Tesis Doctorals - Departament - Genètica, Microbiologia i Estadística

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Role of the trachea in the formation of the abdominal epidermal layer during metamorphosis of Drosophila melanogaster
(Universitat de Barcelona, 2026-01-14) Bisai, Bitarka; Casanova i Roca, Jordi, 1959-; Franch i Marro, Xavier; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] A central challenge in developmental biology is understanding how distinct tissues coordinate their growth and remodeling to produce a functional adult body. This requires the precise integration of systemic hormonal signals with local tissue specific communication, ensuring both temporal precision and spatial alignment. This coordination is particularly critical during insect metamorphosis characterized by a paradigm of extensive tissue reorganization, where larval structures are eliminated and replaced by adult-specific structures. The Drosophila abdomen during metamorphosis provides a powerful model to study such coordination, as histoblasts, muscles, and the tracheal network must remodel within a narrow developmental window to generate a functional adult abdomen. The steroid hormone 20-hydroxyecdysone (20E) is the canonical trigger of developmental transitions, orchestrating many aspects of metamorphosis and pupal development, yet the mechanisms by which local interactions between tissues refine this timing and spatial patterning remain poorly understood. Here, I identify the pupal tracheal system as a dual-function signaling hub that coordinates abdominal morphogenesis through both local endocrine output and paracrine signaling. Using live imaging, genetic knockdown, and tissue-specific manipulations, I show that pupal tracheal remodeling proceeds in two phases, the larval branch degradation followed by adult branch outgrowth, and that it is critically dependent on the temporal transcription factor Broad (Br). Disruption of this remodeling arrests histoblast proliferation and generates a fully penetrant “naked abdomen” phenotype marked by failed epithelial replacement, revealing a unidirectional dependency of epidermal morphogenesis on tracheal signaling. Mechanistically, the pupal trachea expresses Halloween biosynthetic genes and contributes to the pupal 20E pool, a function shared with dorsal internal oblique muscles (DIOMs). Loss of either tissue’s steroidogenic activity yields indistinguishable epidermal defects, highlighting functional interdependence and coordination between local endocrine sources. In parallel, tracheal-derived Pvf1 acts through its receptor Pvr in histoblasts and muscles to coordinate abdominal epidermal patterning and morphogenesis. This establishes a paracrine signaling axis essential for synchronizing epidermal expansion with abdominal remodeling. Together, these findings redefine the trachea from a passive respiratory scaffold to an active developmental coordinator. By integrating systemic hormonal cues with local endocrine and paracrine outputs, the metamorphic trachea synchronizes multi-tissue remodeling in the abdomen, offering a new conceptual framework for inter-organ communication during morphogenesis and highlighting broader principles in developmental biology.
Genomics for conservation: Insights into the ongoing range expansion of sea turtles in the Mediterranean
(Universitat de Barcelona, 2026-01-16) Marín Capuz, Gisela; Carreras Huergo, Carlos; Pegueroles Queralt, Cinta; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Climate change is reshaping species distributions, with sea turtles being particularly vulnerable due to their unique biological and ecological traits. Understanding the origin, mechanisms, and potential future of these changes is essential for predicting species responses and for designing science-based management strategies. This thesis investigates the recent nesting range expansion of green (Chelonia mydas) and loggerhead sea turtles (Caretta caretta) in the central and western Mediterranean, using high-throughput sequencing, satellite telemetry, stable isotopes and long-term field monitoring data. In Chapter 1, we validated the use of umbilical cords (the remaining skin of the yolk sac) as a non-invasive and reliable source of DNA. Genomic analysis confirmed that their genetic profile consistently matches that of the hatchling and not the mother. These tissues are more frequent in artificially incubated nests, a widely used management strategy in emerging sea turtles nesting sites. This method provides an ethical and effective tool for conservation genomics. Chapter 2 focuses on the incipient nesting range expansion of the green turtle. We first developed a genomic baseline to assign individuals to their Regional Management Unit (RMU) of origin. Later, by analysing emerging nests and newly observed adults in the western Mediterranean, we detected simultaneous incipient range expansion from both the South Atlantic and Mediterranean RMUs. These results suggest a climate-driven expansion from multiple regions, in line with the pat-tern already observed in loggerhead turtles. Chapter 3 focuses on the colonisation process of loggerhead turtle in Spain. The long-term analysis of 69 nests laid between 2001 and 2023 showed emerging site fidelity and remigrant females, indicating the establishment of a new nesting colony. Genomic assignments linked two nests to the Northwest Atlantic RMU, six to admixed origins, and the rest to the eastern Mediterranean, mainly Greece and the Levant. We estimated a minimum effective breeding population of 39 females and 47 males and identified genetic and ecological factors affecting reproductive success. Finally, we described the colonisation steps followed by sea turtles, supporting the emergence of a new resident population in the western Mediterranean, likely adapting philopatric behaviour to climate change. We integrate all findings to show how they can inform management plans and support conservation. We recommend standardised protocols and long-term monitoring as essential for effective conservation.
From Multimorbidity to Disease Trajectories: Genetic and Epidemiological Perspectives in Population-Based Cohorts
(Universitat de Barcelona, 2026-01-19) Blay Magrina, Natàlia; Cid Ibeas, Rafael F. de; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Chronic conditions are the leading cause of morbidity and mortality worldwide, often co-occurring as multimorbidity and impacting on individual’s quality of life and healthcare systems. Beyond isolated disease analysis, the study of temporal sequence of diseases, referred to as disease trajectories, may reveal underlying mechanisms and influence disease progression. This doctoral thesis assesses the health effects of chronic conditions and disease trajectories in population-based cohorts, and investigates their genetic and environmental determinants, as well as their possible interaction. First, we assessed the representativeness of the GCAT cohort (n ≈ 20,000) with the general population of Catalonia. We identified the presence of a healthy volunteer bias characterised by a higher proportion of women, higher education and healthier individuals. To mitigate this bias, we developed raked weights based on sociodemographic, lifestyle and healthcare variables, reducing the difference of compared variables by 70% and by 26% in disease prevalence, improving the cohort’s utility for population-level inference. Second, we explored disease trajectories in GCAT, extrapolated them into the general population and validated the finding in UK Biobank (n ≈ 500,000). We identified 300 enriched disease trajectories, many with direction-specific prevalence and mortality rate differences. For example, hypertension → chronic kidney disease was six times more prevalent than the reverse trajectory, or hypertension → chronic obstructive pulmonary disease was three times more lethal than the reverse trajectory. Genetic analysis of 21 disease trajectories in UK Biobank revealed trajectory-specific loci not detected in single-disease GWAS, suggesting the presence of specific underlying mechanisms. Polygenic risk scores derived from GWAS were able to predict disease trajectories risk in GCAT, supporting a genetic component in disease progression. Third, we applied this methodology to long COVID using the COVICAT cohort (n = 8,322). We found that 38 pre-pandemic disease trajectories significantly increased long COVID risk, particularly those involving mental health and neuromuscular disorders, cardiometabolic and digestive diseases, and respiratory conditions. While the genetic correlation between identified diseases and long COVID was low, PRS for headache syndromes and intervertebral disc disorders were associated to long COVID phenotype, suggesting indirect genetic influences. These findings underscore the role of multimorbidity and disease temporality in long COVID susceptibility. Finally, we examined gene-environment interactions in depression in the COVICAT cohort during COVID-19 lockdown. Using poly-environmental risk scores and polygenic risk scores, we observed that social and lifestyle stressors were the strongest predictors of depression severity, and genetic effects showed modest contribution to broad depression phenotypes. Notably, an interaction between environmental stressors and genetic predisposition suggested a potential mitigating effect, highlighting the complexity of mental health vulnerability. In conclusion, this thesis illustrates the complexity of chronic diseases determinants and demonstrates that disease trajectories provide clinically relevant insights beyond traditional multimorbidity approaches, revealing direction-specific risks, different mortality patterns and specific genetic underpinnings. The incorporation of integrative models based on disease trajectories, together with genetic data and environmental factors can improve personalised risk prediction, prevention and management of chronic and emerging conditions such as long COVID.
