Biology subjects

Otaegui, D.

Publications and source records attributed to Otaegui, D..

9 recordsLinked to original sources

bioRxiv2026

Detergent lysis controls are critical for single extracellular vesicle phenotyping in SP-IRIS platforms

Bulk analytical methods are extensively used to identify molecular cargo in extracellular vesicles (EVs), but they cannot distinguish nanovesicles from co-isolated non-vesicular particles or resolve discrete EV subpopulations. Single-particle interferometric reflectance imaging coupled with immunofluorescence (SP-IRIS/IF) has emerged as a platform to simultaneously resolve particle size, count, and cargo at single-EV resolution. However, differentiating authentic EVs from non-vesicular signals requires rigorous methodological controls that remain widely underreported in standard workflows. In this study, we analyzed human neural secretomes and serum EVs across different SP-IRIS/IF setups to evaluate their analytical specificity. By using detergent-mediated lysis controls, we found a critical technical limitation in standard single-particle analysis: the persistence of both interferometric and fluorescent signals after detergent treatment in a subset of presumed single-EV events that were non-vesicular artifacts. These findings indicate that lacking appropriate controls on single-EV platforms risks data misinterpretation, underscoring the indispensable implementation of detergent-mediated lysis for single-particle analysis in SP-IRIS/IF platforms.

cell biology↗

bioRxiv2026

Extrachromosomal circular DNA formation follows a 10.5-bp periodicity reflecting the minor groove orientation around nucleosomes

Extrachromosomal circular DNA is increasingly recognized as a central feature of somatic evolution in cancer, yet the structural determinants of eccDNA formation remain poorly understood. Here, we systematically analysed eccDNA length distributions and breakpoint organization across 25 datasets spanning multiple species, tissues and experimental approaches. Applying a short-time Fourier transform to eccDNA length distributions enabled us to resolve recurrent periodicities at approximately 180 bp and 10.5 bp, consistent with nucleosome length and the helical repeat of DNA, respectively. Mapping eccDNA breakpoints relative to nucleosome dyads revealed preferential breakpoint formation in linker DNA and, within nucleosome-bound DNA, at positions where the minor groove faces towards the histone surface. Our results show that eccDNA formation is shaped not only by nucleosome positioning but also by the rotational orientation of the DNA double helix around the nucleosome. Together, these findings identify chromatin geometry as a determinant of eccDNA biogenesis and provide a structural basis for the length periodicities that characterize eccDNA formation.

genomics↗

bioRxiv2026

MBIOME: A comprehensive, reproducible, and open-source workflow for amplicon-based microbiome data analysis.

Microbiome analysis has become a pivotal tool in understanding the role of microbiota in human health and disease. However, the lack of standardized workflows, together with the limitations of proprietary software solutions, hampers reproducibility and flexibility. Here, we present Mbiome, an open-source, user-friendly and automated workflow designed to streamline amplicon-based microbiome analysis. Built upon QIIME2, Mbiome supports both bacterial (16S rRNA) and fungal (ITS) profiling, and is compatible with raw fastq files generated by Ion Torrent (IT) and Illumina (IL) sequencing platforms. The workflow guides users through an interactive setup process via a simple configuration file, enabling researchers with minimal bioinformatics experience to perform comprehensive analyses without writing code. Once configured, Mbiome automates major steps including quality control, taxonomic assignment, and {beta}-diversity analyses, functional predictions (via q2-metnet), and customizable visualizations and statistical analyses. Mbiome has been validated using real-world datasets from multiple sclerosis research projects, performing a comparison between different microbiome analysis approaches, including 16S hypervariable region reconstruction, amplicon-based strategies, and cross-platform sequencing (IT and IL), as well as against results obtained with Ion Reporter (IR) commercial software. This evaluation demonstrated its versatility and effectiveness across different sequencing platforms. Moreover, Mbiome provided improved flexibility, transparency, and taxonomic resolution compared to IR. By combining accessibility, reproducibility, and cross-platform compatibility, Mbiome lowers the barrier to microbiome data analysis and facilitates high-quality, standardized workflows in both research and applied settings. Mbiome is publicly available at https://github.com/MGorostidi/mbiome.

bioinformatics↗

bioRxiv2026

Oncostatin M cytokine promotes breast cancer progression by remodelling the extracellular matrix and activating integrin signalling in cancer cells

