Biology subjects

Vertesy, A.

Publications and source records attributed to Vertesy, A..

9 recordsLinked to original sources

bioRxiv2026

Connectomics by Sequencing Reveals Self-Organizing Principles of Neuronal Networks in Cerebral Organoids

Functional neural networks emerge as developing neurons form synaptic connections. Revealing how these connections are organized and perturbed in disease requires linking single-neuron connectivity to molecular state across thousands of networks, which is challenging with current methods. We developed COSCO (Connectomics by Sequencing in Cerebral Organoids), combining barcoded monosynaptic rabies tracing with single-cell RNA sequencing to reconstruct over 10,000 networks across organoids. We demonstrate that organoid circuitry self-organizes into stereotyped network motifs governed by cell-intrinsic wiring rules aligning with features of mouse cortical connectivity. Projecting network topology onto transcriptomic space reveals that wiring correlates with cell-adhesion and synaptic gene expression programs. In a Tuberous Sclerosis Complex model, pathological hyperexcitability coincides with topological rewiring: mutant networks become more heterogeneous, driven by the transcriptional distribution of upper layer excitatory cell states. This work establishes cerebral organoids as a model to study self-organization of human connectivity, and its disruption in neurodevelopmental disease. O_FIG O_LINKSMALLFIG WIDTH=199 HEIGHT=200 SRC="FIGDIR/small/755792v1_ufig1.gif" ALT="Figure 1"> View larger version (66K): org.highwire.dtl.DTLVardef@50ade8org.highwire.dtl.DTLVardef@c97320org.highwire.dtl.DTLVardef@1a53a47org.highwire.dtl.DTLVardef@9520ad_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Constitutively active RAS prolongs Cdc42 signalling while attenuating MAPK signalling during fission yeast mating

The small GTPase RAS is a signalling hub activating multiple pathways, which may respond differently to a constitutively active RAS mutation. We explored this issue using fission yeast, where RAS-mediated pheromone signalling (PS) activates the MAPKSpk1 and Cdc42 pathways. We observed that in cells with the yeast RAS mutant, ras1.G17V, the MAPKSpk1 activation was attenuated similarly to that in wildtype cells, whereas Cdc42 activation was prolonged. We built a mathematical model implementing PS negative-feedback circuits and competition between the two Ras1 effectors, MAPKKKByr2 and Cdc42-GEFScd1. The model robustly predicted the MAPKSpk1 activation dynamics of an additional 20 PS mutants. In support of the model, a recombinant Cdc42-GEFScd1 competed with MAPKKKByr2 for Ras1 binding and overexpression of the Ras binding domain of either Cdc42-GEFScd1 or MAPKKKByr2 inhibited both downstream pathways. Our study has established that the constitutively active RAS signalling propagates differently to downstream pathways where the system prevents MAPK overactivation.

cell biology↗

bioRxiv2022

Single-cell brain organoid screening identifies developmental defects in autism

Development of the human brain involves processes that are not seen in many other species, but can contribute to neurodevelopmental disorders (1-4). Cerebral organoids can be used to investigate neurodevelopmental disorders in a human context but are limited by variability and low throughput. We have developed the CRISPR-human organoids-scRNA-seq (CHOOSE) system that utilizes verified pairs of gRNAs, inducible CRISPR/Cas9-based genetic disruption, and single-cell transcriptomics for pooled loss-of-function screening in mosaic organoids. Genetic perturbations of 36 high-risk autism spectrum disorder (ASD) genes related to transcriptional regulation allowed us to identify their effects on cell fate determination and discover developmental stages susceptible to ASD gene perturbations. We construct a developmental gene regulatory network (GRN) of cerebral organoids from single-cell multiomic data including transcriptome and chromatin modalities and identify ASD-associated and perturbation-enriched regulatory modules. We show that perturbing members of the BAF chromatin remodeling complex leads to an expanded population of ventral telencephalon progenitors. Specifically, the BAF subunit ARID1B controls the fate transitions of progenitors to oligodendrocyte precursor cells and interneurons, which we confirmed in patient-specific induced pluripotent stem cell (iPSC) derived organoids. Our study paves the way for phenotypically characterizing disease susceptibility genes in human organoid models with cell type, developmental trajectory, and gene regulatory network readouts.

developmental biology↗

bioRxiv2022

Cellular stress in brain organoids is limited to a distinct and bioinformatically removable subpopulation.