Cancer type specificity and temporal selection patterns of driver genes from large-scale panel sequencing data
(Universitat de Barcelona, 2026-01-20) Colic, Daniel; Supek, Fran; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Progress in cancer genomics, driven by the accessibility of next-generation sequencing as well as the production and analysis of large-scale cancer patient genomic and clinical data, has enabled identification of hundreds of driver genes. This knowledge has been used to gain insights into cancer biology, and in clinical practice to predict patient survival and adequate treatments. However, our understanding of cancer drivers' role across tissues and cancer types, their preferred timing, and their interaction with other drivers remains incomplete. This thesis addresses these fundamental questions by developing and applying novel statistical methods to analyze the selection pressures shaping tumor evolution. We aim to quantify selection pressure on cancer genes across a wide range of cancer types. Next, we aim to unravel the timing of mutations by integrating selection analysis with mutation clonality. Finally, we want to map the epistatic interactions between early and late driver events to understand the cooperative mechanisms that shape cancer evolution. We developed a method to quantify selection pressure by measuring the excess of nonsynonymous mutations compared to effectively synonymous mutations within gene exonic regions, accounting for trinucleotide context and mutational exposure. Applying this approach to 60,000 samples from MSK and DFCI panel sequencing data, we identified distinct selection patterns across cancer types. Beyond the established dichotomy of cancer-specific and universal drivers, we discovered moderate selection signals in several previously considered cancer type-restricted genes, such as KMT2C, MAP2K1, and TET2. These genes can be misclassified in smaller cohorts using methods that separately quantify selection in each cancer type, thereby missing the signal of consistent moderate selection and underestimating rare drivers. Our analysis, supported by recent independent studies, suggests broader oncogenic roles for already annotated cancer driver genes. We then integrated clonality inference with selection analysis, performing pooled analyses grouped by cancer types and genes. Our temporal selection analysis revealed differences between cancer types, broadly grouped into two categories. Cancer types including lung cancers, melanoma, and pancreatic cancer showed strong clonal selection bias, emphasizing the role of early driver events. Conversely, sarcoma, thyroid, and hepatobiliary cancers exhibited continuous selection pressure in the subclonal phase, suggesting adaptation in advanced cancer stages, metastasis, or treatment resistance. At the gene level, we defined a spectrum ranging from preferentially early drivers to time-invariant genes. These findings were validated in whole exome data from The Cancer Genome Atlas. Early genes such as TP53, PTEN, and FBXW7 exhibited decreased selection in the subclonal phase, unlike ERBB2, SETD2, and PIK3CA, which showed equivalent or stronger selection in subclonal phases. Understanding mutation clonality proved valuable for deciphering the temporal dynamics of cancer evolution as an inherently time-dependent process, defining some cancer genes as crucial for initiation and others as accelerators of proliferation in advanced tumors. Finally, we examined the genetic interaction landscape of early and time-invariant genes defined in our previous analysis. We observed differences between these gene classes: time-invariant genes showed a balanced distribution of both positive and negative epistatic interactions, while early genes exhibited predominantly mutually exclusive relationships with other genes, suggesting their distinct roles in cancer evolution. PIK3CA, a frequent and universal oncogene, formed synergistic interactions with PTEN across three cancer types. We propose this mechanism explains late selection pressure, as it depends on the specific genetic background of tumors. Conversely, TP53 showed antagonistic interactions with ATM across five cancer types, exemplifying functional redundancy within the same pathway, the role of early genes in defining evolutionary constraints, and potential therapeutic opportunities. This work establishes a computationally efficient framework for analyzing sparse panel sequencing data and challenges the traditional binary classification of cancer drivers by revealing that cancer evolution is shaped by a spectrum of selection strengths, temporal dynamics, and epistatic interactions that determine how tumors evolve and adapt over time.
Cellular and Molecular Insights into Schaaf-Yang Syndrome: A Multi-Model Analysis from Fibroblasts to iPSC-Derived Neural Cell Types
(Universitat de Barcelona, 2026-01-22) Centeno-Pla, Mónica; Rabionet Janssen, Raquel; Serrano Gimaré, Mercedes; Urreizti, Roser; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Schaaf-Yang syndrome (SYS) is an ultra-rare neurodevelopmental disorder caused by truncating mutations in the maternally imprinted gene MAGEL2. Clinically, SYS shares many features with Prader-Willi syndrome (PWS), such as neonatal hypotonia, feeding difficulties, endocrine dysfunction, neurodevelopmental delay and a consequent intellectual disability, but also presents with distinctive traits including joint contractures and a higher degree of intellectual difficulties. Due to its rarity, SYS remains poorly understood, with limited consensus on management, and molecular mechanisms. This thesis addresses these challenges through an integrated clinical and molecular approach. In this thesis, phenotypic and genetic reviews of all reported SYS cases have been performed, enabling the development of the first clinical guidelines for patient management. A curated catalogue of MAGEL2 variants has been established, resolving misannotations and highlighting the diagnostic importance of parental origin assessment in this imprinted gene. To elucidate disease mechanisms, we have studied the behaviour of truncated MAGEL2 proteins using heterologous expression systems, patient-derived fibroblasts, and induced pluripotent stem cell (iPSC)-based neural models. Our findings show that truncating mutations produce stable proteins that mislocalise to the nucleus rather than being degraded. This aberrant localisation potentially reframes SYS as a dual loss- and gain-of-function disorder distinct from PWS. Multi-omic studies across different patient-derived cell models have shown both overlapping and cell type-specific dysregulations. Specifically, in CNS cell types, transcriptomic profiling has identified disruptions in synaptic and neuronal function, extracellular matrix organisation, and developmental pathways. Preliminary calcium imaging assays have potentially uncovered impaired GABAergic neuronal activity, linking transcriptional dysregulation to altered physiology. Finally, we have observed alterations in amyloid-β secretion across patient-derived cell types, establishing its potential as a robust, disease-relevant biomarker. This work is among the first to use human iPSC-derived neuronal models to investigate SYS, providing direct access to potentially disease-relevant cell types. Using multiple patient-derived platforms, we refine the molecular definition of SYS and reveal potential pathogenic mechanisms alongside candidate biomarkers. By integrating clinical, multi-omic, and functional analyses, this thesis advances our understanding of SYS pathophysiology and establishes a foundation for future studies aimed at dissecting molecular mechanisms and identifying therapeutic targets.
