Biology subjects

Prados, J.

Publications and source records attributed to Prados, J..

15 recordsLinked to original sources

bioRxiv2026

A cellular atlas of Astyanax mexicanus embryogenesis reveals principles of developmental microevolution

The evolution of development is the primary mechanism by which morphological and functional diversity is generated. Comparing the developmental trajectories of single cells in entire embryos across species can reveal how gene expression is conserved and varies, as well as the molecular mechanisms underlying how cell- and tissue-specific changes in developmental regulation generate phenotypic differences. However, this perspective has not addressed the mechanisms of small-scale developmental microevolution, i.e, in closely-related species or even within a single species. The fish Astyanax mexicanus, which comes in two forms adapted to distinct environments - the eyed, river-dwelling morph and the eyeless, cave-adapted morph - is a remarkable model with which to investigate the evolution of gene regulation in a single species. Here, we generated and analyzed a single-nucleus RNA-seq atlas of the development of the two Astyanax morphs, from gastrula to swimming larva. Despite extensive changes in gene expression observed in the eight examined stages, for both ubiquitously-expressed and tissue-specific genes, and in both embryonic and extra-embryonic tissues, the temporal progression of cavefish and surface fish cells was similar in ordinal regression-based pseudotime models using 20-100 core genes. However, transcriptome-wide comparisons of gene expression waves revealed massive heterochronic shifts. In cavefish, some tissues had a majority of genes lagging behind in their transcriptional dynamics (e.g. the optic region), while in others, a majority of genes had accelerated dynamics (e.g. the mesoderm), or else, an equivalent number of genes were found in either category (e.g. the endoderm). Furthermore, cell fate acquisition and differentiation were variably affected in different cavefish lineages, with delays of ~6 hours in the retina, ~2 hours in the axial mesoderm and ~none in the extra-embryonic enveloping layer. Thus, cavefish embryos progress as a mosaic of different tissues, or modules, each following its own transcriptional tempo. Because gene expression differential regulation and time shifts were more prevalent in the early stages and gradually diminished throughout development, we propose that cavefish embryos initially undergo strong decanalisation before becoming re-canalized in line with the shared Astyanax developmental program.

developmental biology↗

bioRxiv2026

Transplantation reveals birthdate-dependent post-mitotic competence in cortical neurons

Cortical glutamatergic neurons are generated through temporally ordered developmental programs that link neuronal birthdate to laminar position and identity. Yet, because differentiation continues after cell-cycle exit, it remains unclear how far post-mitotic identity can be reshaped by the environment and whether this residual competence varies across neuronal types. Here, we used transplantation to uncouple birthdate from final laminar position by placing newborn neurons in ectopic laminae in the postnatal mouse cortex. E13-born (i.e. deep layer-destined) and E15-born (i.e. superficial layer-destined) donor neurons displayed distinct birthdate-associated molecular programs before transplantation. After grafting into P0 hosts, both populations migrated and settled across cortical layers, enabling comparison across laminar environments. We simultaneously profiled transcriptomic, morphological and electrophysiological neuronal identities using Patch-seq. Transplanted neurons developed pyramidal morphologies and active membrane properties, but their final identity was not imposed by their laminar position. Thus, the postnatal cortex supports integration and maturation but does not impose canonical laminar identity. Critically, neuronal competence was asymmetric across birthdates. E15-born neurons remained committed toward a robust superficial layer neuron-like profile independently of cortical position, whereas E13-born neurons retained a broad, non-canonical differentiation rather than either converging on a deep- or superficial-layer reference state. This is consistent with the wider intrinsic repertoire of early-born neurons, which the postnatal environment does not resolve. Thus, post-mitotic identity is constrained by developmental history in a birthdate-dependent manner, with early- and late-born neurons retaining fundamentally different differentiation potentials after cell-cycle exit.

neuroscience↗

bioRxiv2026

Discoidins are cytosolic lectins that shape intracellular host defence by sensing virulence-associated mycobacterial glycolipids and glycopeptidolipids

