que codifica proteïnes solubles en l'ancestre Panartropoda. Finalment, hem estudiat la radiació adaptativa de l’aranya del gènere Dysdera a les Illes Canàries, mitjançant una anàlisi de genòmica de poblacions. Hem determinat que a pesar de ser una espècie endèmica i que viu a illes molt petites, presenta un alt nivell de polimorfisme nucleotídic. Les inferències demogràfiques suggereixen que aquesta espècie té una gran grandària efectiva poblacional, probablement causada per una estructuració poblacional ancestral. Els resultats d'aquesta tesi són un recurs valuós que ens ha permès caracteritzar detalladament les famílies multigèniques del sistema quimiosensorial, a més proporcionar nou coneixement sobre paper relatiu de la selecció positiva en les radiacions adaptatives.

Global food security is endangered by fungal phytopathogens causing devastating crop production losses. Many of these pathogens use specialized appressoria cells to puncture plant cuticles. Here, we unveil a pair of alcohol oxidase–peroxidase enzymes to be essential for pathogenicity. Using Colletotrichum orbiculare, we show that the enzyme pair is cosecreted by the fungus early during plant penetration and that single and double mutants have impaired penetration ability. Molecular modeling, biochemical, and biophysical approaches revealed a fine-tuned interplay between these metalloenzymes, which oxidize plant cuticular long-chain alcohols into aldehydes. We show that the enzyme pair is involved in transcriptional regulation of genes necessary for host penetration. The identification of these infection-specific metalloenzymes opens new avenues on the role of wax-derived compounds and the design of oxidase-specific inhibitors for crop protection. Fungal phytopathogens secrete tandem metalloenzymes that catalyze cuticle oxidation and drive plant cell entry This study was supported by the “Agence Nationale de la Recherche” and by the Natural Sciences and Engineering Research Council of Canada through the ANR-NSERC project “FUNTASTIC” (ANR-17-CE07-0047, STPGP 493781-16). We are grateful to MANE & Fils and the “Association Nationale Recherche Technologie” (ANRT) for funding the Ph.D. fellowship of D.R. (grant no. 2017/1169). Work in Japan was supported by the Japan Society for the Promotion of Science Grants-in-Aid for Scientific Research—KAKENHI, grant numbers 15H05780 and 20H02989 to Y.K., and 20K15529 to S.K. Peer Reviewed "Article signat per 18 autors/es: Bastien Bissaro and Sayo Kodama and Takumi Nishiuchi and Anna Maria Díaz-Rovira and Hayat Hage and David Ribeaucourt and Mireille Haon and Sacha Grisel and A. Jalila Simaan and Fred Beisson and Stephanie M. Forget and Harry Brumer and Marie-Noëlle Rosso and Victor Guallar and Richard O’Connell and Mickaël Lafond and Yasuyuki Kubo and Jean-Guy Berrin "

Breast cancer; Predictive markers; Proteomic analysis Cáncer de mama; Marcadores predictivos; Análisis proteómico Càncer de mama; Marcadors predictius; Anàlisi proteòmic Precision oncology research is challenging outside the contexts of oncogenic addiction and/or targeted therapies. We previously showed that phosphoproteomics is a powerful approach to reveal patient subsets of interest characterized by the activity of a few kinases where the underlying genomics is complex. Here, we conduct a phosphoproteomic screening of samples from HER2-negative female breast cancer receiving neoadjuvant paclitaxel (N = 130), aiming to find candidate biomarkers of paclitaxel sensitivity. Filtering 11 candidate biomarkers through 2 independent patient sets (N = 218) allowed the identification of a subgroup of patients characterized by high levels of CDK4 and filamin-A who had a 90% chance of achieving a pCR in response to paclitaxel. Mechanistically, CDK4 regulates filamin-A transcription, which in turn forms a complex with tubulin and CLIP-170, which elicits increased binding of paclitaxel to microtubules, microtubule acetylation and stabilization, and mitotic catastrophe. Thus, phosphoproteomics allows the identification of explainable factors for predicting response to paclitaxel. M.Q.F. is a recipient of the following grants: AES – PI16/00354 and AES – PI 19/00454 funded by the ISCIII and co-funded by the European Regional Development Fund (ERDF), and B2017/BMD3733 (Immunothercan-CM) – Call for Coordinated Research Groups from the Madrid Region – Madrid Regional Government – ERDF funds. R.C. is a recipient of the ISCIII grant PI17/01865, funded by the ISCIII and co-funded by the European Regional Development Fund (ERDF). Boehringer-Ingelheim contributed with a research grant to this project. This study was also funded by a donation from CRIS Contra el Cancer Foundation.