Evolutionary Genomics using long-read sequencing: A case study in Dysdera spiders
(Universitat de Barcelona, 2026-01-30) Pisarenco, Vadim; Rozas Liras, Julio A.; Guirao Rico, Sara; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Sequencing technologies have undoubtedly revolutionised the life sciences within just a few decades, transforming the tedious effort of sequencing a single gene into the routine generation of entire genomes. This rapid progress has been accompanied by the steady development of bioinformatic methods and tools needed to process and analyse these vast amounts of data. The significance of this progress for evolutionary biology is indisputable: it has established connections between distant biological fields and provided unprecedented opportunities to test population genetics’ theoretical predictions. The impact is far-reaching; evolutionary studies have moved beyond purely descriptive narratives and now integrate multidisciplinary approaches to explore in depth the patterns and processes underlying biological phenomena and to propose explanatory mechanisms. Grounded within the framework of the neutral theory of molecular evolution and comparative genomics, this thesis aims to advance our understanding of the evolution of genome size and chemoreceptor gene families, using spiders of the genus Dysdera as a model system. We generated three high-quality, chromosome-level genome assemblies, one from a mainland species (Dysdera catalonica) and two for Canary Island endemics (D. silvatica and D. tilosensis), and provided a structural and functional annotation of protein-coding genes and repetitive elements. These assemblies represent the first chromosome-level genomic resources for the Synspermiata spider clade, encompassing 17 spider families, and thus constitute a valuable resource for the scientific community. Using these assemblies, we performed a comprehensive comparative genomic analysis to investigate the genome size differences between mainland and insular species. We uncovered an unexpected case of genome size reduction in insular species, driven by large deletions that primarily affected the repetitive fraction of the genome as well as genetic elements under low functional constraint. Our results further suggest that a large effective population size in insular species enhanced purifying selection against slightly deleterious mutations, such as transposable element insertions. Overall, these findings support the non-adaptive mutation hazardous hypothesis of genome size evolution. We also developed GALEON, a bioinformatic tool designed to assist in the identification, analysis, and visualisation of gene clusters. This software facilitates the study of gene families by enabling exploration of the relationship between physical and evolutionary distances. Additionally, we extended its application to explore putative co-evolution between gene families. Finally, we conducted a comprehensive characterisation of the main arthropod chemoreceptor gene families (gustatory (Gr) and ionotropic (Ir) receptors) in six Dysdera species, employing chromosome-level assemblies and reference-guided scaffolding data. We found that the continental species D. catalonica encodes the largest number of Ir genes reported among chelicerates, while the size of the Gr family falls within the range documented for other chelicerates. We showed that the chemosensory gene families are organised into genomic clusters; however, the genome of D. catalonica harbours larger clusters containing more genes than those of the insular species. Our analysis suggests that unequal crossing-over is the primary mechanism driving gene duplication, supporting the birth-and-death model of gene family evolution. Intriguingly, the X chromosome is consistently depleted of chemoreceptors compared to the autosomes. Our study also shows that a substantial fraction of gene family members remain on small scaffolds and thus, could not be assigned to chromosomes, highlighting the persistent challenges of analysing large, repetitive gene families even with high-quality genomes.
Aproximaciones genómicas y estudios funcionales de variantes genéticas asociadas con trastornos esqueléticos con baja y alta masa ósea
(Universitat de Barcelona, 2026-01-30) Patiño-Salazar, Juan David; Rabionet Janssen, Raquel; Balcells Comas, Susana; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[spa] Las displasias esqueléticas (DiE), son un grupo heterogéneo de enfermedades con una incidencia poco frecuente y diversas manifestaciones clínicas, que están asociadas a múltiples genes y que pueden presentar fenotipos de alta o baja masa ósea. El diagnóstico preciso de las DiE es difícil desde el punto de vista clínico y genético. Los avances de los últimos años en técnicas de secuenciación y procesamiento de datos así como los estudios GWAS, nos han dado luces sobre las bases genéticas que pueden estar regulando la fisiología ósea en contextos patológicos como el de las DiE. Sin embargo, la información obtenida mediante NGS o GWAS, debe ser validada funcionalmente. En esta tesis doctoral utilizamos aproximaciones genómicas mediante WES y panel de genes para analizar las bases genéticas que subyacen a trastornos esqueléticos con baja masa ósea como la osteoporosis posmenopáusica (OP), o casos de displasia esquelética familiar; y estudios funcionales in silico e in vitro para evaluar las implicaciones de variantes genéticas raras asociadas a trastornos esqueléticos con alta masa ósea y de otras patologías con fenotipo óseo como la enfermedad de Gaucher tipo 1 (EG1). En cuanto a los análisis de trastornos esqueléticos con baja masa ósea, el estudio realizado mediante panel de genes en una selección truncada de casos de la cohorte BARCOS con valores de DMO extremos, nos permitió identificar variantes en 6 genes (TCIRG1, COL1A2, SEC24D, LRP4, ANO5 y LMNA), una de ellas patogénica en heterocigosis en el gen COL1A2 p.(Gly751Ser). Además, mediante análisis WES identificamos una variante missense p.(Arg296Pro) en el gen PORCN en un paciente de 29 años con una displasia esquelética sin diagnosticar, asociada con LBM y fracturas por fragilidad en su infancia y adolescencia. Análisis adicionales a partir de la DMO de este paciente y revisión posterior de literatura, nos llevaron a sugerir que la hipoplasia dérmica focal (FDH), causada principalmente por variantes genéticas en este gen, es probablemente una causa monogénica poco reconocida de LBM debida a una señalización defectuosa de la vía de Wnt. Respecto de los trastornos esqueléticos con alta masa ósea, estudiamos funcionalmente las variantes en SEM4AD, TBX18, PTCH1, PTK7, y ADGRE5 identificadas previamente mediante WES, en un caso familiar de displasia ósea esclerosante caracterizado por lesiones óseas lucentes y HBM, logrando determinar que ninguna de las variantes afectó la estructura conformacional de la proteína ni comprometió la estabilidad proteica. Además, los estudios in vitro de la variante p.Arg349Gln en TBX18 nos permitieron observar que no afectaba su actividad transcripcional, y los ensayos de diferenciación de osteoblastos realizados para validar funcionalmente la variante p.Val212Met en SEMA4D no nos permitieron obtener resultados concluyentes. Finalmente, la generación y caracterización de modelos celulares osteoblástico y osteoclástico para estudiar el fenotipo óseo de la EG1, nos permitió sugerir que quizá haya un defecto osteoclástico intrínseco en la EG1, lo cual podría implicar que los osteoclastos serían los principales impulsores de la patología ósea, junto con la alteración de la comunicación entre osteoblastos y osteoclastos.