Mycobacteria are surrounded by a dynamic, glycolipid-rich envelope that controls both virulence and immune recognition, yet how cytosolic lectins detect and interpret the organization of surface glycans during intracellular infection remains unclear. Here, we show that Dictyostelium discoideum discoidins act as cytosolic sensors of mycobacterial envelope organization. During Mycobacterium marinum infection, Discoidin A and Discoidin E assemble into foci on intracellular bacilli and bind surface-associated glycan patches. Using transposon mutagenesis, synthetic mycobacterial glycan arrays, biochemical fractionation and targeted envelope mutants, we find that discoidins recognize a restricted set of lipid-linked glycans enriched in methylated L-rhamnose motifs, including structures associated with LOS, PGL and GPL. Binding depends on the H-type lectin {beta}-galactoside-binding pocket and is inhibited by point mutations and TDG, a soluble disaccharide competitor, demonstrating glycan-dependent recognition. Specific assays led to exclusion of major structural carbohydrates, including AG-PG, AM/LAM, -glucan and TDM, while protease treatment of capsular material and fractionation of polar lipids, identified both GPL and LOS-like glycolipids as dominant discoidin ligands. Disruption of LOS biosynthesis or PDIM/PGL-dependent envelope organization reduced discoidin binding to intact bacteria even though some ligands remained detectable by dot blot in various envelope extracts. Thus, discoidins do not simply detect ligand abundance, but binding depends on envelope perturbations and unmasking of glycolipids and glycopeptidolipids. These findings reveal pathogen envelope glycans as direct targets of cytosolic lectin surveillance and establish discoidins as probes of mycobacterial envelope remodelling during intracellular infection. Author SummaryMany disease-causing bacteria are surrounded by a protective outer layer that helps them survive inside host cells and avoid being eliminated. This surface is not static, it can be remodelled during infection, changing which molecules are exposed to the host. How host cells detect these changes is still not fully understood. We studied this question using Dictyostelium discoideum, a single-celled amoeba that shares many cellular defence mechanisms with human immune phagocytes. We focused on discoidins, a family of proteins that bind sugars. We found that discoidins accumulate on intracellular Mycobacterium marinum, a close relative of Mycobacterium tuberculosis, the bacterium that causes tuberculosis, and recognize specific sugar-containing molecules exposed at the bacterial surface. Importantly, discoidins do not simply detect whether these molecules are present. Instead, they respond to how they are displayed and exposed on the bacterial surface. Changes in the organization of the bacterial outer layer strongly affected discoidin binding, masking or revealing specific target molecules. Our findings show that discoidins act as sensors of bacterial surface remodelling during infection. More broadly, they reveal an ancient mechanism by which host cells can monitor pathogens by detecting changes in the sugars exposed on their surface.

microbiology↗

bioRxiv2026

Prenatal assembly of functional cortical circuits

The extent to which early brain maturation requires interactions with the outside world is a central question in neurobiology. Because much of cortical maturation occurs after birth in Mus, it has often been viewed as dependent on postnatal experience. However, studies addressing this issue have largely relied on postnatal sensory deprivation paradigms, which perturb normal development and cannot determine to what extent postnatal experience per se drives maturation. To address this question, here we compare two related rodents with markedly different gestation lengths: the precocial Acomys dimidiatus (39-day gestation) and the altricial Mus musculus (19-day gestation). By generating a novel Acomys reference genome and using histology, cellular birth dating, electrophysiology, single-nucleus transcriptomics, and quantitative behavior, we show that Acomys preserves the canonical sequence and timing of cortical development, while shifting major milestones of neuronal, circuit and behavioral maturation into prenatal life. At birth, Acomys cortex already shows advanced cytoarchitecture, neuronal physiology, thalamocortical barrels, transcriptional states, and sensorimotor behavior, with postnatal molecular programs in Mus unfolding prenatally in Acomys. Thus, birth is not a prerequisite for early cortical maturation. Instead, evolutionarily conserved developmental programs unfold across birth, with birth occurring at different stages of these programs, reflecting a species-specific balance between neonatal competence and prolonged postnatal plasticity.

