Biology subjects

Nicolis, S. K.

Publications and source records attributed to Nicolis, S. K..

6 recordsLinked to original sources

bioRxiv2026

An improved CRISPR-LICHT screen of 120 Tdark genes illuminates new biologically relevant SOX2 targets in human brain organoids

SOX2 is a transcription factor and central regulator of neural progenitor identity, yet the downstream molecular networks through which it supports human brain development remain incompletely characterized. Here, we applied CRISPR-lineage tracing at cellular resolution in heterogeneous tissue (CRISPR-LICHT) to human cerebral organoids to functionally interrogate SOX2-dependent regulatory networks. Using a SOX2-focused library targeting SOX2 together with conserved neurodevelopmental regulators expressed in neural progenitor cells, we showed that CRISPR-LICHT detects reproducible lineage- depletion phenotypes across genes already known to be involved in progenitor maintenance, fate specification, glial differentiation and neuronal maturation. This extends the application of CRISPR-LICHT beyond the severe neural progenitor phenotypes previously characterized for microcephaly-associated genes and demonstrates its ability to detect perturbation effects across a broader set of neurodevelopmental regulators, involving SOX2 targets previously defined in mouse and now open to further understanding in a human model setting. We further screened 120 evolutionarily conserved, poorly characterized SOX2-dependent genes classified as Tdark within the Illuminating the Druggable Genome framework and identified 28 candidate hits whose disruption impaired neural progenitor lineage expansion and/or survival. These candidates included proteins associated with cytoskeletal organization, membrane dynamics and intracellular architecture, suggesting that SOX2-dependent developmental programmes may engage cellular processes beyond transcriptional regulation. Individual downregulation of CDC42EP4, a poorly characterized CDC42 effector linked to cytoskeletal organization, revealed altered distribution of targeted progenitors between ventricular- and subventricular- like compartments, with changes consistent with altered progenitor-state progression. Together, these findings expand the functional landscape of SOX2-dependent regulation in human neural progenitors and identify cellular architecture as a potential component of SOX2-associated developmental programs.

developmental biology↗

bioRxiv2023

SOX2 and NR2F1 coordinate the gene expression program of the early postnatal visual thalamus

The thalamic dorsolateral geniculate nucleus, (dLGN) receives visual input from the retina via the optic nerve, and projects to the cortical visual area, where eye-derived signals are elaborated. The transcription factors SOX2 and NR2F1 are directly involved in the differentiation of dLGN neurons, based on mouse work and patient mutations leading to vision defects. However, whether they regulate each other, or control common targets is still unclear. By RNA-seq analysis of neonatal dLGN from thalamo-specific Sox2 and Nr2f1 mouse mutants, we found a striking overlap of deregulated genes. Among them, VGF, a cytokine transported along thalamic-cortical axons is strongly downregulated in both mutants. CUT&RUN analysis of SOX2 binding in dLGN chromatin identified a binding pattern characteristic of the dLGN. Collectively, the SOX2 and NR2F1- coregulated genes, and cognate SOX2 binding sites, contribute as a basis to understand the gene regulatory network driving the differentiation and connectivity of thalamic neurons.

developmental biology↗

bioRxiv2022

Bridging between mouse and human enhancer-promoter long-range interactions in neural stem cells, to understand enhancer function in neurodevelopmental disease

Non-coding variation in complex human disease has been well established by genome-wide association studies, and it is thought to involve regulatory elements, such as enhancers, whose variation affects the expression of the gene responsible for the disease. The regulatory elements often lie far from the gene they regulate, or within introns of genes differing from the regulated gene, making it difficult to identify the gene whose function is affected by a given enhancer variation. Enhancers are connected to their target gene promoters via long-range physical interactions (loops). In our study, we re-mapped, onto the human genome, more than 10,000 enhancers connected to promoters via long-range interactions, that we had previously identified in mouse brain-derived neural stem cells by RNApolII-ChIA-PET analysis, coupled to ChIP-seq mapping of DNA/chromatin regions carrying epigenetic enhancer marks. These interactions are thought to be functionally relevant. We discovered, in the human genome, thousands of DNA regions syntenic with the interacting mouse DNA regions (enhancers+connected promoters). We further annotated these human regions regarding their overlap with sequence variants (single nucleotide polymorphisms, SNPs; copy number variants, CNVs), that were previously associated with neurodevelopmental disease in human. We document various cases in which the genetic variant, associated in human to neurodevelopmental disease, affects an enhancer involved in long-range interactions: SNPs, previously identified by genome-wide association studies to be associated with schizophrenia, bipolar disorder, and intelligence, are located within our human syntenic enhancers, and alter transcription factors recognition sites. Similarly, CNVs associated to autism spectrum disease and other neurodevelopmental disorders overlap with our human syntenic enhancers. Some of these enhancers are connected (in mouse) to homologs of genes already associated to the human disease, strengthening the hypothesis that the gene is indeed involved in the disease. Other enhancers are connected to genes not previously associated with the disease, pointing to their possible pathogenetic involvement. Our observations provide a resource for further exploration of neural disease, in parallel with the now widespread genome-wide identification of DNA variants in patients with neural disease.

neuroscience↗

bioRxiv2020

An early Sox2-dependent gene expression program required for hippocampal dentate gyrus development