El mostreig passiu per a la detecció i caracterització de virus en aigües: aplicacions en vigilància epidemiològica i gestió de recursos hídrics
(Universitat de Barcelona, 2026-06-26) Mejías Molina, Cristina; Bofill Mas, Silvia; Rusiñol Arantegui, Marta; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[cat] Molts dels virus que infecten humans són excretats al medi a través de la femta o de l’orina, independentment que l’individu infectat presenti o no símptomes. L’estudi dels virus presents en l’aigua residual permet, per tant, conèixer la circulació d’aquests dins d’una comunitat concreta. En aquesta tesi s’ha avaluat un sistema de mostreig passiu per a la detecció i caracterització per metagenòmica de virus en aigua residual amb l’objectiu principal d’aplicar-lo en contextos poblacionals petits i/o de difícil accés amb als sistemes tradicionals de mostreig, com ara edificis. Un cop validat com a mètode sensible per a la caracterització dels virus excretats, s’ha utilitzat per analitzar el viroma d’aigües residuals provinents d’edificis amb perfils demogràfics diferenciats, amb la finalitat d’identificar les diferències existents entre aquests grups poblacionals. Els virus presents a les aigües residuals poden convertir-se en una font de contaminació del medi aquàtic a través d’abocaments d’aigua residual no tractada derivats de fuites de clavegueram o de fosses sèptiques, però també a través dels efluents de les estacions depuradores d’aigua residual (EDARs) que tot i els diferents passos de tractament, alliberen contínuament virus al medi per mitjà dels efluents. Paral·lelament, el context actual marcat per una creixent escassetat de recursos hídrics fa necessari avançar cap a una gestió més eficient i sostenible dels recursos, això implica tenir en compte l’estat de fonts d’aigua potencialment contaminades i de l’aigua regenerada potencialment reutilitzada. Per aquest motiu, en aquesta tesi s’ha aplicat per primera vegada el mostreig passiu per al monitoratge de virus en aigües regenerades i aigües subterrànies, entorns on els virus es troben habitualment en concentracions molt baixes. Els resultats obtinguts a l’aqüífer demostren que aquesta metodologia és una alternativa viable per detectar contaminació fecal d’origen humà i millorar la gestió d’aquesta font d’aigua mitjançant la detecció de patògens vírics La seqüenciació massiva ha permès per primera vegada en aquest compartiment la descripció del viroma de l’aigua subterrània. Per altra banda, l’ús de mostrejadors passius per al monitoratge de l’aigua regenerada, produïda en sistemes basats en la natura com els aiguamolls construïts, fa possible monitoritzar de manera senzilla la presència de patògens víric d’importància per a la salut pública. En conjunt, els resultats d’aquesta tesi posen de manifest que el mostreig passiu és una metodologia versàtil per a la detecció i caracterització de virus en aigües amb diferents graus de contaminació i que amplia la capacitat de vigilància en aquells entorns on les tècniques convencionals presenten limitacions o són difícils d’implementar.
Development of an iPSC-based model system for exploring the biological impact of NF1, CDKN2A and SUZ12 loss on malignant peripheral nerve sheath tumor initiation and identification of new therapeutic strategies
(Universitat de Barcelona, 2025-12-19) Uriarte Arrazola, Itziar; Serra Arenas, Eduard; Carrió i Llach, Meritxell; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive soc tissue sarcomas arising from peripheral nerve sheath cells. Nearly half of all cases are associated with Neuroﬁbromatosis type 1 (NF1), a hereditary syndrome that predisposes to the development of benign and malignant tumors. MPNSTs account for 3–10% of all soc tissue sarcomas, and NF1 patients face a 10–15% lifetime risk of developing them, making MPNSTs the leading cause of mortality in this collective. No eﬀective therapies exist besides a timely surgery. In the context of NF1, MPNSTs commonly initiates through the stepwise inactivation of diﬀerent tumor suppressor genes (TSGs). Benign plexiform neuroﬁbromas (PNFs) arise in large nerves from neural crest (NC)–derived Schwann cell precursors (SCP) following NF1 loss. Subsequent inactivation of the CDKN2A locus gives rise to pre-malignant nodular lesions termed atypical neuroﬁbromatous neoplasms of uncertain biological potential (ANNUBPs). Progression towards an MPNST involves the disruption of the chromatin remodeler polycomb repressive complex 2 (PRC2) through SUZ12 or EED complete inactivation. Acer TSG loss, MPNSTs undertake an extensive genomic reorganization, including copy number alterations and structural variants. This PhD thesis aimed to investigate the biological consequences of the inactivation of NF1-CDKN2A-SUZ12 key TSGs during the PNF-ANNUBP-MPNST progression. It intended (i) to elucidate the mechanistic insights of this progression by the generation and characterization of induced pluripotent stem cell (iPSC)-based models of each tumor progression step; and (ii) to used the developed models to identify new potential therapeutic epigenetic compounds for MPNSTs. We successfully developed diﬀerent iPSC-derived NC models through stepwise inactivation of NF1, CDKN2A and SUZ12 using gene editing technologies. These models faithfully capture the progression from benign to malignant peripheral nerve sheath tumors. NF1–CDKN2A double-knockout (2KO) neural crest cells (NCs) retained glial diﬀerentiation capacity and generated neuroﬁbroma-like tumors in vivo, requiring the inactivation of both p14ARF and p16INK4a. NF1–CDKN2A-PRC2 triple-knockout (3KO) NCs demonstrated that the TSG inactivation order driving PNF-MPNST progression is biologically constrained. We conﬁrmed that PRC2 function is essential for maintaining pluripotency. iPSCs without PRC2 function were stably maintained as NCs, acquiring mesenchymal stem cell–like properties and extensively remodeling chromatin accessibility, with profound eﬀects on cell identity. We observed that 3KO NC cells cannot diﬀerentiate into SCs due to SOX10 epigenetic silencing, while they re-express developmental neurogenesis programs, due to chromatin opening. Part of these genes were poised and others required factors only present in MPNST cells. Thus, 3KO NCs represent a genuine early-stage MPNST-like tumor model, since they capture the glial-to-mesenchymal transition observed in the PNF-MPNST progression and form MPNST-like tumors when engraced in the sciatic nerve of nude mice. Since PRC2 loss completely modify chromatin accessibility, we next used 3KO NC spheroids to perform high-throughput screening (HTS) of an epigenetic compound library for the identiﬁcation of new potential MPNST drugs. We identiﬁed poly(ADP-ribose) polymerase inhibitors (PARPi) as a novel class of compounds selectively active in PRC2-deﬁcient models. Combination assays revealed modest synergy between Selumetinib (MEKi) and HDAC, BET, or PARP inhibitors. We tested those combinations in 2D MPNST cell lines, observing nearly identical viability readouts for the BETi and HDACi combinations, thereby validating our system. In contrast, PARPi did not reduce viability in 2D MPNST cell lines. Importantly, in a patient-derived orthotopic xenograc (PDOX) mouse model, the Olaparib–Selumetinib (PARPi-MEKi) combination was well tolerated and signiﬁcantly inhibited MPNST growth, corroborating the therapeutic potential identiﬁed in our 3D model. In conclusion, we have demonstrated that our iPSC-based NC system functions both as a robust platorm for studying PNF-MPNST progression, and as a scalable and reliable 3D preclinical tool for drug discovery.
RNA-editing: an evolutionary perspective through bioinformatics and machine learning.
(Universitat de Barcelona, 2025-11-11) Zawisza Alvarez, Mikel; Garcia Fernández, Jordi; Herrera Úbeda, Carlos; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] RNA-editing is a post-transcriptional modification of the sequence of an RNA molecule at the level of particular nucleotides, in contrast to the more widely known and studied process of splicing which acts by removing larger regions of the sequence. Adenosine to inosine (A-to-I) editing by the ADAR (Adenosine deaminase acting on RNA) family of proteins is the most prominent type of RNA-editing in metazoans, with a wide range of biological consequences, including notable functions related to the immune and nervous systems. While it is known that the secondary structure of the RNA plays an important role in the decision to edit a particular adenosine, a deep understanding of ADAR’s targeting mechanism is still elusive. Differences in the regulation of RNA-editing between mammals and birds poses questions about the evolutionary relevance of this mechanism. In this thesis I attempt to tackle the technical difficulties of RNA-editing detection with the intention of studying the process from an evolutionary perspective, while also advancing the knowledge of ADAR’s targeting mechanism. To achieve this, I developed various novel bioinformatics methods and tools, including orthology-based and machine-learning based approaches, with the highlight being a method we call cross-testing which we propose to study the functional conservation of ADAR’s targeting mechanism in silico, beyond the sequence conservation of the functional domains of the proteins.