[Background]: Amphioxus are non-vertebrate chordates characterized by a slow morphological and molecular evolution. They share the basic chordate body-plan and genome organization with vertebrates but lack their 2R whole-genome duplications and their developmental complexity. For these reasons, amphioxus are frequently used as an outgroup to study vertebrate genome evolution and Evo-Devo. Aside from whole-genome duplications, genes continuously duplicate on a smaller scale. Small-scale duplicated genes can be found in both amphioxus and vertebrate genomes, while only the vertebrate genomes have duplicated genes product of their 2R whole-genome duplications. Here, we explore the history of small-scale gene duplications in the amphioxus lineage and compare it to small- and large-scale gene duplication history in vertebrates. [Results]: We present a study of the European amphioxus (Branchiostoma lanceolatum) gene duplications thanks to a new, high-quality genome reference. We find that, despite its overall slow molecular evolution, the amphioxus lineage has had a history of small-scale duplications similar to the one observed in vertebrates. We find parallel gene duplication profiles between amphioxus and vertebrates and conserved functional constraints in gene duplication. Moreover, amphioxus gene duplicates show levels of expression and patterns of functional specialization similar to the ones observed in vertebrate duplicated genes. We also find strong conservation of gene synteny between two distant amphioxus species, B. lanceolatum and B. floridae, with two major chromosomal rearrangements. [Conclusions]: In contrast to their slower molecular and morphological evolution, amphioxus’ small-scale gene duplication history resembles that of the vertebrate lineage both in quantitative and in functional terms., RR acknowledges support from the Swiss National Science Foundation grant 31003A/173048, and from the Etat de Vaud. FMar acknowledges support from the Royal Society grant URF\R1\191161. MI acknowledges support from European Research Council ERC-StG-LS2-637591. HE acknowledges support from the Centre National de la Recherche Scientifique and Agence Nationale de la Recherche (ANR-16-CE12-0008-01 and ANR-19-CE13-0011). PP acknowledges support from the French Government under the “Investissements d’avenir” program managed by the Agence Nationale de la Recherche (Méditerranée Infection 10-IAHU-03). IM acknowledges support from the Spanish Ministry of Science and Innovation and the European Union (RYC-2016-20089 and PGC2018-099392-A-I00). JJT acknowledges support from the ERC (Grant Agreement No. 740041) and the Spanish Ministerio de Ciencia e Innovación (PID2019-103921GB-I00).

The authors thank the Hematopathology Collection registered at the Biobank of Hospital Clínic—Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS) as well as Sílvia Martín for the technical support. This study was supported by the “la Caixa” Foundation (CLLEvolution-LCF/PR/HR17/52150017, Health Research 2017 Program HR17-00221, to EC), the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (810287, BCLLatlas, to EC, and HH), CERCA Program/Generalitat de Catalunya, Generalitat de Catalunya Suport Grups de Recerca AGAUR 2017-SGR-1142 (to EC), CIBERONC (CB16/12/00225 to EC), Ministerio de Ciencia e Innovación PID2020-117185RB-I00 (to XSP), FEDER: European Regional Development Fund “Una manera de hacer Europa”, and Fundación Asociación Española Contra el Cáncer FUNCAR-PRYGN211258SUÁR (to XSP). The authors thankfully acknowledge the computer resources at MareNostrum4 and the technical support provided by Barcelona Supercomputing Center (RES activity BCV-2018-3-0001). FN acknowledge research support from the American Association for Cancer Research (2021 AACR-Amgen Fellowship in Clinical/Translational Cancer Research, Grant Number 21-40-11-NADE), the European Hematology Association (EHA Junior Research Grant 2021, Grant Number RG-202012-00245), and the Lady Tata Memorial Trust (International Award for Research in Leukemia 2021–2022, Grant Number LADY_TATA_21_3223). EC is an Academia Researcher of the “Institució Catalana de Recerca i Estudis Avançats” (ICREA) of the Generalitat de Catalunya. This work was partially developed at the Centre Esther Koplowitz (CEK, Barcelona, Spain). Peer Reviewed "Article signat per 11 autors/es: Romina Royo, Laura Magnano, Julio Delgado, Sara Ruiz-Gil, Josep Ll. Gelpí, Holger Heyn, Malcom A. Taylor, Tatjana Stankovic, Xose S. Puente, Ferran Nadeu & Elías Campo"

