Biology subjects

Sanchez-Cabo, F.

Publications and source records attributed to Sanchez-Cabo, F..

At least 19 recordsLinked to original sources

bioRxiv2026

Abnormal ventricular wall patterning precedes and drives MYBPC3 hypertrophic cardiomyopathy

BACKGROUNDExcessive trabeculations and myocardial crypts are recurrent features across cardiomyopathies, yet their developmental origins and clinical significance remain poorly defined. To reveal the link between cardiac morphogenesis and disease, we generated humanized mouse models carrying patient-derived MYBPC3 frameshift mutations associated with overlapping hypertrophic cardiomyopathy (HCM) and left ventricular non-compaction (LVNC). METHODSWe applied CRISPR-Cas9 to introduce distinct MYBPC3 frameshift alleles into the mouse genome and performed comprehensive phenotypic and transcriptomic profiling from fetal life through adulthood. RESULTSAdult homozygous Mybpc3 frameshift mutant mice like humans displayed hallmark HCM; however, without LVNC. Fetal and neonatal mutant hearts exhibited markedly enlarged ventricular trabeculae and crypts that progressed postnatally into the observed adult hypertrophy. Transcriptomic analysis revealed stage-specific dysregulation of oxidative metabolism, nonsense-mediated decay (NMD), and cell cycle pathways, peaking at postnatal days 1 and 7, indicating that these stages represent critical time points in disease onset. The persistent NMD signature, also observed in phenotype-negative heterozygotes, suggests a compensatory stress response. Enlarged trabeculae exhibited 2-fold increased trabecular cardiomyocyte proliferation, reversing the normal compact-trabecular proliferative gradient and leading to impaired ventricular compaction in neonates. Hey2CreERT2 lineage tracing demonstrated invasion of Hey2+ compact cardiomyocytes into the trabeculae and ectopic trabecular expression of the Prdm16 transcription factor, indicating defective ventricular wall patterning and maturation. Postnatally, Hey2+-derived cardiomyocytes became restricted to the outer/compact myocardium in mutants, while the inner/trabecular myocardium underwent accelerated hypertrophy concurrent with Prdm16 downregulation. Mice with a Mybpc3 missense variant also exhibited Hey2+ myocardial lineage expansion into trabeculae but no increased proliferation, implicating additional mechanisms beyond Hey2 regulation. Postnatal Prdm16 restoration, via transgenic expression in Mybpc3-null mice effectively attenuated hypertrophy, establishing a causal link between Mybpc3 loss, Prdm16 decline, and pathological remodeling. CONCLUSIONSMybpc3 governs ventricular wall maturation by regulating cardiomyocyte proliferation, patterning, and maturation, partly via Prdm16. Disruption of these developmental programs precedes and drives adult HCM, highlighting a developmental role for sarcomeric proteins, and revealing postnatal Prdm16 modulation as an antihypertrophic therapeutic strategy.

developmental biology↗

bioRxiv2025

Functional rescue of critical-size bone defect using molecular network analysis of axolotl limb regeneration.

The salamander limb has served as a canonical model for successful regeneration that has yielded numerous molecular insights on its basic mechanism. Harnessing such information to induce regeneration in a non-regenerative setting has been a long-sought goal. The amputated salamander limb efficiently regenerates all of its missing bones, but paradoxically, a large bone gap without amputation - commonly called a Critical Size Defect (CSD) - is not regenerated, similarly to other vertebrates1. This non-regenerating injury provides a human-relevant setting to understand how to rescue lack-of-regeneration. Satoh and colleagues demonstrated in axolotl that transplantation of blastema cells from an amputated limb into a CSD yields cartilage bridging of the CSD2. This work provided a roadmap for rescuing the CSD. Here we asked, what are the crucial molecular differences in cells populating a regenerating blastema versus the CSD and can they be used to elicit CSD bridging? Previous genetic fate mapping and single cell transcriptomics showed that limb regeneration occurs via migration and dedifferentiation of fibroblastic, soft connective tissue (CT) cells that form a multipotent, skeletal stem cell 3, 4. Here using single cell transcriptomics and genetic fate mapping we found that the CSD is populated by CT cells that undergo a divergent molecular transition compared to the blastema. Using gene regulatory network (GRN) modeling, and lipid nanoparticle (LNP) delivery of mRNA we could express a single molecular factor, Wnt3a, to induce cartilage bridging of the CSD. Our results demonstrate the power of using molecular information from successful regeneration to rescue a non-regenerating injury.