Tailing Free: The Evolution of Tail Muscle in Oikopleura dioica Sheds Light on the Transition to a Fully Free-Swimming Lifestyle in Appendicularian Tunicates
(2025-11-13) Fabregà Torrus, Marc; Cañestro García, Cristian; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Tunicates constitute a large and diverse group of marine invertebrates encompassing a wide range of life strategies, from the fully free-swimming lifestyle of appendicularians to the biphasic lifestyle of ascidians, which are characterized by motile larvae and sessile adults. However, whether the ancestral tunicate resembled a biphasic, ascidian-like form or a fully free-swimming appendicularian remains an open evolutionary question. The free-swimming appendicularian lifestyle is tightly associated with the evolutionary innovation of the filtering house and the retention and specialization of a functional tail, which is maintained throughout the entire life cycle. This tail enables complex swimming behaviours, in contrast to the simpler dispersal function of the ascidian larval tail, which is lost during metamorphosis. To better understand how the evolution of the tail facilitated the emergence of the appendicularian lifestyle, in this PhD project, we investigate tail muscle development in the appendicularian Oikopleura dioica through embryonic lineage tracing, gene expression analysis, and comparative genomic surveys of the myofibrillar gene catalogue. Our findings reveal a profound remodelling of tail muscle lineages compared to ascidians, along with a massive expansion and diversification of cardioparaxial sarcomeric Myh genes (Myh-Scp), which exhibit temporally and spatially specific expression patterns. This diversification of myofibrillar genes suggest and increased diversity of muscle cell identities with distinct contractile properties along the anteroposterior axis of the tail, enabling the complex repertoire of movements observed in appendicularians. Finally, our results allow us to propose a new evolutionary model for the transition in lifestyle among tunicates, based on the evolution of the tail. This model, named “tailing free”, emphasizes the central role of the tail in enabling a transition from a biphasic ascidian-like ancestor to a fully free-swimming appendicularian lifestyle. Furthermore, it supports evidence of our previous work suggesting that the ancestral tunicate exhibited a biphasic lifestyle with a sessile, post-metamorphic adult stage, similar to ascidians. Metaphorically, appendicularians “break free” from their ancestral ties to a sessile, substrate-attached life and adapt to a free open-water existence, ultimately becoming one of the most abundant and ecologically important components of the mesozooplankton in marine ecosystems.
Aneuploidy in neural stem cells: linking proteostasis failure and mitochondrial dysfunction to microcephaly
(Universitat de Barcelona, 2025-10-03) González Blanco, Amanda; Milán, Marco; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] The precise acquisition of chromosome number is fundamental for proper brain development and function. Alterations in this process result in aneuploidy, defined as an abnormal chromosome number. Aneuploidy has profound detrimental effects at both the cellular and organismal levels. Several human conditions, such as Down syndrome and Mosaic Variegated Aneuploidy (MVA), are caused by aneuploidy and are associated with severe health problems, such as cognitive impairment, developmental delay and microcephaly. Unfortunately, therapeutic strategies for aneuploidy-related conditions are still lacking, highlighting the need for further investigation into the molecular mechanisms underlying these disorders. In this thesis, we investigated the cellular and molecular consequences of aneuploidy in the context of brain development. We used Drosophila neural stem cells (neuroblasts, NBs) as a robust model system to study both simple aneuploidies (trisomies) and chromosomal instability (CIN)-induced aneuploidy. We determined that just by downregulating one spindle assembly checkpoint gene, as observed in MVA human patients, we were able to recapitulate the microcephaly phenotype. Our findings revealed that CIN-induced aneuploidy caused a progressive decrease in the number of NBs and their proliferative capacity, as well as in the number of progeny cells, consequently resulting in smaller brains compared to controls. We explored the underlying mechanisms contributing to this impairment, distinguishing between the consequences of DNA damage and aneuploidy in the cells. We dissected the differences between complex (CIN) and simple aneuploidies (trisomies) and demonstrated that increasing the degree and complexity of aneuploidy correlates with a more detrimental outcome for NBs. Furthermore, we deeply characterized the cellular process by which NBs are lost from the tissue upon CIN. We assessed a mild contribution of apoptosis to the resulting microcephaly and discarded other types of cell death. Conversely, we propose that NBs undergo a novel mechanism of loss of stem cell identity, becoming cells that are neither stem cells nor fully differentiated. These cells lose typical NB markers, making them undetectable in the tissue, become non-functional, and no longer contribute to the generation of nervous system cells, ultimately leading to microcephaly. Finally, we detected several cellular stresses in the NBs that arise as a consequence of CIN, such as protein accumulation, proteasome saturation, autophagy activation and saturation, and mitochondrial dysfunction. Importantly, we demonstrated that alleviating these cellular stresses in CIN NBs improves brain health, thereby opening promising avenues for the development of targeted therapeutic strategies aimed at enhancing the well-being of patients affected by aneuploidies.
Development of Experimental Methodologies for Modeling Bacterial Infections
(Universitat de Barcelona, 2025-10-03) Admella Pedrico, Joana; Torrents Serra, Eduard; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] The rise of antimicrobial resistance has made bacterial infections a global and signiﬁcant health threat. Additionally, infections associated with bioﬁlms are typically persistent and hinder the penetration and eﬃcacy of traditional antibiotics. Therefore, the development of new antimicrobial therapies relies on a comprehensive understanding of bacterial pathogenesis, supported by reliable laboratory models and methodologies. First, various in vitro lung cell models were developed and optimized to recreate bacterial lung infections. Within these systems, Pseudomonas aeruginosa pathogenesis, bioﬁlm formation, virulence, host-pathogen interactions, and the eﬃcacy of diﬀerent antibiotics were investigated. Given that respiratory infections are usually polymicrobial, coinfections involving P. aeruginosa and other clinically relevant species, such as Staphylococcus aureus and Candida albicans were also modeled. This research explored how microbial interactions shape both virulence and antibiotic susceptibility. On the other hand, several methodologies were optimized and established in the Galleria mellonella in vivo model. Among its many advantages, this insect shares signiﬁcant similarities with the human innate immune system, being widely employed in both infection and toxicity studies. In vivo bacterial pathogenesis was investigated by analyzing pathogen dissemination and monitoring the expression of key bacterial genes using a simple bioluminescence-based system. Furthermore, novel protocols, such as the primary culture of hemocytes, were developed for nanotoxicology evaluation. Both nanomedicine and phage therapy represent promising alternative therapeutic approaches and can be easily assessed in G. mellonella. In summary, this thesis presents the optimization of various in vitro and in vivo methodologies for modeling bacterial infections caused by clinically relevant pathogens, while providing diverse tools for the evaluation of novel antimicrobial therapies.