Background The periderm is a protective barrier crucial for land plant survival, but little is known about genetic factors involved in its development and regulation. Using a transcriptomic approach in the cork oak (Q. suber) periderm, we previously identified an RS2-INTERACTING KH PROTEIN (RIK) homologue of unknown function containing a K homology (KH)-domain RNA-binding protein, as a regulatory candidate gene in the periderm. Results To gain insight into the function of RIK in the periderm, potato (S. tuberosum) tuber periderm was used as a model: the full-length coding sequence of RIK, hereafter referred to as StRIK, was isolated, the transcript profile analyzed and gene silencing in potato performed to analyze the silencing effects on periderm anatomy and transcriptome. The StRIK transcript accumulated in all vegetative tissues studied, including periderm and other suberized tissues such as root and also in wounded tissues. Downregulation of StRIK in potato by RNA interference (StRIK-RNAi) did not show any obvious effects on tuber periderm anatomy but, unlike Wild type, transgenic plants flowered. Global transcript profiling of the StRIK-RNAi periderm did show altered expression of genes associated with RNA metabolism, stress and signaling, mirroring the biological processes found enriched within the in silico co-expression network of the Arabidopsis orthologue. Conclusions The ubiquitous expression of StRIK transcript, the flower associated phenotype and the differential expression of StRIK-RNAi periderm point out to a general regulatory role of StRIK in diverse plant developmental processes. The transcriptome analysis suggests that StRIK might play roles in RNA maturation and stress response in the periderm This work was supported by the Ministerio de Educación y Ciencia (AGL2006–07342: identification of the gene and obtention of the transgenic lines), the Ministerio de Innovación y Ciencia [AGL2009–13745, FPI grant to P.B.: Transcript profile of the gene in potato, selection of the transgenic lines], the Ministerio de Economía y Competitividad [AGL2012–36725; AGL2015–67495-C2–1-R: analysis of the periderm anatomy, periderm transcriptome and cell localization of the protein], Departament d’Universitats, Investigació i Societat de la Informació de Catalunya [PhD grant to M.S.], the University of Girona [PhD grant to S.F.-P] and The European Regional Development Fund (ERDF)

During the last century, plant breeding of crops and ornamentals has become an important business. Mendel’s principles of inheritance of traits laid the foundation of today’s modern genetics. Although intra- and interspecific cross breeding, chromosome doubling and mutation breeding still form the cornerstone of ornamental plant breeding, the speed of plant breeding has significantly increased since Mendel’s time. New genomic tools now offer possibilities for advanced precision breeding and selection towards more difficult to assess or quantitative traits. Today’s ornamental plant breeding is a complicated exercise, characterized by new and rapidly-changing challenges. Partnerships between industry and research will be essential to implement available technologies in minor crops. Resumo No último século, o melhoramento de plantas de culturas e de ornamentais tornou-se um negócio importante. Os princípios de herança de características de Mendel lançaram as bases da genética moderna de hoje. Embora os cruzamentos intra e interespecífico, a duplicação de cromossomos e o cruzamento de mutações ainda constituam a base do melhoramento de plantas ornamentais, a velocidade do melhoramento de plantas aumentou significativamente desde a época de Mendel. Novas ferramentas genômicas agora oferecem possibilidades para precisão avançada de melhoramento e seleção para características mais difíceis de serem acessadas ou características quantitativas. Atualmente, o melhoramento de plantas ornamentais é um complicado exercício, caracterizado por novos e rápidos desafios. Parcerias entre indústria e pesquisa serão essenciais para implementar as tecnologias disponíveis em culturas menores.

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de Lectura: 18-11-2022, Esta Tesis tiene embargado el acceso al texto completo hasta el 18-05-2024