cell biology↗

bioRxiv2025

HDL-associated phosphatidylserine blunts myeloid activation and protects from atherosclerosis

Background and aimsLipids play a critical role in atherosclerosis. Low-density lipoprotein (LDL)-cholesterol and certain lipid classes like sphingomyelins are associated with inflammation and poor cardiovascular outcomes. Phosphatidylserine (PS), on the other hand, is a negatively charged anti-inflammatory phospholipid class involved in efferocytosis. In this study, we sought to investigate its anti-atherosclerotic properties through a combination of complementary human lipidomics analyses, in vitro assays and in vivo experiments in Apoe-/- mice. MethodsHuman lipidomics studies were performed on the 300OB cohort comprising 300 obese and overweight individuals at risk of cardiovascular disease. In vitro assays were carried out using human monocytes and macrophages, and in vivo experiments included histopathological, immunophenotyping and single-cell transcriptomic analyses. ResultsIn humans, we identified PS as an anti-inflammatory and atheroprotective biomarker. Hence, we developed a high-density lipoprotein (HDL)-like formulation enriched in PS to exploit its properties in a targeted fashion in mice. In vitro, this formulation potently inhibited inflammatory cytokine production on human myeloid cells. Our in-depth in vivo experiments provided evidence of the formulations potent plaque-stabilizing and anti-inflammatory actions. These effects were mediated by a shift in the monocyte/macrophage compartment toward homeostatic/repairing phenotypes. ConclusionsCollectively, our results demonstrate that HDL-associated PS potently suppresses inflammation and atheroprogression, and holds promise as a viable approach to improve immunomodulatory therapies.

immunology↗

bioRxiv2025

Titin cleavage is a driver of cardiomyocyte disengagement and reactive myocardial fibrosis

Myocardial remodeling including cardiomyocyte-death-independent, reactive fibrosis and disconnection of cardiomyocytes is at the basis of prevalent cardiac conditions converging into arrhythmias and heart failure. However, the molecular mechanisms behind these pathogenic responses remain incompletely understood limiting therapeutic opportunities. Here, we find that a molecular event common to unrelated heart diseases, namely the cleavage of the sarcomeric protein titin, is enough to trigger fast myocardial remodeling. Using an engineered system based on the expression of tobacco etch virus protease (TEVp) in mice, we show that 30% mosaic cardiac titin cleavage leads to global cardiomyocyte disengagement, activation of cardiac fibroblasts and interstitial collagen deposition. These effects are concurrent, involve ERK1/2 signaling, and are expected to contribute, at least, to myocardial remodeling in chemotherapy-induced cardiotoxicity and ischemia damage.

physiology↗

bioRxiv2025

CIRCADIAN MODULATION OF NEUTROPHIL FUNCTION DETERMINES COLLATERAL PERFUSION AND OUTCOME AFTER ISCHEMIC STROKE

Stroke is a leading cause of mortality and disability, driven by complex and time-dependent mechanisms that aggravate ischemic damage. Among them, collateral perfusion determines the initial size of the ischemic core, the rate of its expansion, and the extent of the penumbra both at stroke onset and over time. Insufficiency of collaterals may occur due to genetic factors or other determinants, such as aging or cardiovascular risk factors, which reduce the number of collaterals or the diameter of those that remain. But aspects of less structural nature could also affect the effectiveness of these pathways by decreasing their patency. We hereby show that diurnal fluctuations in infarct volume in ischemic stroke mouse models are neutrophil phenotype-dependent, since differences in infarct volumes were abolished by depleting neutrophils or blocking their circadian clock, and linked to the collateral circulation: during the inactive phase of mice (daytime), collateral perfusion in the ipsilesional hemisphere was reduced, coinciding with an increase in intravascular neutrophil accumulation, suggestive of microvascular stalling. Single-cell transcriptomics, ex vivo functional assays and in vivo pharmacological and genetic strategies confirmed enhanced neutrophil extracellular traps (NETs) formation at this time. Importantly, in a cohort of human stroke patients, we identified diurnal oscillations in neutrophil and NET-related biomarkers, peaking during the human inactive phase (evening/night), and similarly associated with reduced collateral flow and poorer clinical outcomes. These findings underscore the critical role of neutrophils, their circadian dynamics and NET release in driving collateral insufficiency and ischemic brain damage, suggesting novel personalized therapeutic strategies based on circadian rhythms for the treatment of stroke.