Characterization of alternative polyadenylation sites at single cell resolution and its impact on Alzheimer Disease
(Universitat de Barcelona, 2025-10-23) Ake, Franz Arnold; Plass Pórtulas, Mireya; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Alternative polyadenylation (APA) is a widespread mechanism of gene regulation that that affects more than 70% of human genes. mRNA isoforms generated though APA have distinct 3ʹ ends, which can affect mRNA regulation as well as the resulting proteins. APA is well known to be regulated during cell differentiation and it is a major source of gene regulation in the brain. Yet, it is not known till which extend APA contributes to the transcriptomic variability across individual cell populations. While bulk RNA sequencing has provided important insights into APA, it does not provide the resolution needed to resolve the cell-type-specific dynamics essential for understanding development and disease. The rapid development of 3’ tagged-based single-cell RNA sequencing (scRNA-seq) has enabled the study of gene expression and the implementation of methods for describing isoform usage at single cell resolution. However, challenges such as dropout effects or technical biases arising from the experimental setup have to be surmonted. In this thesis, we developed a novel computational approach to characterize isoform diversity at single-cell resolution using 3'-tagged scRNA-seq data, thereby enhancing the understanding of APA dynamics at single-cell resolution. First, I systematically evaluated the impact of various single-cell transcriptomics sample preservation methods on neural and glial cells derived from induced pluripotent stem cells. Secondly, I implemented and benchmarked SCALPEL, a novel computational Nextflow workflow designed for quantifying isoforms using standard 3′-tagged scRNA-seq data. Our results show that while DMSO provides the highest cell quality in terms of RNA molecules and genes detected per cell, it strongly affects the cellular composition and induces the expression of stress and apoptosis genes. In contrast, methanol fixed samples display a cellular composition similar to fresh samples and provide a good cell quality and little expression biases, indicating that methanol fixation is the method of choice for performing droplet-based single-cell transcriptomics experiments on neural cell populations. Furthermore, SCALPEL demonstrated high sensitivity and precision in quantifying isoforms, and the ability to detect differential isoform usage (DIU) in both synthetic and experimental datasets. In real datasets, SCALPEL predictions have a high agreement with other tools and can be experimentally validated. The use of SCALPEL on real datasets reveals novel cell populations undetectable using single-cell gene expression data, confirms known 3’ UTR length changes during cell differentiation, and identifies cell-type specific miRNA signatures regulating isoform expression. Additionally, we show that SCALPEL improves isoform quantification using paired long- and short-read scRNA-seq data. Overall, SCALPEL expands the current scRNA-seq toolkit to explore post-transcriptional gene regulation across species, tissues, and technologies, advancing our understanding of gene regulatory mechanisms at the single-cell level.
Fagos CrAssBcn: los nuevos virus crAssphage indicadores de contaminación fecal humana
(Universitat de Barcelona, 2025-10-31) Gómez Gómez, Clara; Muniesa Pérez, Ma. Teresa; García Aljaro, Cristina; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[spa] El estudio de la microbiota intestinal es esencial para comprender su impacto en la salud humana. Este ecosistema dinámico se estructura mediante complejas interacciones entre bacterias, bacteriófagos y otros microorganismos presentes en la microbiota. Los procesos evolutivos de las diferentes poblaciones y la acción de los fagos sobre las poblaciones bacterianas regulan su estabilidad y diversidad. En este contexto, el descubrimiento de crAssphage, posiblemente el fago más abundante en el intestino humano, ha revolucionado el conocimiento del viroma intestinal. El aislamiento de crAssphage mediante técnicas convencionales presenta desafíos significativos relacionados con su dificultad de propagación in vitro y la escasa visibilidad de las calvas de lisis que producen. A pesar de ello, disponer de crAssphage aislados es esencial para profundizar en su conocimiento. En la presente tesis doctoral, se ha logrado aislar 25 nuevos fagos tipo crAss en aguas residuales de Cataluña. Los fagos crAss aislados (crAssBcn) presentan una notable diversidad genética, posiblemente impulsada por la presión selectiva y la interacción con los sistemas de defensa de su hospedador bacteriano, Bacteroides intestinalis. La disponibilidad de viriones aislados ha permitido realizar ensayos experimentales más precisos sobre su ciclo de vida, superando así las limitaciones inherentes a los estudios realizados sólo con datos metagenómicos. Se observó la coexistencia de estos fagos virulentos con su hospedador bacteriano, cuyas poblaciones presentan fenotipos sensibles y resistentes a la infección. Los resultados sugieren que dicha resistencia, independiente del receptor del fago, está vinculada a una región genética en el hospedador B. intestinalis que está sujeta a variación de fase, la cual modula diferencialmente la expresión de polisacáridos capsulares (CPS) que, según se expresen o no, permiten la infección de los fagos. Los fagos del tipo crAss destacan como potenciales marcadores universales de contaminación fecal humana debido a su abundancia en heces, su ubicuidad y su resistencia a los tratamientos de desinfección. Sin embargo, buscando la selección de un candidato universal representativo de este grupo tan abundante y heterogéneo, los análisis de genomas completos, previamente anotados en bases de datos (tanto aislados como no), revelan la necesidad de considerar fagos tipo crAss específicos para ser aplicados en cada región geográfica. La ausencia de una relación filogenética entre ellos, su posible presencia en heces de origen animal y la inexistencia de un candidato con distribución global uniforme refuerzan la hipótesis de considerar variantes locales para su aplicación. Siguiendo esta premisa, se llevaron a cabo estudios experimentales más exhaustivos sobre la persistencia de los viriones crAssBcn autóctonos, siendo el primer estudio que ha evaluado la inactivación natural de fagos tipo crAss aislados. Los resultados confirman su mayor persistencia en comparación con otros indicadores microbiológicos e, incluso, con ciertos virus entéricos. En conjunto, los hallazgos obtenidos en esta tesis subrayan la necesidad de profundizar en la comprensión del papel ecológico de estos virus, con el propósito de desarrollar nuevas estrategias para el control ambiental, contribuyendo a mejorar la gestión de la calidad del agua, al tiempo que permitirá explorar nuevas aproximaciones diagnósticas y terapéuticas para abordar desequilibrios en la microbiota intestinal.
Evaluation of fecal pollution impact from a One Health perspective, in the southern region of Sétif (Algeria)
(Universitat de Barcelona, 2026-01-15) Boulainine, Dalal; García Aljaro, Cristina; Mezaache Aichour, Samia; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Water plays a fundamental role in sustaining life and supporting human social and economic development. However, its availability is unevenly distributed across the globe, and many regions are increasingly facing challenges related to water scarcity. These challenges are further exacerbated by rapid population growth, urbanization, and intensified agricultural and industrial activities. Among the regions most affected, the Mediterranean has seen a marked decline in water resources in recent years a trend expected to intensify due to the projected impacts of climate change. In this context, effective and sustainable water management has become a global priority, particularly through the lens of One Health and Global Health approaches, which recognize the interdependence of human, animal, and environmental health. A critical component of such management strategies is the monitoring of water quality, especially regarding microbial risks that pose threats to public health and ecosystem integrity. Within this framework, the primary objective of this doctoral thesis is to evaluate the impact of fecal pollution in aquatic environments, with the aim of identifying anthropogenic influences and associated public health risks. This research has been developed in various studies, resulting in three scientific articles. These studies have evaluated the dynamics and analysis of the main sources of fecal microbial contaminants, examining the presence of fecal indicators, antibiotic resistance genes (ARGs), and antibiotic-resistant bacteria (ARBs). They also examined the effectiveness of traditional and modern nature-based treatment techniques to reduce human fecal microbial contaminants, their impact on river community composition, and the natural deactivation processes of fecal indicator organisms. They also have examined environmental bacterial isolates from different environments including human and animal wastewaters, aquatic environments and vegetables, to understand the environmental reservoirs and transport pathways of antibiotic resistance genes ARGs and ARBs. The results obtained provide valuable information on the behavior of various fecal microbial indicators in the environment, and revealed that fecal pollution was widespread across the watershed, with higher concentrations primarily linked to urban wastewater discharges and animal sources. Changes in the structure and diversity of bacterial communities were also observed in this river. However, it was observed that the structure and diversity of bacterial communities can recover in the reservoir, demonstrating a high resilience to anthropogenic influences, as well as the persistence of ARGs and ARB in treated sewage, surface waters, and even fresh crops. Finally, the results of this research contribute to the evaluation of new water management tools in Algeria based on One Health and Global Health. It also highlights the urgent need for integrated monitoring, improved wastewater and agricultural practices, and a One Health approach to mitigate the public health risks associated with fecal contamination and antimicrobial resistance in Algerian watersheds.