neuroscience↗

bioRxiv2026

Deletion of TNFR1 in astrocytes restores memory in aged Alzheimer's disease mice

Astrocytes participate in local inflammation and cognitive decline in Alzheimers disease (AD). Aberrant cytokine TNF signaling via astrocyte type-1 receptor (aTNFR1) could causally link the two AD pathology aspects. To verify this hypothesis, we crossed transgenic AD mice with mice enabling astrocyte-specific conditional TNFR1 deletion (aTNFR1KO). Induction of aTNFR1KO at early AD stages, preserved memory and reduced {beta}-amyloid load and astrogliosis in the aged mice. Induction of aTNFR1KO at late AD stages, in mice already memory-impaired, surprisingly produced rapid memory rescue, without affecting {beta}-amyloid load and astrogliosis. Single nucleus-RNA-seq analysis of all hippocampal cell populations revealed that late-stage aTNFR1KO rapidly modifies gene expression mainly in neurons, primarily targeting synaptic pathways, causing combined glutamatergic downregulation and GABAergic up-regulation. Consistently, hippocampal EEGs showed a pro-inhibitory effect of aTNFR1KO, which thus restores memory by "rebalancing" hippocampal circuitry excitability. This pro-memory effect identifies a new mechanism and astrocyte target against cognitive decline in AD.

neuroscience↗

bioRxiv2026

Early metabolic reprogramming licenses Streptococcus pneumoniae for Influenza-driven superinfection

Bacterial pneumonia remains a major cause of morbidity and mortality following influenza A virus (IAV) infection. However, the adaptive mechanisms that enable pathogen expansion in the post-viral lung remain poorly defined. Here, using a mouse model of IAV-Streptococcus pneumoniae superinfection, we characterize bacterial transcriptional reprogramming in vivo. We identify alcohol dehydrogenases (AdhA and AdhE), that support NAD regeneration during mixed-acid fermentation, as key determinants of bacterial fitness specifically in the IAV-primed lung. Genetic deletion of these results in a pronounced fitness defect during superinfection but not in primary bacterial pneumonia. Consistent with this requirement, pharmacological inhibition of alcohol dehydrogenases limits bacterial expansion and dissemination following IAV infection. Mechanistically, we show that IAV infection profoundly remodels the lung environment, inducing hypoxia and increasing the availability of alternative carbon sources, which together impose a metabolic dependency on Adh for bacterial expansion. Our findings place metabolic adaptation as a central driver of pneumococcal outgrowth following viral infection and reveal exploitable vulnerabilities for therapeutic intervention.

microbiology↗

bioRxiv2025

Neurogliaform cells mediate interhemispheric modulation of sensory-evoked activity in cortical pyramidal neurons

Integration of bilateral sensory inputs requires effective communication between brain hemispheres. This interhemispheric communication is essential for sensory perception and involves reciprocal connections between homotopic sensory areas. A key role in this process is attributed to interhemispheric inhibition which, owing to its long-lasting form, is posited to operate largely through neurogliaform cells (NGCs). However, direct evidence on the role of NGCs in interhemispheric inhibition is missing. Here we show that NGCs in the mouse barrel cortex (BC) are engaged by interhemispheric callosal projections to modulate pyramidal neuron (PN) activity and sensory perception. Using optogenetics, ex vivo whole-cell recordings, and in vivo calcium imaging, we found that layer 1 and layer 2/3 (L1-3) NGCs are strongly activated by the callosal and suppressed by the thalamocortical pathway, suggesting that NGCs encode ipsilateral rather than contralateral whisker stimuli. We also found that direct stimulation of L1-3 NGCs modulates whisker-evoked activity in L2/3 and L5b PNs, and increases the perceptual threshold in a whisker-deflection detection task. Furthermore, these effects were recapitulated by direct stimulation of callosal projections and deflection of the ipsilateral whiskers respectively, suggesting that the effect of the callosal pathway on sensory perception is mediated by NGCs. Our results not only prove that NGCs mediate interhemispheric inhibition, but also demonstrate their role in sensory perception via modulation of the main units involved in cortical input and output.

neuroscience↗

bioRxiv2025

Cross-species comparative modelling of antimicrobial host responses ex vivo in human and murine precision cut lung slices and in vivo in mice