La tuberculosis (TB) es una enfermedad crónica causada por la bacteria intracelular facultativa Mycobacterium tuberculosis (Mtb), un patógeno altamente exitoso. A pesar de contar con más de 100 años de investigación, la TB actualmente es la enfermedad infecciosa más mortífera por detrás del COVID-19. Se calcula que un cuarto de la población mundial se encuentra infectada latentemente con este patógeno. Los últimos reportes de la organización Mundial de la Salud (OMS) estimaron que alrededor 1.6 millones de personas murieron durante 2021 a causa de esta enfermedad. En particular, las cepas multirresistentes (MDR, resistentes a isoniacida y rifampicina) y extremadamente resistentes (Mtb XDR, resistente a isoniacida, rifampicina, una fluoroquinolona y un aminoglucósido inyectable de segunda línea), representan un problema serio para los sistemas de salud. Argentina, en relación a otros países, tiene una carga media de esta enfermedad. Alrededor de 10500 nuevos casos de TB son notificados anualmente, con cerca de 1000 decesos anuales. Aunque las mismas suelen tratarse y resolverse con una terapia estándar con al menos 3 antibióticos diferentes durante 6-9 meses, no es el caso con las infecciones con Mtb XDR, que requiere un tratamiento más costoso y extendido en el tiempo y muchas veces es de pronóstico reservado. En este trabajo se desarrolló un flujo de trabajo bioinformático (pipeline) con el objetivo de analizar las bases moleculares de resistencia a antibióticos de M. tuberculosis y se aplicó al estudio de los aislamientos XDR circulantes en Argentina entre los años 2008-2016, cuyos genomas fueron obtenidos por técnicas de secuenciación masiva. Las variantes genotípicas obtenidas para cada aislamiento fueron contrastadas contra bases de datos de variantes asociadas a resistencia y se realizó una análisis filogenético sobre las mismas. Con los datos procesados, no se observaron orígenes comunes para las mutaciones de resistencia más frecuentes, pero sí dentro de determinados grupos monofiléticos para algunas drogas. Dentro de las variantes màs frecuentes para las drogas de primera línea se pueden mencionar katG Ser315Thr y fabG1 C-15T para INH, rpoB Ser450Trp y Asp435Val para RIF, embB Gly406Asp y Met306Ile para EMB y variantes en el gen de pncA para PZA. En el marco de la emergencia provocada por las cepas resistentes, multirresistentes y extremadamente resistentes, se requiere con urgencia el desarrollo de nuevos antimicrobianos. A pesar de esta situación crítica, el diseño y la producción de nuevas drogas ha resultado ineficaz por diferentes razones que incluyen el elevado costo asociado al desarrollo de las mismas y una tasa de éxito relativamente baja. El desarrollo de drogas es un proceso complejo que requiere entre 10 y 20 años para atravesar las distintas etapas de investigación y una inversión aproximada de 1500 millones de dólares. En esta situación resulta esencial la adecuada selección inicial del blanco molecular a ser inhibido por el antibiótico en desarrollo. En este sentido, los métodos de secuenciación masiva han sido una enorme fuente de datos, creando nuevas oportunidades para el diseño y desarrollo de drogas para combatir a los agentes infecciosos, incluyendo a los resistentes y multirresistentes. En la presente tesis se ha desarrollado Target-Pathogen, una aplicación web diseñada y desarrollada específicamente como un recurso online que permite la integración y valoración de diferentes características de todas las proteínas de un genoma (función, rol metabólico, homología con proteínas humanas, drogabilidad estructural, esencialidad, expresión) para facilitar la identificación y priorización de proteínas como blancos adecuados. En la base de datos se incluyen los genomas de 25 de los microorganismos más relevantes para la salud humana en continua expansión y actualización. Utilizando Target-Pathogen se seleccionaron y priorizaron una serie de blancos moleculares prometedores para el desarrollo de drogas para Mycobacterium tuberculosis en la fase latente, entre los que se destaca ino1, una proteìna que participa de vìa de la síntesis de micotiol. Tuberculosis (TB) is a chronic disease caused by the intracellular bacterium Mycobacterium tuberculosis (Mtb), a highly successful pathogen. Despite having more than 100 years of research, Mtb is the second leading infectious killer after COVID-19. It is estimated that a quarter of the world population is latently infected with this pathogen. The latest reports from the World Health Organization (WHO) estimated that around 1.6 million people died during 2021 from this disease. In particular, multiresistant strains (MDR, resistant to isoniazid and rifampin) and extremely resistant (Mtb XDR, resistant to isoniazid, rifampin, a fluoroquinolone, and a second-line injectable aminoglycoside), represent a serious problem for health systems. Argentina, in relation to other countries, has a medium burden of this disease. About 10,500 new TB cases are reported annually, with about 1,000 deaths annually. Although they are usually treated and resolved with standard therapy with at least 3 different antibiotics for 6-9 months, this is not the case with Mtb XDR infections, which require more expensive and extended treatment over time and is often guarded prognosis. In this work, a bioinformatic pipeline was developed with the aim of analyzing the molecular bases of resistance to antibiotics in M. tuberculosis and it was applied to the study of XDR strains circulating in Argentina between the years 2008-2016, whose genomes were obtained by techniques of massive sequencing. The genotypic variants obtained for each isolate were compared against databases of variants associated with resistance and on phylogenetic analyses, both locally and globally. With the processed data, no common origins were observed for the most frequent resistance mutations. Among the most frequent variants for first-line drugs, we can mention KatG Ser315Thr and fabG1 C-15T for INH, rpoB Ser450Trp and Asp435Val for RIF, embB Gly406Asp and Met306Ile for EMB, and variants in the pncA gene for PZA. In the context of the emergency caused by resistant, multiresistant and extremely resistant strains, the development of new antimicrobials is urgently required. Despite this critical situation, the design and production of new drugs has been ineffective for different reasons, including the high cost associated with their development and a relatively low success rate. Drug development is a complex process that requires between 10 and 20 years to go through the different stages of research and an investment of approximately 1.5 billion dollars. In this workflow, the adequate initial selection of the molecular target to be inhibited by the antibiotic in development is essential. In this sense, massive sequencing methods have been an enormous source of data, creating new opportunities for the design and development of drugs to combat infectious agents, including resistant and multi-resistant ones. In this thesis we have developed Target-Pathogen, a web platform specifically designed and developed as an online resource that allows the integration and evaluation of different characteristics of all the proteins of a genome (function, metabolic role, homology with human proteins, structural druggability, essentiality, expression) to facilitate the identification and prioritization of proteins as suitable targets. The database includes the genomes of 23 of the most relevant microorganisms for human health in continuous expansion and updating. Using Target-Pathogen, promising molecular targets for the development of drugs for Mycobacterium tuberculosis in the latent phase were selected and prioritized, among which ino1 stands out, a protein that participates in the synthesis of mycothiol. Fil: Sosa, Ezequiel Jorge. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