immunology↗

bioRxiv2024

Epigenetic priming of embryonic lineages in the mammalian epiblast

Understanding the diversification of mammalian cell lineages is an essential to embryonic development, organ regeneration and tissue engineering. Shortly after implantation in the uterus, the pluripotent cells of the mammalian epiblast generate the three germ layers: ectoderm, mesoderm and endoderm1. Although clonal analyses suggest early specification of epiblast cells towards particular cell lineages2-4, single-cell transcriptomes do not identify lineage-specific markers in the epiblast5-11 and thus, the molecular regulation of such specification remains unknow. Here, we studied the epigenetic landscape of single epiblast cells, which revealed lineage priming towards endoderm, ectoderm or mesoderm. Unexpectedly, epiblast cells with mesodermal priming show a strong signature for the endothelial/endocardial fate, suggesting early specification of this lineage aside from other mesoderm. Through clonal analysis and live imaging, we show that endothelial precursors show early lineage divergence from the rest of mesodermal derivatives. In particular, cardiomyocytes and endocardial cells show limited lineage relationship, despite being temporally and spatially co-recruited during gastrulation. Furthermore, analysing the live tracks of single cells through unsupervised classification of cell migratory activity, we found early behavioral divergence of endothelial precursors shortly after the onset of mesoderm migration towards the cardiogenic area. These results provide a new model for the phenotypically silent specification of mammalian cell lineages in pluripotent cells of the epiblast and modify current knowledge on the sequence and timing of cardiovascular lineages diversification12,13.

developmental biology↗

bioRxiv2023

An ATF4-centric regulatory network is required for the assembly and function of the OXPHOS system

Identifying the factors that determine mammalian cell viability when oxidative phosphorylation (OXPHOS) function is impaired poses challenges due to the diverse cellular responses and limited clinical material availability. Moreover, animal models often fail to replicate human phenotypes. To address these challenges, this study conducted comprehensive analyses involving multiple defects and species by comparing the RNA-Seq expression profiles of human and murine cell lines with distinct nuclear backgrounds, representing both normal and OXPHOS-deficient models. To minimize species-specific variation, the study employed clustering techniques to group murine genes affected by OXPHOS dysfunction and identified crucial regulators like ATF4, UCP1, and SYVN1. ATF4 consistently displayed activation in response to OXPHOS defects, not only in murine but also in human cells, confirming its pivotal role in the cellular response to mitochondrial dysfunction. By integrating human and murine data, the study unveiled a conserved regulatory network encompassing genes related to the mTOR pathway and folate metabolism. Remarkably, the study uncovered an unexpected finding: the depletion of ATF4 in both mouse and human cells impairs OXPHOS assembly and supercomplex organization. This impairment primarily stems from a severe disruption in complex I assembly in the absence of ATF4, even under non-stress conditions.

molecular biology↗

bioRxiv2023

SpatialDDLS: An R package to deconvolute spatial transcriptomics data using neural networks

SummarySpatial transcriptomics has changed our way to study tissue structure and cellular organization. However, there are still limitations in its resolution, and most available plaXorms do not reach a single cell resolution. To address this issue, we introduce SpatialDDLS, a fast neural network-based algorithm for cell type deconvolution of spatial transcriptomics data. SpatialDDLS leverages single-cell RNA sequencing (scRNA-seq) data to simulate mixed transcriptional profiles with predefined cellular composition, which are subsequently used to train a fully-connected neural network to uncover cell type diversity within each spot. By comparing it with two state-of-the-art spatial deconvolution methods, we demonstrate that SpatialDDLS is an accurate and faster alternative to the available state-of-the art tools. Availability and implementationThe R package SpatialDDLS is available via CRAN-The Comprehensive R Archive Network: https://CRAN.R-project.org/package=SpatialDDLS. A detailed manual of the main functionalities implemented in the package can be found at https://diegommcc.github.io/SpatialDDLS. Contactfscabo@cnic.es Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2023