The hippocampus is a brain area central for cognition. Mutations in the human SOX2 transcription factor cause neurodevelopmental defects, leading to intellectual disability and seizures, together with hippocampal dysplasia. We generated an allelic series of Sox2 conditional mutations in mouse, deleting Sox2 at different developmental stages. Late Sox2 deletion (from E11.5, via Nestin-Cre) affects only postnatal hippocampal development; earlier deletion (from E10.5, Emx1-Cre) significantly reduces the dentate gyrus, and the earliest deletion (from E9.5, FoxG1-Cre) causes drastic abnormalities, with almost complete absence of the dentate gyrus. We identify a set of functionally interconnected genes (Gli3, Wnt3a, Cxcr4, p73 and Tbr2), known to play essential roles in hippocampal embryogenesis, which are downregulated in early Sox2 mutants, and (Gli3 and Cxcr4) directly controlled by SOX2; their downregulation provides plausible molecular mechanisms contributing to the defect. Electrophysiological studies of the Emx1Cre mouse model reveal altered excitatory transmission in CA1 and CA3 regions.

developmental biology↗

bioRxiv2020

Sox2 controls neural stem cell self-renewal through a Fos-centered gene regulatory network

The Sox2 transcription factor is necessary for the long-term self-renewal of neural stem cells (NSC). Its mechanism of action is still poorly defined. To identify molecules regulated by Sox2, and acting in mouse NSC maintenance, we transduced, into Sox2-deleted NSC, genes whose expression is strongly downregulated following Sox2 loss (Fos, Jun, Egr2), individually or in combination. Fos alone rescued long-term proliferation, as shown by in vitro cell growth and clonal analysis. Further, pharmacological inhibition of the FOS/JUN AP1 complex binding to its targets, decreased cell proliferation and expression of the putative target Suppressor of cytokine signaling 3 (Socs3). Additionally, Fos requirement for efficient long-term proliferation was demonstrated by the reduction of NSC clones capable of long-term expansion following CRISPR/Cas9-mediated Fos inactivation. Previous work showed that the Socs3 gene is strongly downregulated following Sox2 deletion, and its reexpression by lentiviral transduction rescues long-term NSC proliferation. Fos appears to be an upstream regulator of Socs3, possibly together with Jun and Egr2; indeed, Sox2 reexpression in Sox2-deleted NSC progressively activates both Fos and Socs3 expression; in turn, Fos transduction activates Socs3 expression. Based on available SOX2 ChIPseq and ChIA-PET data, we propose a model whereby Sox2 is a direct activator of both Socs3 and Fos, as well as possibly Jun and Egr2; further, we provide direct evidence for FOS and JUN binding on Socs3 promoter, suggesting direct transcriptional regulation. These results provide the basis for developing a model of a network of interactions, regulating critical effectors of NSC proliferation and long-term maintenance. Significance statementProliferation and maintenance of NSC are essential during normal brain development, and, postnatally, for the maintenance of hippocampal function and memory until advanced age. Little is known about the molecular mechanisms that maintain the critical aspects of NSC biology (quiescence and proliferation) in postnatal age. Our work provides a methodology, transduction of genes deregulated following Sox2 deletion, that allows to test many candidate genes for their ability to sustain NSC proliferation. In principle, this may have interesting implications for identifying targets for pharmacological manipulations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=186 SRC="FIGDIR/small/995621v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@e9241forg.highwire.dtl.DTLVardef@4276deorg.highwire.dtl.DTLVardef@18a2058org.highwire.dtl.DTLVardef@151b2d2_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2020

Sox2-dependent maintenance of mouse oligodendroglioma involves the Sox2-mediated downregulation of Cdkn2b, Ebf1, Zfp423 and Hey2

Cancer stem cells (CSC) are essential for tumorigenesis. The transcription factor Sox2 is overexpressed in brain tumors. In gliomas, Sox2 is essential to maintain CSC. In mouse high-grade glioma pHGG, Sox2 deletion causes cell proliferation arrest and inability to reform tumors in vivo; 134 genes are significantly derepressed. To identify genes mediating the effects of Sox2 deletion, we overexpressed into pHGG cells nine among the most derepressed genes, and identified four genes, Cdkn2b, Ebf1, Zfp423 and Hey2, that strongly reduced cell proliferation in vitro and brain tumorigenesis in vivo. CRISPR/Cas9 mutagenesis, or pharmacological inactivation, of each of these genes, individually, showed that their activity is essential for the proliferation arrest caused by Sox2 deletion. These Sox2-inhibited antioncogenes also inhibited clonogenicity in primary human glioblastoma-derived cancer stem-like cell lines. These experiments identify critical anti-oncogenic factors whose inhibition by Sox2 is involved in CSC maintenance, defining new potential therapeutic targets for gliomas. Table of Contents Image O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=176 SRC="FIGDIR/small/976811v1_ufig1.gif" ALT="Figure 1"> View larger version (61K): org.highwire.dtl.DTLVardef@1c42c8forg.highwire.dtl.DTLVardef@496b73org.highwire.dtl.DTLVardef@2d4c31org.highwire.dtl.DTLVardef@1e5367e_HPS_FORMAT_FIGEXP M_FIG C_FIG Main PointsSox2 maintains glioma tumorigenicity by repressing the antioncogenic activity of a regulatory network involving the Ebf1, Hey2, Cdkn2b and Zfp423 genes. Mutation of these genes prevents the cell proliferation arrest of Sox2-deleted glioma cells.

cancer biology↗