Genome architecture and function in tissue regeneration
(Universitat de Barcelona, 2025-12-16) Llorens Giralt, Palmira; Corominas, Montserrat (Corominas Guiu); Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] Regenerative capacity differs not only across species, but also between organs and developmental stages within the same organism. Understanding why this variability exists remains a major challenge in developmental biology. Studies in regenerative animals have shown that genes involved in development are often re-expressed during regeneration and tend to be evolutionary conserved. Therefore, differences in regenerative potential may depend on the ability of a given tissue to reactivate these programs in a context-dependent manner, rather than on the presence or absence of necessary genes. This transcriptional reactivation is orchestrated through multiple layers of regulation, including transcription factor activity, cis-regulatory DNA elements, chromatin remodeling, histone modifications, and the three-dimensional organization of the genome. In this work, we explore how chromatin organization and genome activity influence regenerative responses by studying two epithelial tissues that regenerate through compensatory proliferation. First, we use a highly regenerative mammalian model, the two-thirds partial hepatectomy in mouse liver, to study changes in chromatin structure during regeneration. By integrating chromatin accessibility and transcriptomic data, we show that transcriptional reprogramming is driven by a wide range of cis-regulatory elements, including regeneration-responsive regulatory elements (RREs). These RREs, which comprise both regeneration-specific and reactivated developmental enhancers, activate genes involved in hepatocyte priming and proliferation. In contrast, enhancers linked to energy-intensive functions of quiescent hepatocytes become repressed, leading to the downregulation of metabolic genes, particularly those involved in lipid metabolism. We further construct a gene regulatory network that uncovers a cascade of transcription factor activation, which modulates expression of repair genes in a time-specific manner. Altogether, we provide a genome-wide atlas of enhancer-gene interactions that offers new insights into the regulatory mechanisms driving liver regeneration. To investigate whether and how chromatin structure and genome folding influence regenerative potential, an area that remains largely unexplored, we use the wing imaginal disc of Drosophila melanogaster as a model. We find that the spatial organization of the genome in wing disc cells undergoes subtle but significant reorganization during regeneration, including reduced compartmentalization. Notably, we observe increased contact frequency across three long-range chromatin loops, termed meta-loops, whose anchors span over 3 megabases. Using chromosomal deficiencies and CRISPR/Cas9-mediated knockouts, we demonstrate that these loops are required for regeneration but are dispensable during development. Finally, we identify the architectural protein Cp190 as a key factor in the formation of these meta-loops. The results of this thesis reveal critical roles for chromatin states and higher-order genome architecture in regeneration, highlighting conserved regulatory principles underlying tissue repair.
Análisis transcripcional y epigenético de las interacciones de factores de transcripción ectópicos en nuevos entornos de cromatina
(2025-05-23) López Fernández, Maria José; Maeso, Ignacio; Almuedo Castillo, María; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[spa] Las entidades biológicas interactúan continuamente con otras estructuras, estableciendo redes complejas a través de las cuales se intercambia información biológica. Estas redes de interacción, como las complejas redes reguladoras de genes que controlan el desarrollo animal, se ensamblan y mantienen durante el curso de la evolución. Sin embargo, ¿qué ocurre cuando de repente se incorporan por primera vez nuevos elementos a estos sistemas? La hipótesis planteada es que las propiedades intrínsecas de los nuevos elementos y el entorno en el que se expresan por primera vez influyen en el resto de su proceso coevolutivo. Sin embargo, sólo se pueden investigar aquellos encuentros moleculares que tuvieron éxito y sobrevivieron. Para superar esto, en este trabajo utilizamos un enfoque sintético en el que ponemos en contacto estructuras moleculares que nunca antes habían interactuado, para generar encuentros reguladores de genes por primera vez. En primer lugar, expresamos factores de transcripción (FT) específicos de mamíferos y plantas en el entorno de cromatina completamente extraño de embriones de pez cebra y estudiamos estas interacciones mediante técnicas ómicas. Dos de ellos, LEUTX y WRKY40, detienen el desarrollo temprano del pez cebra durante la gastrulación. Además, mediante experimentos de RNA-seq de embriones inyectados con LEUTX demostramos que la activación del genoma cigótico se ve afectada, y los experimentos de ATAC-seq muestran que reconoce el mismo motivo de ADN en el pez cebra que en las células humanas. Por otra parte, el estudio de enriquecimiento de motivos de los picos diferencialmente accesibles de WRKY40 reveló el motivo característico de la familia de genes apoptóticos p53, p63 y p73. Finalmente, ARGFX está asociado con un motivo característico de la familia de factores de transcripción T-box. Estos resultados sugieren que tanto WRKY40 como ARGFX se podrían estar uniendo a la cromatina de forma indirecta, interaccionando con los genes de las familias p53 y Tbx, respectivamente. Por otro lado, estudiamos las interacciones desde la perspectiva del entorno de la cromatina, analizando el impacto del cambio en la accesibilidad de la cromatina a través de 2 estrategias diferentes, las líneas transgénicas Gal4 y UAS, y la administración de diferentes drogas. La primera estrategia revela que el silenciamiento de transgenes regulados por UAS es un desafío técnico para el campo del pez cebra y es necesario más esfuerzo de investigación en este campo. Con respecto a la administración de drogas, establecemos que SAHA y entinostato a las concentraciones de 50 ng/μL producen un aumento del nivel de acetilación en el pez cebra y, por tanto, son 2 buenos candidatos para estudiar sus efectos a nivel genómico y para aplicarlos en los embriones inyectados con los FT estudiados. En definitiva, estos resultados sugieren que las estructuras moleculares como los FT tienen capacidades inherentes para interpretar y leer la información contenida en otros sistemas biológicos, incluso cuando estos encuentros son completamente inesperados. Así, durante las primeras etapas del origen de nuevas interacciones biológicas, estas propiedades intrínsecas pueden tener un impacto fundamental en el desenlace evolutivo final de las novedades moleculares.