Tumours reshape their surrounding extracellular matrix (ECM), creating a microenvironment with altered chemical and mechanical properties. Integrins detect these changes, linking the ECM to the intracellular cytoskeleton and promoting cell survival, motility, invasion and differentiation, and further ECM remodelling. However, the molecular mechanisms by which tumours remodel their ECM are not well understood. Here, we found that the cytokine oncostatin M (OSM) promotes breast cancer progression by activating ECM remodelling and integrin signalling in cancer cells, as shown by combining complementary in vitro, in ovo and in vivo models, and transcriptomic and proteomic analyses. We demonstrated that OSM induces fibrosis, characterized by increased collagen deposition and hydroxylation, together with activation of ECM and ECM-associated proteins and modifiers such as fibronectin, tenascin C, LOX, PLOD2 and collagen prolyl hydroxylases. OSM also promoted the expression of integrins. Integrin alpha 5 (ITGA5) was identified as an important mediator of OSM-effects. ITGA5 blockade, by means of small interference RNA and therapeutic inhibition with a blocking peptide, abrogated OSM-induced cancer cell migration, invasion and in vivo tumour growth. In addition, OSM blockade with a specific inhibitor reduced tumour growth in an immunocompetent mouse model. Our results are clinically relevant as the expression of integrins and matrisome genes strongly correlated with OSM and its receptor OSMR in breast cancer clinical samples; and co-expression of OSMR and ITGA5 associated with decreased survival in basal breast cancer patients. Collectively, our data reinforce the potential of the OSM-ITGA5 axis as a therapeutic target in this breast cancer subtype, which shows the highest mortality rates.

cancer biology↗

bioRxiv2026

Circular RNA-associated QTLs show stronger association with splicing-QTLs than with expression-QTLs

IntroductionCircular RNA quantitative trait locus (circQTLs) have emerged as a class of regulatory variants, but their mechanistic basis remains poorly characterized. Understanding how genetic variation influences circRNA biogenesis is essential to clarify their role in post-transcriptional gene regulation. MethodsWe systematically compared circQTLs with matched splicing (sQTL) and expression (eQTL) datasets. Using bootstrap-based Jaccard similarity analyses, we quantified genomic overlap patterns and assessed their statistical significance. We further validated these findings across independent circQTL studies. In addition, we analyzed the genomic distribution of circQTLs to identify enrichment patterns across functional genomic regions. ResultscircQTLs exhibited a statistically significant but modestly stronger genomic overlap with sQTLs compared to eQTLs. This pattern was consistent across independent datasets despite limited reproducibility of individual circQTL signals. Genomic annotation revealed distinct distributional patterns, including depletion in exonic regions and relative enrichment in non-coding genomic contexts compared to other QTL classes. DiscussionTogether, these results suggest that circRNA-associated regulatory variation is preferentially linked to splicing-related mechanisms rather than transcriptional control of host genes. However, the modest effect size indicates that this relationship is not exclusive, and likely reflects a mixture of shared splice-site regulatory effects and additional mechanisms specific to back-splicing that are not captured by conventional sQTL or eQTL frameworks. This dual architecture positions circRNA biogenesis at the interface between splicing dynamics, RNA structure, and higher-order genomic organization, supporting circQTLs as a distinct layer of post-transcriptional gene regulation.

genetics↗

bioRxiv2026

A dual-function variant on chromosome 17 regulates circRNA expression and splicing in multiple sclerosis

Multiple sclerosis (MS) is a chronic autoimmune demyelinating disease of the central nervous system with a complex etiology. Recent genomic studies highlight the contribution of expression quantitative trait loci (eQTLs) in modulating gene expression and disease susceptibility. Given the emerging role of circular RNAs (circRNAs) in MS, we hypothesized that genetic variants may regulate circRNA expression through circRNA-specific eQTLs (circ-eQTLs). We performed a cis-circ-eQTL analysis integrating circRNA expression and whole-genome genotyping data from 30 MS patients and 18 healthy controls using a linear regression model adjusted for disease status and sex. Candidate circ-eQTLs were prioritized based on MS-associated regions and known splicing QTLs (sQTLs) from GTEx and validated in an independent cohort (67 MS, 64 controls). Association analysis in a larger cohort (2831 MS, 3191 controls) evaluated two candidate variants for MS risk. We identified 42,077 significant cis-circ-eQTLs and validated three. Two SNPs, rs7214410 and rs11079784, modulated hsa_circ_0106983 expression, and rs7214410 also acted as an sQTL affecting EFCAB13 splicing. rs7214410 showed stronger association with MS than rs11079784. Our findings reveal extensive genetic regulation of circRNA expression and highlight rs7214410 as a dual-function variant refining the MS susceptibility locus on chromosome 17.