Organoids enable disease modeling in complex and structured human tissue, in vitro. Like most 3D models, they lack sufficient oxygen supply, leading to cellular stress. These negative effects are particularly prominent in complex models, like brain organoids, where they can prevent proper lineage commitment. Here, we analyze brain organoid and fetal single cell RNA sequencing (scRNAseq) data from published and new datasets totaling over 190,000 cells. We describe a unique stress signature found in all organoid samples, but not in fetal samples. We demonstrate that cell stress is limited to a defined organoid cell population, and present Gruffi, an algorithm that uses granular functional filtering to identify and remove stressed cells from any organoid scRNAseq dataset in an unbiased manner. Our data show that adverse effects of cell stress can be corrected by bioinformatic analysis, improving developmental trajectories and resemblance to fetal data.

neuroscience↗

bioRxiv2020

Cerebral organoid model reveals excessive proliferation of human caudal late interneuron progenitors in Tuberous Sclerosis Complex

Although the intricate and prolonged development of the human brain critically distinguishes it from other mammals1, our current understanding of neurodevelopmental diseases is largely based on work using animal models. Recent studies revealed that neural progenitors in the human brain are profoundly different from those found in rodent animal models2-5. Moreover, post-mortem studies revealed extensive migration of interneurons into the late-gestational and post-natal human prefrontal cortex that does not occur in rodents6. Here, we use cerebral organoids to show that overproduction of mid-gestational human interneurons causes Tuberous Sclerosis Complex (TSC), a severe neuro-developmental disorder associated with mutations in TSC1 and TSC2. We identify a previously uncharacterized population of caudal late interneuron progenitors, the CLIP-cells. In organoids derived from patients carrying heterozygous TSC2 mutations, dysregulation of mTOR signaling leads to CLIP-cell over-proliferation and formation of cortical tubers and subependymal tumors. Surprisingly, second-hit events resulting from copy-neutral loss-of-heterozygosity (cnLOH) are not causative for but occur during the progression of tumor lesions. Instead, EGFR signaling is required for tumor proliferation, opening up a promising approach to treat TSC lesions. Our study demonstrates that the analysis of developmental disorders in organoid models can lead to fundamental insights into human brain development and neuropsychiatric disorders.

neuroscience↗

bioRxiv2019

Dynamics of Meiotic Sex Chromosome Inactivation and Pachytene Activation in Mice Spermatogenesis

During germ cell development, cells undergo a drastic switch from mitosis to meiosis to form haploid germ cells. Sequencing and computational technologies now allow studying development at the single-cell level. Here we developed a multiplexed trajectory reconstruction to create a high-resolution developmental map of spermatogonia and prophase-I spermatocytes from testes of a Dazl-GFP reporter mouse. We identified three main transitions in the meiotic prophase-I: meiotic entry, the meiotic sex chromosome inactivation (MSCI), and concomitant pachytene activation. We validated the key features of these transitions in vivo using single molecule FISH. Focusing on MSCI, we found that 34% of sex chromosomal genes are induced shortly before MSCI, that silencing time is diverse and correlates with specific gene functions. These highlight a previously underappreciated level of regulation of MSCI. Finally, we found that spermatozoal genes in pachytene are activated in a temporal pattern reflecting the future anatomic and functional order of the sperm cell. Altogether we highlighted how precise and sequential changes in gene expression regulate cellular states in meiotic prophase-I.

genomics↗

bioRxiv2018

Spindle checkpoint silencing at kinetochores with submaximal microtubule occupancy