Tissue resident host responses to microbial infections in the respiratory tract are highly dynamic in space and time and rely on the interaction of a multitude of cell types. In an attempt to model these multicellular responses reliably in cell culture, we compare here the global transcriptional antimicrobial response to infection with influenza A virus (IAV) in precision cult lung slices (PCLS), volume defined organ discs largely maintaining the cellular composition and 3D architecture of the donor lung. To permit a fair comparison of host responses in an isogenic background we first challenged mice in vivo and murine PCLS (mPCLS) and assess host transciptomic changes by unbiased RNAseq. While core antiviral responses overlapped substantially, mPCLS lacked certain features--such as type II interferon expression--likely due to the absence of infiltrating immune cells responses. Importantly, when expanding our findings to immune experienced human precision cut lung slices (hPCLS), we find a much broader antiviral response after IAV challenge, including type I, II and III interferons, suggesting the presence of responsive tissue resident lymphocytes. To prove specificity of this response we infected hPCLS with Streptococcus pneumoniae. Ex vivo tissues responded with a distinct proinflammatory gene profile including IL1A, IL1B and IL17 expression. Blocking of IL-1 signaling partially inhibited the proinflammatory response, suggesting cellular cross-talk and a complex and specific antimicrobial reaction in this ex vivo model. In conclusion diversified tissue resident immune cell compartment distinguishes the human ex vivo model, making it an ideal system for microbiological and immunological research. ImportancePathogen interactions with the lung are very dynamic processes. In biomedical research it is paramount to model these processes in the laboratory as accurately as possible. Influenza A virus has been extensively studied in epithelial cell culture models, including advanced organoids and organ on a chip systems. We use here ex vivo cultured PCLS and use transcriptomics to assess the global tissue resident host response to viral and bacterial challenge. Our data show 1) that murine PCLS faithfully reflect core responses to viral infection, while missing proinflammatory responses linked to infiltrating immune cells and 2) that human PCLS show a highly diversified tissue resident immune response to viral infection due to previous exposures of the host to this pathogen. These responses are clearly distinct from antibacterial gene profiles. Our data advertise PCLS as a complex and realistic model to study tissue resident immune responses to microbes in a human system.

immunology↗

bioRxiv2023

Human Smc5/6 recognises transcription-generated positive DNA supercoils

Beyond its essential roles in ensuring faithful chromosome segregation and genomic stability, the human Smc5/6 complex acts as an antiviral factor. It binds to and impedes the transcription of extrachromosomal DNA templates; an ability which is lost upon chromosomal DNA integration. How the complex distinguishes among different DNA templates is unknown. Here we show that, in human cells, Smc5/6 preferentially binds to circular rather than linear extrachromosomal DNA. We further show that this binding is unlikely due to differences in the chromatin composition. Instead, the transcriptional process, per se, and more specifically the accumulation of DNA secondary structures known to be substrates for topoisomerases, is responsible for Smc5/6 recruitment. Those findings, in conjunction with our genome-wide Smc5/6 binding analysis showing that Smc5/6 localizes at few but highly transcribe chromosome loci, reveal a previously unforeseen role of Smc5/6 in DNA topology management during transcription.

molecular biology↗

bioRxiv2023

Temporal genome-wide fitness analysis of Mycobacterium marinum during infection reveals genetic requirement for virulence and survival in amoebae and microglial cells

Tuberculosis remains the most pervasive infectious disease and the recent emergence of multiple or even fully drug-resistant strains increases the risk and emphasizes the need for more efficient and better drug treatments. A key feature of mycobacteria pathogenesis is the metabolic switch during infection and expression of virulence genes is often adapted to specific infection conditions. This study aims to identify genes that are involved in the establishment and maintenance of the infection. To answer these questions, we have applied Transposon Sequencing (Tn-Seq) in M. marinum, an unbiased genome-wide strategy that combines saturation insertional mutagenesis and high throughput sequencing. This approach allowed us to precisely identify the localization and relative abundance of insertions in pools of Tn mutants. The essentiality and fitness cost, in terms of growth advantage and disadvantage of over 105 mutants were quantitatively compared between in vitro and different stages of infection in two evolutionary distinct hosts, D. discoideum and BV2 microglial cells. We found that 57% of TA sites in the M. marinum genome were disrupted and that 568 genes (10.2%) are essential for M. marinum, which is comparable to previous Tn-Seq studies on M. tuberculosis. The major pathways involved in the survival of M. marinum during infection of D. discoideum were related to vitamin metabolism, the esx-1 operon, as well as the mce1 operon.

microbiology↗

bioRxiv2022

Cellulosic copper nanoparticles and a hydrogen peroxide-based disinfectant protect Vero E6 cells against infection by viral pseudotyped particles expressing SARS-CoV-2, SARS-CoV or MERS-CoV Spike protein.