Incongruence between transcriptional and vascular pathophysiological cell states

The Notch pathway is a major regulator of transcriptional specification and vascular biology. Previous studies have suggested that targeting the ligand Dll4 or the Notch-receptors results in similar molecular and angiogenesis outcomes. Here, we analyzed single and compound genetic mutants for all Notch signaling members and found very significant differences in the way ligands and receptors regulate vascular homeostasis. Loss of Notch receptors, leads to minor vascular pathology featuring hypermitogenic MAPK-driven cell-cycle arrest and senescence. In contrast, loss of Dll4 triggers a strong Myc-driven switch towards cell proliferation and sprouting and major organ pathology. Targeting of Myc completely suppressed the proliferative and tip-cell angiogenic states induced by Dll4 loss-of-function, however, this did not avoid vascular pathology. Only VEGF blockade prevented the pathology induced by Dll4 loss, but without fully suppressing its transcriptional and metabolic programs. This study shows incongruence between single-cell transcriptional states and adult vascular phenotypes and related pathophysiology.

physiology↗

bioRxiv2021

Single cell clonal analysis identifies an AID-dependent pathway of plasma cell differentiation

Germinal centers (GC) are microstructures where B cells that have been activated by antigen can improve the affinity of their B cell receptors and differentiate into memory B cells (MBCs) or antibody secreting plasma cells. Activation Induced Deaminase (AID) initiates antibody diversification in GCs by somatic hypermutation and class switch recombination. Here we have addressed the role of AID in the terminal differentiation of GC B cells by combining single cell transcriptome and immunoglobulin clonal analysis in a mouse model that traces AID-experienced cells. We identified 8 transcriptional clusters that include dark zone and light zone GC subsets, plasmablasts/plasma cells (PB), 4 subsets of MBCs and a novel prePB subset, which shares the strongest clonal relationships with PBs. Mice lacking AID have various alterations in the size and expression profiles of these transcriptional clusters. We find that AID deficiency leads to a reduced proportion of prePB cells and severely impairs transitions between the prePB and the PB subsets. Thus, AID shapes the differentiation fate of GC B cells by enabling PB generation from a prePB state.

immunology↗

bioRxiv2020

Variant pathogenic prediction by locus variability, the importance of the last picture of evolution.

Accurate pathogenic detection for single nucleotide variants (SNVs) is a key problem to perform variant ranking in whole exome sequencing studies. Several in silico tools have been developed to identify deleterious variants. Locus variability, computed as Shannon entropy from gnomAD/helixMTdb variant allele frequencies can be used as pathogenic variants predictor. In this study we evaluate the use of Shannon entropy in non-coding mitochondrial DNA and also in coding regions with an additional selective pressure other than that imposed by the genetic code, as are splice-sites. To benchmark this functionality in non-coding mitochondrial variants, Shannon entropy was compared with HmtVar disease score, outperforming it in non-coding SNVs (AUCH=0.99 in ROC curve and PR-AUCH=1.00 in Precision-recall curve). In the same way, for splice-sites variants, Shannon entropy was compared against two state-of-the-art ensemble predictors ada score and rf score, matching their overall performance both in ROC curves (AUCH=0.95) and Precision-recall curves (PR-AUC=0.97). Therefore, locus variability could aid in variant ranking process for these specific types of SNVs. Contactjlcabreraa@cnic.es; fscabo@cnic.es

bioinformatics↗

bioRxiv2020

Impaired miRNA degradation by post-transcriptional addition of 3' cytosine and adenine in T cell activation

MiRNA repertoire of T cells undergoes extensive changes in response to activation. Whereas global miRNA downregulation occurs few hours after activation, some individual miRNAs are specifically up- or down-regulated. In this study, we have assessed miRNA expression and post-transcriptional modification kinetics in human primary CD4+ T cells upon short-term stimulation with CD3CD28 or IFN I using Next Generation Sequencing. Multiple miRNAs not related before with T cell activation profile have been identified as differentially expressed. Downregulated miRNAs presented higher 3 uridylation. Dis3L2 and Eri1 (3 to 5 exoribonucleases that prefer uridylated RNA as substrates) increased their expression upon TCR stimulation, probably generating an adverse environment for miRNAs. Remarkably, non-templated cytosine additions to 3 end, previously unknown to be a relevant post-transcriptional modification mechanism, were overrepresented in upregulated miRNAs, together with high levels of adenylation. In the midst of an increasing presence of exoribonucleases, miRNAs multiplying their levels may successfully escape degradation due to 3 cytosine and adenine addition. These protective signals open a new avenue to improve miRNA stability for therapy in T cells.