Advances in genomics and sequencing technologies demand faster and more scalable analysis methods that can process longer sequences with higher accuracy. However, classical pairwise alignment methods, based on dynamic programming (DP), impose impractical computational requirements to align long and noisy sequences like those produced by PacBio, and Nanopore technologies. The recently proposed Wavefront Alignment (WFA) algorithm paves the way for more efficient alignment tools, improving time and memory complexity over previous methods. Notwithstanding the advantages of the WFA algorithm, modern high performance computing (HPC) platforms rely on accelerator-based architectures that exploit parallel computing resources to improve over classical computing CPUs. Hence, a GPU-enabled implementation of the WFA could exploit the hardware resources of modern GPUs and further accelerate sequence alignment in current genome analysis pipelines. This thesis presents two GPU-accelerated implementations based on the WFA for fast pairwise DNA sequence alignment: eWFA-GPU and WFA-GPU. Our first proposal, eWFA-GPU, computes the exact edit-distance alignment between two short sequences (up to a few thousand bases), taking full advantage of the massive parallel capabilities of modern GPUs. We propose a succinct representation of the alignment data that successfully reduces the overall amount of memory required, allowing the exploitation of the fast on-chip memory of a GPU. Our results show that eWFA-GPU outperforms by 3-9X the edit-distance WFA implementation running on a 20 core machine. Compared to other state-of-the-art tools computing the edit-distance, eWFA-GPU is up to 265X faster than CPU tools and up to 56 times faster than other GPU-enabled implementations. Our second contribution, the WFA-GPU tool, extends the work of eWFA-GPU to compute the exact gap-affine distance (i.e., a more general alignment problem) between arbitrary long sequences. In this work, we propose a CPU-GPU co-design capable of performing inter and intra-sequence parallel alignment of multiple sequences, combining a succinct WFA-data representation with an efficient GPU implementation. As a result, we demonstrate that our implementation outperforms the original WFA implementation between 1.5-7.7X times when computing the alignment path, and between 2.6-16X when computing only the alignment score. Moreover, compared to other state-of-the-art tools, the WFA-GPU is up to 26.7X faster than other GPU implementations and up to four orders of magnitude faster than other CPU implementations.

Este artículo contiene 11 páginas, 4 figuras., Mortality at early life stages of fishes is common in nature and can be shaped by stochastic and selective processes. Selective mortality has rarely been assessed in natural conditions but can now be studied by combining genomic data with information on different life stages that realates to fitness. Here we investigate selective mortality between settlers and six-month survivors of the sharpsnout seabream by genotype-phenotype/environmental association studies in three localities along a geographic gradient. We gathered information on 105 individuals at 85,031 SNPs, obtained from individual based 2b-RAD libraries, as well as 9 phenotypic and environmental variables derived from individual otolith readings. We found common signals across localities for potential selection, such as lower survival rates for individuals hatching earlier, growing faster and experiencing higher temperatures during their planktonic phase. We identified 122 loci with parallel significant association to phenotypic and environmental variables. Importantly, one of these loci mapped to the exonic region of the il20rb, a gene involved in immune response, in the phylogenetically closest reference genome, showing parallel frequency changes in non-synonymous mutations in the three studied populations. Further temporal assessments are needed to understand how polymorphisms that are key to selective mortality are maintained., This research was funded by projects PopCOmics (CTM2017-88080, funded by MCIN/ AEI/10.13039/501100011033 and by “ERDF A way of making Europe’’ of the European Union) and MarGeCh (PID2020-118550RB, funded by MCIN/AEI/10.13039/501100011033) from the Spanish Government. The authors CC and MP are members of the research group SGR2017-1120 and EM of SGR2017-378 (Catalan Government). HT was supported by a PhD scholarship funded by the Spanish Ministry of Science, Innovation and Universities (FPU15/02390).