The spindle assembly checkpoint (SAC) ensures proper chromosome segregation by monitoring kinetochore-microtubule interactions. SAC proteins are shed from kinetochores once stable attachments are achieved. Human kinetochores consist of hundreds of SAC protein recruitment modules and bind up to 20 microtubules, raising the question how the SAC responds to intermediate attachment states. We show that the RZZS-MAD1/2 module of the SAC is removed from kinetochores at low microtubule occupancy and remains absent at higher occupancies, while the BUB1/R1 module is retained at substantial levels irrespective of attachment states. Artificially tuning the affinity of kinetochores for microtubules further shows that ~50% occupancy is sufficient to shed MAD2 and silence the SAC. Kinetochores thus responds as a single unit to shut down SAC signaling at submaximal occupancy states, but retains one SAC module. This may ensure continued SAC silencing on kinetochores with fluctuating occupancy states while maintaining the ability for fast SAC re-activation.

cell biology↗

bioRxiv2018

Constitutively active RAS in S. pombe causes persistent Cdc42 signalling but only transient MAPK activation

The small GTPase RAS is a signalling hub for many pathways and oncogenic human RAS mutations are assumed to over-activate all of its downstream pathways. We tested this assumption in fission yeast, where, RAS-mediated pheromone signalling (PS) activates the MAPKSpk1 and Cdc42 pathways. Unexpectedly, we found that constitutively active Ras1.G17V induced immediate but only transient MAPKSpk1 activation, whilst Cdc42 activation persisted. Immediate but transient MAPKSpk1 activation was also seen in the deletion mutant of Cdc42-GEFScd1, a Cdc42 activator. We built a mathematical model using PS negative-feedback circuits and competition between the two Ras1 effectors, MAPKKKByr2 and Cdc42-GEFScd1. The model robustly predicted the MAPKSpk1 activation dynamics of an additional 21 PS mutants. Supporting the model, we showed that a recombinant Cdc42-GEFScd1 fragment competes with MAPKKKByr2 for Ras1 binding. Our study has established a concept that the constitutively active RAS propagates differently to downstream pathways where the system prevents MAPK overactivation. HighlightsO_LIConstitutively active Ras1.GV prolongs Cdc42 activation in S. pombe pheromone signalling C_LIO_LIRas1.GV results in an immediate but only transient MAPKSpk1 activation C_LIO_LIThe RAS effector pathways MAPKSpk1 and Cdc42 compete with each other for active Ras1 C_LIO_LIPredictive modelling explains MAPKSpk1 activation dynamics in 24 signaling-mutants C_LI eTOC BlurbS. pombe Ras1 activates the MAPKSpk1 and Cdc42 pathways. Kelsall et al. report that the constitutively active Ras1.G17V mutation, which causes morphological anomalies, induces prolonged Cdc42 activation but only a transient MAPKSpk1 activation followed by attenuation. Mathematical modelling and biochemical data suggest a competition between the MAPKSpk1 and Cdc42 pathways for active Ras1.

molecular biology↗

bioRxiv2018

Spatial transcriptomics of C. elegans males and hermaphrodites identifies novel fertility genes

To advance our understanding of the genetic programs that drive cell and tissue specialization, it is necessary to obtain a comprehensive overview of gene expression patterns. Here, we have used RNA tomography to generate the first high-resolution, anteroposterior gene expression maps of C. elegans males and hermaphrodites. To explore these maps, we have developed computational methods for discovering region and tissue-specific genes. Moreover, by combining pattern-based analysis with differential gene expression analysis, we have found extensive sex-specific gene expression differences in the germline and sperm. We have also identified genes that are specifically expressed in the male reproductive tract, including a group of uncharacterized genes that encode small secreted proteins that are required for male fertility. We conclude that spatial gene expression maps provide a powerful resource for identifying novel tissue-specific gene functions in C. elegans. Importantly, we found that expression maps from different animals can be precisely aligned, which opens up new possibilities for transcriptome-wide comparisons of gene expression patterns.

genomics↗