immunology↗

bioRxiv2020

Functional impact and regulation of global alternative splicing patterns in heart development and disease

Alternative splicing (AS) plays a major role in the generation of transcript diversity. In the heart, roles have been described for some AS variants and individual regulatory RNA binding proteins (RBPs); however, the global impact and regulation of AS patterns in cardiac pathophysiology is poorly understood. Here, we studied the AS profiles in heart disease, their relationship with heart development and the regulatory mechanisms control-ling AS dynamics in the mouse heart using a total of 136 RNA-seq samples. We found that AS and gene expression changes affect different genes, which are also involved in distinct biological functions. Developmental AS changes were more abundant and had stronger predicted impact on the encoded protein than those taking place during heart disease. However, AS changes in heart disease significantly modified protein interaction patterns and rewire the protein-protein interaction network. Using a database of experimentally determined binding sites of a large collection of RNA binding proteins, we studied the regulatory proteins associated to AS changes in each condition. Computational modelling revealed that developmental transitions were mainly driven by the up-regulation of MBNL1, whereas disease associated AS changes were driven by a more complex regulatory network, characterized by the interaction of different RNA binding proteins, with PTBP1 as the largest individual modulator. In adult mice, PTBP1 over-expression was sufficient to induce cardiac hypertrophy and diastolic dysfunction and significantly alter the AS profile. Overall, our study provides new in-sights into the functional impact of AS patterns in cardiac physiology and how computationally driven hypotheses can help to improve our understanding of RNA regulation and its contribution to heart disease.

bioinformatics↗

bioRxiv2020

Interplay between UNG and AID governs intratumoral heterogeneity in mature B cell lymphoma

ABSTRACTMost B cell lymphomas originate from B cells that have germinal center (GC) experience and bear chromosome translocations and numerous point mutations. GCs B cells remodel their immunoglobulin (Ig) genes by somatic hypermutation (SHM) and class switch recombination (CSR) in their immunoglobulin (Ig) genes. Activation Induced Deaminase (AID) initiates CSR and SHM by generating U:G mismatches on Ig DNA that can then be processed by Uracyl-N-glycosylase (UNG). AID promotes collateral damage in the form of chromosome translocations and off-target SHM, however, the exact contribution of AID activity to lymphoma generation and progression is not completely understood. Here we show using a conditional knock-in strategy that AID supraactivity alone is not sufficient to generate B cell transformation. In contrast, in the absence of UNG, AID supra-expression increases SHM and promotes lymphoma. Whole exome sequencing revealed that AID heavily contributes to lymphoma SHM, promoting subclonal variability and a wider range of oncogenic variants. Thus, our data provide direct evidence that UNG is a brake to AID-induced intratumoral heterogeneity and evolution of B cell lymphoma.View Full Text

cancer biology↗

bioRxiv2020

Axial skeleton anterior-posterior patterning is regulated through feedback regulation between Meis transcription factors and retinoic acid

Vertebrate axial skeletal patterning is controlled by coordinated collinear expression of Hox genes and axial level-dependent activity of Hox protein combinations. Transcription factors of the Meis family act as cofactors of Hox proteins and profusely bind to Hox complex DNA, however their roles in mammalian axial patterning have not been established. Similarly, retinoic acid (RA) is known to regulate axial skeletal element identity through the transcriptional activity of its receptors, however, whether this role is related to Meis/Hox activity in axial patterning remains unknown. Here we study the role of Meis in axial skeleton formation and its relationship to the RA pathway by characterizing Meis1, Meis2 and Raldh2 mutant mice. Meis elimination produces axial skeleton defects without affecting Hox gene transcription, including vertebral homeotic transformations and rib mis-patterning associated to defects in the hypaxial myotome. While Raldh2 and Meis positively regulate each other, Raldh2 elimination largely recapitulates the defects associated to Meis-deficiency and Meis overexpression rescues the axial skeletal defects in Raldh2 mutants. We propose a Meis-RA positive feedback loop whose output is Meis levels and is essential to establish anterior-posterior identities and pattern of the vertebrate axial skeleton.