The linear sequence of DNA provides invaluable information about genes and their regulatory elements along chromosomes. However, to fully understand gene function and regulation, we need to dissect how genes physically fold in the three-dimensional nuclear space. Here we describe immuno-OligoSTORM, an imaging strategy that reveals the distribution of nucleosomes within specific genes in super-resolution, through the simultaneous visualization of DNA and histones. We combine immuno-OligoSTORM with restraint-based and coarse-grained modeling approaches to integrate super-resolution imaging data with Hi-C contact frequencies and deconvoluted micrococcal nuclease-sequencing information. The resulting method, called Modeling immuno-OligoSTORM, allows quantitative modeling of genes with nucleosome resolution and provides information about chromatin accessibility for regulatory factors, such as RNA polymerase II. With Modeling immuno-OligoSTORM, we explore intercellular variability, transcriptional-dependent gene conformation, and folding of housekeeping and pluripotency-related genes in human pluripotent and differentiated cells, thereby obtaining the highest degree of data integration achieved so far to our knowledge.

The contribution of germline copy number variants (CNVs) to risk of developing cancer in individuals with pathogenic BRCA1 or BRCA2 variants remains relatively unknown. We conducted the largest genome-wide analysis of CNVs in 15,342 BRCA1 and 10,740 BRCA2 pathogenic variant carriers. We used these results to prioritise a candidate breast cancer risk-modifier gene for laboratory analysis and biological validation. Notably, the HR for deletions in BRCA1 suggested an elevated breast cancer risk estimate (hazard ratio (HR) = 1.21), 95% confidence interval (95% CI = 1.09–1.35) compared with non-CNV pathogenic variants. In contrast, deletions overlapping SULT1A1 suggested a decreased breast cancer risk (HR = 0.73, 95% CI 0.59-0.91) in BRCA1 pathogenic variant carriers. Functional analyses of SULT1A1 showed that reduced mRNA expression in pathogenic BRCA1 variant cells was associated with reduced cellular proliferation and reduced DNA damage after treatment with DNA damaging agents. These data provide evidence that deleterious variants in BRCA1 plus SULT1A1 deletions contribute to variable breast cancer risk in BRCA1 carriers.

Background Tracking person-to-person SARS-CoV-2 transmission in the population is important to understand the epidemiology of community transmission and may contribute to the containment of SARS-CoV-2. Neither contact tracing nor genomic surveillance alone, however, are typically sufficient to achieve this objective. Aim We demonstrate the successful application of the integrated genomic surveillance (IGS) system of the German city of Düsseldorf for tracing SARS-CoV-2 transmission chains in the population as well as detecting and investigating travel-associated SARS-CoV-2 infection clusters. Methods Genomic surveillance, phylogenetic analysis, and structured case interviews were integrated to elucidate two genetically defined clusters of SARS-CoV-2 isolates detected by IGS in Düsseldorf in July 2021. Results Cluster 1 (n = 67 Düsseldorf cases) and Cluster 2 (n = 36) were detected in a surveillance dataset of 518 high-quality SARS-CoV-2 genomes from Düsseldorf (53% of total cases, sampled mid-June to July 2021). Cluster 1 could be traced back to a complex pattern of transmission in nightlife venues following a putative importation by a SARS-CoV-2-infected return traveller (IP) in late June; 28 SARS-CoV-2 cases could be epidemiologically directly linked to IP. Supported by viral genome data from Spain, Cluster 2 was shown to represent multiple independent introduction events of a viral strain circulating in Catalonia and other European countries, followed by diffuse community transmission in Düsseldorf. Conclusion IGS enabled high-resolution tracing of SARS-CoV-2 transmission in an internationally connected city during community transmission and provided infection chain-level evidence of the downstream propagation of travel-imported SARS-CoV-2 cases.

In patients with early-stage endocrine receptor-positive (ER+) breast cancer (BC), adjuvant endocrine therapy (ET) for 5 years is the standard of care. However, for some patients, the risk of recurrence remain high for up to 15 years after diagnosis and extended ET beyond 5 years may be a reasonable option. Nevertheless, this strategy significantly increases the occurrence of side effects. Here we summarize the available evidence from randomized clinical trials on the efficacy and safety profile of extended ET and discuss available clinical and genomic tools helpful to select eligible patients in daily clinical practice.