Severe acute respiratory syndrome (SARS) is a viral respiratory infection caused by human coronaviruses (HuCoV) that include SARS-CoV-2, SARS-CoV, and Middle East respiratory syndrome coronavirus (MERS-CoV). Although their primary mode of transmission is through contaminated respiratory droplets from infected carriers, the deposition of expelled virus particles onto surface and fomites could contribute to viral transmission. Here, we use replication-deficient murine leukemia virus (MLV) pseudoviral particles expressing SARS-CoV-2, SARS-CoV, or MERS-CoV Spike (S) protein on their surface. These surrogates of native coronavirus counterparts serve as a model to analyze the S-mediated entry into target cells. Carboxymethyl cellulose (CMC) nanofibers that are combined with copper (Cu) exhibit strong antimicrobial properties. S-pseudovirions that are exposed to CMC-Cu nanoparticles (30 s) display a dramatic reduction in their ability to infect target Vero E6 cells, with [~]97% less infectivity as compared to untreated pseudovirions. In contrast, addition of the Cu chelator tetrathiomolybdate protects S- pseudovirions from CMC-Cu-mediated inactivation. When S-pseudovirions were treated with a hydrogen peroxide-based disinfectant (denoted SaberTM) used at 1:16 dilution, their infectivity was dramatically reduced by [~]98%. However, the combined use of SaberTM and CMC-Cu is the most effective approach to restrict infectivity of SARS-CoV-2-S, SARS-CoV-S, and MERS-CoV-S pseudovirions in Vero E6 cell assays. Together, these results show that cellulosic Cu nanoparticles enhance the effectiveness of diluted SaberTM sanitizer, setting up an improved strategy to lower the risk of surface- and fomite-mediated transmission of enveloped respiratory viruses.

microbiology↗

bioRxiv2021

MiR-137 and miR-122, two outer subventricular zone-enriched non-coding RNAs, regulate basal progenitor expansion and neuronal differentiation

Cortical expansion in the primate brain relies on the presence and the spatial enlargement of multiple germinal zones during development and on a prolonged developmental period. In contrast to other mammals, which have two cortical germinal zones, the ventricular zone (VZ) and subventricular zone (SVZ), gyrencephalic species display an additional germinal zone, the outer subventricular zone (OSVZ), which role is to increase the number and types of neurons generated during corticogenesis. How the OSVZ emerged during evolution is poorly understood but recent studies suggest a role for non-coding RNAs, which allow tight regulations of transcriptional programs in time and space during development (Dehay et al. 2015; Arcila et al., 2014). Here, using in vivo functional genetics, single-cell RNA sequencing, live imaging and electrophysiology to assess progenitor and neuronal properties in mice, we identify two ferret and human OSVZ-enriched microRNAs (miR), miR-137 and miR-122, which regulate key cellular features associated with cortical expansion. MiR-137 promotes basal progenitor self-replication and superficial layer neuron fate, while miR-122 slows down neuronal differentiation pace. Together, these findings support a cell-type specific role for miR-mediated transcriptional regulation in cortical expansion.