developmental biology↗

bioRxiv2019

Frequency Conservation Score (FCS): the power of conservation and allele frequency for variant pathogenic prediction

BackgroundPrediction of pathogenic variants is one of the biggest challenges for researchers and clinicians in the time of next-generation sequencing technologies. Stratification of individuals based on truly pathogenic variants might lead to improved, personalized treatments.\n\nResultsWe present Frequency Conservation Score (FCS) and Frequency Conservation Score for Mitochondrial DNA (FCSMt) two methods for the detection of pathogenic single nucleotide variants in nuclear and mitochondrial DNA, respectively. These scores are based in a random forest model trained over a set of potentially relevant predictors: (i) conservation scores (PhastCons and phyloP); (ii) locus variability at each genomic position built from gnomAD database and (iii) physicochemical distance for amino acids substitutions and the impact/consequence over the canonical transcript. FCS showed an AUC of 98% for deleteriousness in an independent validation dataset, outperforming other scores such as metaLR, metaSVM, REVEL, DANN, CADD, SIFT, PROVEAN or FATHMM-MKL. Moreover, FCSMt presented an AUC=0.92 for pathogenic mitochondrial SNVs detection. The tool is available at http://bioinfo.cnic.es/FCS\n\nConclusionsFCS and FCS-Mt improve pathogenic mutation detection, allowing the prioritization of relevant variants in Whole Exome and Whole Genome Sequencing Analysis.

bioinformatics↗

bioRxiv2019

Coronary arterial development is regulated by a Dll4-Jag1-EphrinB2 signaling cascade

Coronaries are essential for myocardial growth and heart function. Notch is crucial for mouse embryonic angiogenesis, but its role in coronary development remains uncertain. We show Jag1, Dll4 and activated Notch1 receptor expression in sinus venosus (SV) endocardium. Endocardial Jag1 removal blocks SV capillary sprouting, while Dll4 inactivation stimulates excessive capillary growth, suggesting that ligand antagonism regulates coronary primary plexus formation. Later endothelial ligand removal, or forced expression of Dll4 or the glycosyltransferase MFng, blocks coronary plexus remodeling, arterial differentiation, and perivascular cell maturation. Endocardial deletion of Efnb2 phenocopies the coronary arterial defects of Notch mutants. Angiogenic rescue experiments in ventricular explants, or in primary human endothelial cells, indicate that EphrinB2 is a critical effector of antagonistic Dll4 and Jag1 functions in arterial morphogenesis. Thus, coronary arterial precursors are specified in the SV prior to primary coronary plexus formation and subsequent arterial differentiation depends on a Dll4-Jag1-EphrinB2 signaling cascade.

developmental biology↗

bioRxiv2019

Nanog regulates Pou3f1 expression and represses anterior fate at the exit from pluripotency.

Pluripotency is regulated by a network of transcription factors that maintains early embryonic cells in an undifferentiated state while allowing them to proliferate. NANOG is a critical factor for maintaining pluripotency and its role in primordial germ cell differentiation has been well described. However, Nanog is expressed during gastrulation across all the posterior epiblast, and only later in development its expression is restricted to primordial germ cells. In this work, we unveiled a previously unknown mechanism by which Nanog specifically represses the anterior epiblast lineage. Analysis of transcriptional data from both embryonic stem cells and gastrulating mouse embryos revealed Pou3f1 expression to be negatively correlated with that of Nanog during the early stages of differentiation. We have functionally demonstrated Pou3f1 to be a direct target of NANOG by using a dual transgene system for the controlled expression of Nanog. Use of Nanog null ES cells further demonstrated a role for Nanog in repressing anterior neural genes. Deletion of a NANOG binding site (BS) located nine kilobases downstream of the transcription start site of Pou3f1 revealed this BS to have a specific role in the regionalization of the expression of this gene in the embryo. Our results indicate an active role of Nanog inhibiting the neural fate by repressing Pou3f1 at the onset of gastrulation.

developmental biology↗