El splicing se encuentra presente a lo largo de las distintas especies de organismos eucariotas, representando una pieza clave en la inmensa mayoría de los procesos de regulación. Para que el mismo pueda llevarse a cabo, debe darse el reconocimiento de una serie de señales. En particular, los sitios donores y aceptores de splicing definen los límites del intrón y su reconocimiento constituye uno de los primeros pasos dentro del ciclo de splicing. A lo largo del genoma, estos sitios presentan una cierta variabilidad en su secuencia, la cual se encuentra relacionada con la posibilidad de regular su reconocimiento y establecer diversos patrones de splicing alternativo. Por otro lado, este reconocimiento se encuentra influido por la acción de una gran variedad de factores en trans que son reclutados al lugar donde se está llevando a cabo el splicing. En la primera parte de esta tesis, buscamos analizar la variabilidad de secuencia que presentan los sitios donores a lo largo de diversas especies eucariotas. Para esto construimos un modelo estadístico de máxima entropía para determinar patrones de correlación no triviales entre los distintos pares de posiciones que constituyen los sitios donores de splicing, en busca de determinar cuáles de éstos son comunes entre las 30 especies analizadas, y cuáles son característicos de solo algunos grupos. Si bien el proceso de splicing conserva un gran número de elementos en común en los organismos eucariotas, en los últimos años múltiples estudios han destacado la posible relevancia funcional de pequeñas diferencias entre las especies. Así logramos establecer patrones de correlación característicos en los sitios donores de splicing que distinguen a las especies vegetales de los metazoos y los hongos. En la segunda parte, nos abocamos al estudio del efecto sobre el splicing de PRMT5, un factor que, mediante la metilación de proteínas, participa de la regulación de múltiples procesos moleculares. Se ha propuesto que la acción de PRMT5 podría favorecer el reconocimiento de sitios donores débiles. Con el objetivo de determinar en qué medida el efecto que tiene PRMT5 sobre los patrones de splicing se encuentra influído por señales en cis, se realizaron experimentos de secuenciación de ARN (RNA-Seq) en plantas de Arabidopsis thaliana mutantes de PRMT5, pertenecientes a dos ecotipos distintos: Columbia (Col-0) y Landsberg erecta (Ler). De esta manera se buscó analizar el efecto que tiene la mutación ante la variabilidad genética que presentan estos ecotipos. Para poder discriminar los cambios relacionados con variaciones de las secuencias en cis, se analizó también híbridos F1 Col-0 X Ler y Ler X Col-0. Mediante estos análisis se llegó a la conclusión de que una parte importante de los patrones de splicing afectados por la mutación de PRMT5 dependen de señales en cis, teniendo una particular relevancia la fortaleza del sitio donor. Splicing is present throughout eukaryotic organisms, representing a key part in a large number of regulatory processes. To be carried out, there must be recognition of a series of signals present both within and around the intron. In particular, the splicing donor and acceptor sites define the boundaries of the intron and are critical for its proper recognition. Throughout the genome, these sites present a certain variability in their sequence, which is related to the possibility of regulating their recognition and establishing different patterns of alternative splicing. On the other hand, this recognition is influenced by the action of a wide variety of factors in trans that are recruited. In the first part of this work, we analyzed the sequence variability of donor sites across various eukaryotic species. We built a maximum entropy statistical model to determine non-trivial correlation patterns between the different pairs of positions that constitute the splicing donor sites, seeking to determine which of these are common among the 30 species analyzed, and which are characteristic of just some groups. Although the splicing process preserves a large number of elements in common between the different species, in recent years multiple studies have highlighted the possible functional relevance of small interspecific differences. Using this approach, we were able to establish characteristic correlation patterns in splice donor sites that distinguish plant species from metazoans and fungi. In the second part of this thesis, we study the effect of PRMT5 on splicing. PRMT5 is a Protein arginine methyltransferase that participates in the regulation of multiple molecular processes, such as chromatin modification and alternative splicing. It has been proposed that the action of PRMT5 could favor the recognition of weak donor sites. In order to deter mine to what extent the effect PRMT5 has on splicing patterns is influenced by cis signals, sequencing experiments were performed (RNA-Seq) in PRMT5 wildtype and mutant Arabi-dopsis thaliana plants belonging to two different ecotypes: Columbia (Col-0) and Landsberg erecta (Ler). In this way, we sought to analyze the effect that the mutation has on the genetic variability that these ecotypes have. In order to discriminate the changes related to variations in the cis sequences, F1 Col-0 X Ler and Ler X Col-0 hybrids were also analyzed. Through these analyses, it was concluded that an important part of the splicing patterns affected by the PRMT5 mutation depend on cis signals, having a particular relevance the strength of the donor site. Fil: Beckel, Maximiliano Sebastián. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