The role of RBFOX1 and its regulatory network in major depression and comorbid psychiatric phenotypes
(Universitat de Barcelona, 2025-05-29) Adel, Maja Rebecca; Fernàndez Castillo, Noèlia; Freudenberg, Florian; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[eng] RBFOX1 is a master regulator of large transcriptomic networks with pleiotropic effects on numerous neurodevelopmental and psychiatric disorders, including major depressive disorder (MDD). MDD is a complex disorder characterized by symptoms such as anhedonia, hopelessness, feelings of emptiness and guilt. It frequently co-occurs with other psychiatric disorders, which can be partially explained by an overlapping genetic basis. Its aetiology involves complex interactions of biological predispositions, including a highly polygenic genetic basis, and environmental risk factors. In this doctoral thesis we aimed to study the role of RBFOX1 and its regulatory network on MDD and comorbid psychiatric phenotypes and characterize the underlying mechanisms through which this gene exerts its pleiotropic effects by using zebrafish models. To obtain a comprehensive overview, we followed a multi-faceted approach investigating the effects of RBFOX1 at the levels of genetics, transcriptomics, neurotransmission, brain activity, and behaviour. Our results show that RBFOX1 is an essential regulator of a large network of differentially spliced and expressed genes, which substantially contribute to the shared genetic basis of MDD, anxiety, irritability, and neuroticism. The RBFOX1-regulated genes associated with MDD and these comorbid traits show a substantial overlap and are mainly related to the neurodevelopment and neurotransmission. We furthermore identified enrichment of RBFOX1-regulated genes among genes causal for rare disorders with symptoms of depression, anxiety and irritability, suggesting that RBFOX1 contributes to these symptoms through shared pathways in psychiatric and rare disorders. Our integrative analyses highlighted 19 genes regulated by RBFOX1, including CADM2, CTSB, MAPT, NSF, PAX6, SCAI, SNCA, SP4, and TCF4, with a high pleiotropic effect on MDD and its comorbid traits. Experiments in two rbfox1 mutant zebrafish lines showed that the effects of rbfox1 on behavioural changes mirror various symptoms of different psychiatric disorders. We observed a range of consistent behavioural alterations in both lines, including hyperactivity, thigmotaxis, social impairments, and aggression in only one. The behavioural alterations in rbfox1 mutant zebrafish could be partially reversed through acute exposure to fluoxetine, a commonly prescribed antidepressant targeting the serotonergic system. In line with our findings of the involvement of RBFOX1-regulated genes in neurotransmission, we observed reduced levels of serotonin, dopamine, GABA, and glutamate in forebrain regions of adult rbfox1 mutant zebrafish. Interestingly, in vivo whole-brain imaging analyses showed increased neuronal activity in midbrain and hindbrain regions of rbfox1 mutant zebrafish larvae. Transcriptomic analyses of adult rbfox1 mutant zebrafish revealed numerous differentially spliced and expressed genes in all brain regions, and were enriched in cell development processes and DNA repair mechanisms. These transcriptomic changes likely explain the alterations in neurotransmission and neural activity.
Genòmica de la conservació aplicada a la baldriga balear (Puffinus mauretanicus)
(2025-05-09) Cuevas Caballé, Cristian; Riutort León, Marta; Rozas Liras, Julio A.; Universitat de Barcelona. Departament de Genètica, Microbiologia i Estadística
[cat] La baldriga balear (Puffinus mauretanicus) és l’ocell més amenaçat d’Europa. Les poblacions de l’espècie decreixen a un ritme de 7-14% anual, principalment degut a la mortalitat causada per la captura accidental (bycatch) per part de la flota pesquera, principalment del palangre, i per la depredació per part de mamífers invasors. Diferents estudis han previst que es podria extingir en només 60 anys si no es prenen ràpidament accions de conservació. L’objectiu principal d’aquesta tesi és l’aplicació de la genòmica de poblacions a l’estudi de la baldriga balear per tal d’augmentar el nostre coneixement sobre l’espècie i les seves poblacions, sempre tenint en compte que el coneixement generat haurà de servir per tal de fonamentar les accions de recuperació i conservació de les poblacions. Al Capítol 1, hem assemblat i anotat per primera vegada un genoma de referència de P. mauretanicus. Mitjançant la combinació de llibreries de seqüenciació de reads llargs i curts, el genoma assemblat per a la baldriga balear és d’elevada qualitat i completesa, comparable als genomes d’ocells d’alta qualitat disponibles. Aquest genoma és la pedra angular de tota la tesi. Al Capítol 2, mitjançant una anàlisi de genòmica de poblacions, hem identificat 1.304.832 SNPs que han mostrat la baixa diferenciació al llarg de tot el genoma entre la baldriga balear (P. mauretanicus) i la baldriga mediterrània (P. yelkouan). No obstant això, hem sigut capaços de delimitar 5 Unitats Evolutivament Significatives (ESU, en anglès): Pitiüses, Mallorca i Cabrera (pertanyents al llinatge P. mauretanicus) i Menorca i Yelkouan (pertanyents al llinatge P. yelkouan). De manera sorprenent, els resultats mostren que els individus mostrejats a Menorca, que fins ara es pensava que eren híbrids entre P. mauretanicus i P. yelkouan, pertanyen a P. yelkouan. Via anàlisis demogràfiques i de modelització per a diferents hipòtesis d’estructura poblacional i flux gènic, hem inferit que la separació entre la baldriga balear i la baldriga mediterrània va ocórrer durant l’últim màxim glacial (LGM), però durant el període interglacials de l’holocè s’ha facilitat la hibridació entre aquests tàxons, conduint a una reversió de l’especiació. Aquests contactes secundaris han permès mitigar una eventual depressió endogàmica a P. mauretanicus i que podria experimentar degut al seu baix cens poblacional. Tot i això, hem estimat el grau d’endogàmia i heterozigositat i no suposa a curt termini un risc per a la viabilitat de l’espècie. D’altra banda, les anàlisis de selecció mostren una regió de 200 kb on s’hi troba l’empremta genòmica d’una escombrada selectiva. En aquesta zona trobaríem haplotips adaptatius que tindrien a veure amb les estratègies migratòries diferents que presenten P. mauretanicus i P. yelkouan, i que s’han introgressat repetidament entre aquests llinatges. Combinant els paràmetres demogràfics prèviament estudiats per a la baldriga balear amb el grau de diversitat genètica que hem caracteritzat en aquesta tesi , hem simulat diferents projeccions del cens poblacional de P. mauretanicus sota diferents combinacions de flux gènic i intensitat del bycatch. Aquestes simulacions suggereixen que si no es pren acció immediata per tal de mitigar el bycatch, la baldriga balear s’extingirà en només 60 anys. A més, posen de manifest que inclús en els escenaris on s’aconsegueix mitigar el bycatch, el flux gènic entre la baldriga balear i P. yelkouan és crucial per a garantir la diversitat genètica suficient per a la supervivència de l’espècie. En resum, l’aplicació de la genòmica de poblacions a la baldriga balear permet posar èmfasi en que és una necessitat màxima protegir també el flux gènic entre els llinatges mauretanicus – yelkouan, que té lloc principalment a les illes de Menorca i Cabrera. Al Capítol 3 hem aprofitat tota aquesta caracterització de la diversitat genètica a nivell genòmic per a dissenyar un panell d’SNPs capaç de determinar el sexe i la colònia de procedència dels cadàvers de bycatch. Tot i la baixa diferenciació genètica entre les diferents colònies mostrejades (FST mitjana = 0,006), el panel proposat, de només 61 SNPs, ha sigut capaç d’assignar correctament la colònia de tots els individus mostrejats en provar-ne la seva validesa. El genoma de la baldriga balear ha sigut la primera espècie seqüenciada sota el paraigua de la la Iniciativa Catalana per a l’Earth BioGenome Project (CBP). Aquest genoma ha contribuït a comprendre els processos demogràfics que han modelat l’evolució de P. mauretanicus i P. yelkouan. A més, ha proporcionat un coneixement molt valuós per tal de poder guiar la presa de decisions per la conservació de la baldriga balear, alhora que ha proporcionat una eina pràctica que permetrà la determinació de l’origen del seu bycatch.

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