evolutionary biology↗

bioRxiv2018

Genetic resistance to DEHP-induced transgenerational endocrine disruption

Di(2-ethylhexyl)phthalate (DEHP) interferes with sex hormones signaling pathways (SHP). C57BL/6J mice prenatally exposed to DEHP develop a testicular dysgenesis syndrome (TDS) at adulthood, but similarly-exposed FVB/N mice are not affected. Here we aim to understand the reasons behind this drastic difference that should depend on the genome of the strain. In both backgrounds, pregnant female mice received per os either DEHP or corn oil vehicle and the male filiations were examined. Computer-assisted sperm analysis showed a DEHP-induced decreased sperm count and velocities in C57BL/6J. Sperm RNA sequencing experiments resulted in the identification of the 62 most differentially expressed RNAs. These RNAs, mainly regulated by hormones, produced strain-specific transcriptional responses to prenatal exposure to DEHP; a pool of RNAs was increased in FVB, another pool of RNAs was decreased in C57BL/6J. In FVB/N, analysis of non-synonymous SNP impacting SHP identified rs387782768 and rs387782768 respectively associated with absence of the Forkhead Box A3 (Foxa3) RNA and increased expression of estrogen receptor 1 variant 4 (NM_001302533) RNA. Analysis of the role of SNPs modifying SHP binding sites in function of strain-specific responses to DEHP revealed a DEHP-resistance allele in FVB/N containing an additional FOXA1-3 binding site at rs30973633 and four DEHP-induced beta-defensins (Defb42, Defb30, Defb47 and Defb48). A DEHP-susceptibility allele in C57BL/6J contained five SNPs (rs28279710, rs32977910, rs46648903, rs46677594 and rs48287999) affecting SHP and six genes (Svs2, Svs3b, Svs4, Svs3a, Svs6 and Svs5) epigenetically silenced by DEHP. Finally, targeted experiments confirmed increased methylation in the Svs3ab promoter with decreased SEMG2 persisting across generations, providing a molecular explanation for the transgenerational sperm velocity decrease found in C57BL/6J after DEHP exposure. We conclude that the existence of SNP-dependent mechanisms in inbred mice may confer resistance to transgenerational endocrine disruption.

genomics↗

bioRxiv2018

Single-cell transcriptional dynamics and origins of neuronal diversity in the developing mouse neocortex

During cortical development, distinct subtypes of glutamatergic neurons are sequentially born and differentiate from dynamic populations of progenitors. The neurogenic competence of these progenitors progresses as corticogenesis proceeds; likewise, newborn neurons transit through sequential states as they differentiate. Here, we trace the developmental transcriptional trajectories of successive generations of apical progenitors (APs) and isochronic cohorts of their daughter neurons using parallel single-cell RNA sequencing between embryonic day (E) 12 and E15 in the mouse cerebral cortex. Our results identify the birthdate- and differentiation stage-related transcriptional dynamics at play during corticogenesis. As corticogenesis proceeds, APs transit through embryonic age-dependent molecular states, which are transmitted to their progeny to generate successive initial daughter cell identities. In neurons, essentially conserved post-mitotic differentiation programs are applied onto these distinct AP-derived ground states, allowing temporally-regulated sequential emergence of specialized neuronal cell types. Molecular temporal patterning of sequentially-born daughter neurons by their respective mother cell thus underlies emergence of neuronal diversity in the neocortex.\n\nOne Sentence SummaryDuring corticogenesis, temporally dynamic molecular birthmarks are transmitted from progenitors to their post-mitotic progeny to generate neuronal diversity.

neuroscience↗

bioRxiv2018

Single-cell molecular connectomics of intracortically-projecting neurons

The neocortex is organized into distinct areas, whose interconnectivity underlies sensorimotor transformations and integration1-7. These behaviorally critical functions are mediated by intracortically-projecting neurons (ICPN), which are a heterogeneous population of cells sending axonal branches to distinct cortical areas as well as to subcortical targets8-10. Although population-based11-14 and single-cell15-19 intracortical wiring diagrams are being identified, the transcriptional signatures corresponding to single-cell axonal projections of ICPN to multiple sites remain unknown. To address this question, we developed a high-throughput approach, \"ConnectID\", to link connectome and transcriptome in single neurons. ConnectID combines MAPseq projection mapping17,20 (to identify single-neuron multiplex projections) with single-cell RNA sequencing (to identify corresponding gene expression). Using primary somatosensory cortex (S1) ICPN as proof-of-principle neurons, we identify three cardinal targets: (1) the primary motor cortex (M1), (2) the secondary somatosensory cortex (S2) and (3) subcortical targets (Sub). Using ConnectID, we identify transcriptional modules whose combined activities reflect multiplex projections to these cardinal targets. Based on these findings, we propose that the combinatorial activity of connectivity-defined transcriptional modules serves as a generic molecular mechanism to create diverse axonal projection patterns within and across neuronal cell types.

neuroscience↗