[ES] La detección de variantes de ADN es una tarea importante y difícil en la Genómica. Las tecnologías de secuenciación de una sola molécula han conseguido revolucionar la detección de variantes genéticas, el ensamblaje de genomas complejos y la detección de marcas epigenéticas (marcas químicas que se añaden al ADN y permiten su correcto funcionamiento) en los últimos años. En el caso de la secuenciación de una sola molécula, se tiene una tasa de error por nucleótido de entre 5 y 15%. Por lo que a pesar de los rápidos avances en las tecnologías de secuenciación, es difícil detectar variantes del genoma de un individuo a partir de miles de millones de lecturas de secuencias cortas y erróneas. Debido a la masiva cantidad de lecturas del genoma y la alta tasa de error de secuenciación, aplicar Deep Learning para el aprendizaje de relaciones entre las secuencias de ADN, es el enfoque que mejores resultados obtiene, superando a las herramientas de última generación y métodos tradicionales de detección de variantes. La naturaleza de la tarea que se propone enmarca este Trabajo Final de Máster dentro de la Genómica, concretamente en el problema de Detección de Variantes Genómicas aplicando Técnicas de Inteligencia Artificial y del Reconocimiento de Formas. Como primer objetivo se realiza un estudio de las diferentes herramientas de última generación, enfoques y particularidades. El segundo objetivo se centra en el tratamiento y manipulación de los datos genómicos con el fin de desarrollar una metodología robusta. En la metodología se incluye el análisis y diseño de arquitecturas de Redes Neuronales Artificiales para el problema que se trata y sus correspondientes experimentos. Finalmente, se discuten los resultados obtenidos y se declaran futuros desarrollos para continuar nuevas investigaciones en esta línea., [EN] DNA variant detection is an important and challenging task in Genomics. Single-molecule sequencing technologies have revolutionized the detection of genetic variants, the assembly of complex genomes and the detection of epigenetic marks (chemical marks that are added to DNA and allow it to function correctly) in recent years. In the case of single molecule sequencing, there is a per nucleotide error rate of between 5 and 15%. So despite rapid advances in sequencing technologies, it is difficult to detect variants of an individual's genome from billions of short and erroneous sequence reads. Due to the massive amount of genome reads and the high sequencing error rate, applying Deep Learning for learning relationships between DNA sequences is the approach that yields the best results, outperforming state-of-the-art tools and traditional variant detection methods. The nature of the proposed task frames this Master's Thesis within Genomics, specifically in the problem of Genomic Variant Calling by applying Artificial Intelligence and Shape Recognition Techniques. The first objective is a study of the different state-of-the-art tools, approaches and particularities. The second objective focuses on the treatment and manipulation of genomic data in order to develop a robust methodology. The methodology includes the analysis and design of Artificial Neural Network architectures for the problem at hand and their corresponding experiments. Finally, the results obtained are discussed and future developments are stated in order to continue new research in this line.

Over the last decade, molecular systematics has undergone a change of paradigm as high-throughput sequencing now makes it possible to reconstruct evolutionary relationships using genome-scale datasets. The advent of “big data” molecular phylogenetics provided a battery of new tools for biologists but simultaneously brought new methodological challenges. The increase in analytical complexity comes at the price of highly specific training in computational biology and molecular phylogenetics, resulting very often in a polarized accumulation of knowledge (technical on one side and biological on the other). Interpreting the robustness of genome-scale phylogenetic studies is not straightforward, particularly as new methodological developments have consistently shown that the general belief of “more genes, more robustness” often does not apply, and because there is a range of systematic errors that plague phylogenomic investigations. This is particularly problematic because phylogenomic studies are highly heterogeneous in their methodology, and best practices are often not clearly defined. The main aim of this article is to present what I consider as the ten most important points to take into consideration when planning a well-thought-out phylogenomic study and while evaluating the quality of published papers. The goal is to provide a practical step-by-step guide that can be easily followed by nonexperts and phylogenomic novices in order to assess the technical robustness of phylogenomic studies or improve the experimental design of a project., J.L.-F. was supported by a Juan de la Cierva Incorporación fellowship (Ministerio de Ciencia e Innovación, IJC2018-035237-I).