genetics↗

bioRxiv2025

Comparative analysis of eccDNA and circRNA tools shows increased accuracy of tool combination

IntroductionCircular nucleic acids such as extrachromosomal circular DNA (eccDNA) and circular RNA (circRNA) are increasingly recognized for their biological relevance and potential as biomarkers in disease contexts. Despite their growing importance, their detection remains challenging due to tool-specific biases, limited validation frameworks, and high variability in performance across datasets. MethodsWe benchmarked 10 circle detection tools across diverse conditions using both simulated and biological datasets. Our evaluation included classical performance metrics and a novel internal measure of read distribution symmetry ({Delta}CJ) to assess circle prediction confidence. We explored the impact of sequencing protocols, filtering strategies, and combined tool consensus. ResultsWe found that detection accuracy was highly influenced by sequencing depth, alignment algorithm, and experimental enrichment protocols. {Delta}CJ proved effective in flagging potential false positive circles, showing improved accuracy of Intersect (circles detected by all tools) and Rosette (circles detected by[≥] 2 tools) combinations. DiscussionThis study offers a broad evaluation of circular detection tools, suggesting that the combination of[≥] 3 tools is necessary for a correct prediction. These insights will inform future experimental design and data analysis pipelines in both experimental and clinical settings.

bioinformatics↗

bioRxiv2024

A proposed workflow to analyze bacterial transcripts in RNAseq from blood extracellular vesicles of people with Multiple Sclerosis

MotivationThe taxonomical characterisation of bacterial species derived from genetic material blood, including reads derived from bacterial extracellular vesicles (bEVs) poses certain challenges, such as the proper discrimination of "true" reads from contaminating reads. This is a common issue in taxa profiling and can lead to the false discovery of species that are present in the sample. To avoid such biases a careful approximation to taxa profiling is necessary. ResultsIn this work we propose a workflow to analyze the presence of bacterial transcripts as indicative of putative bEVs circulating in the blood of people with MS (pwMS). The workflow includes several reference mapping steps against the host genome and a consensus selection of genera based on different taxa profilers. The consensus selection is performed with a flagging system that removes species with low abundance or with high variation across profilers. Additionally, the inclusion of biological samples from known cultured species as well as the generation of artificial reads constitute two key aspects of this workflow. AvailabilityThe workflow is available at the following repository: https://github.com/NanoNeuro/EV_taxprofiling. Contactdavidangel.otaeguibichot@osakidetza.eus

bioinformatics↗

bioRxiv2022

Peripheral Myeloid-Derived Suppressor Cells are good biomarkers of the efficacy of Fingolimod in Multiple Sclerosis

The increasing number of treatments that are now available to manage patients with multiple sclerosis (MS) highlights the need to develop biomarkers that can be used within the framework of individualized medicine. Fingolimod is a disease-modifying treatment that belongs to the sphingosine-1-phosphate receptor modulators. In addition of inhibiting T cell egression from lymph nodes, fingolimod promotes the immunosuppressive activity of Myeloid-Derived Suppressor Cells (MDSCs), a cell type that can be used as a biomarker of disease severity, and of the degree of demyelination and extent of axonal damage in MS. In the present study, we have assessed whether the abundance of circulating monocytic-MDSCs (M-MDSCs) may represent a useful biomarker of fingolimod efficacy. As such, blood immune cells were analyzed at disease onset in the experimental autoimmune encephalomyelitis (EAE) MS mouse model. Fingolimod treated animals presented a milder EAE course with less demyelination and axonal damage, although a few animals did not respond well to treatment and they invariably had fewer M-MDSCs prior to initiating the treatment. Remarkably, M-MDSC abundance was also found to be an important and specific parameter to distinguish EAE mice prone to better fingolimod efficacy. Finally, in a translational effort, M-MDSCs were quantified in MS patients at baseline and correlated with different clinical parameters after 12 months of fingolimod treatment. The data obtained indicated that the M-MDSCs at baseline were highly representative of a good therapeutic response to fingolimod, i.e. patients who met at least two of the criteria used to define non-evidence of disease activity (NEDA-3) 12 months after treatment, providing relevant information of intention-to-treat MS patients. Collectively, our data indicate that M-MDSCs might be a useful predictive biomarker of the response of MS patients to fingolimod.

immunology↗