Biology subjects

Erdin, S.

Publications and source records attributed to Erdin, S..

11 recordsLinked to original sources

bioRxiv2025

CRISPR-engineered deletion of POGZ alters transcription factor binding at promoters of genes involved in synaptic signaling

One of the seminal discoveries from genetic studies of autism spectrum disorder and related neurodevelopmental disorders (NDD) has been that loss-of-function (LoF) mutations in many genes that impact chromatin and transcriptional regulation confer substantial liability to NDD. Haploinsufficiency of the epigenetic regulator POGZ represents one of the strongest such associations; however, little is known about the direct or indirect regulatory targets of POGZ, or the mechanisms by which loss of this chromatin modifier alters early neuronal development and synaptic functions. Here, we created an allelic series of CRISPR-engineered human induced pluripotent stem cell (hiPSC) clones harboring mono- and biallelic POGZ deletions. In hiPSC-derived neural stem cells (NSC) and Neurogenin 2-induced neurons (iN), POGZ LoF altered the expression of genes associated with critical cellular processes and neuronal functions, including synaptic and intracellular signaling and extracellular matrix organization. Our multiomics profiling also showed altered footprinting of critical transcription factors (e.g., activator protein 1 complexes) that were enriched at promoters of differentially expressed genes associated with synaptic function. To further interrogate the shared molecular changes in neuronal development associated with NDD and POGZ regulation, we compared our results to deletions of the transcription factor MEF2C and the sodium channel gene SCN2A that we generated in these same isogenic iN. These analyses revealed strong enrichment of extracellular matrix and intracellular signaling disruption associated with POGZ and MEF2C deletion, whereas POGZ and SCN2A haploinsufficiency exhibited shared transcriptional effects on gene modules enriched for NDD-associated genes with opposing regulatory effects. Notably, we also observed alterations to synaptic firing rate and neurite extension with biallelic deletions, but not heterozygous lines, suggesting subtle effects in neuronal development associated with haploinsufficiency. Overall, these shared molecular consequences suggest key points of convergence that connect epigenetic regulation to neuronal function in the etiology of neurodevelopmental pathologies.

genomics↗

bioRxiv2025

Aberrant recursive splicing in a human disease locus

Recursive splice sites are rare motifs postulated to facilitate splicing across massive introns and shape isoform diversity, especially for long, brain-expressed genes. The necessity of this unique mechanism remains unsubstantiated, as does the role of recursive splicing (RS) in human disease. From analyses of rare copy number variants (CNVs) from almost one million individuals, we previously identified large, heterozygous deletions eliminating an RS site (RS1) in the first intron of CADM2 that conferred substantial risk for attention deficit hyperactivity disorder (ADHD) and other neurobehavioral traits. CADM2 encodes a neuronally expressed cell adhesion molecule that has repeatedly been associated with ADHD and numerous similar traits. To explore the molecular impact of RS ablation in CADM2, we used CRISPR to model patient deletions and to target a smaller region ([~]500 base pairs) containing RS1 in both human induced neurons (iNs) and rats. Transcriptome analyses in unedited iNs provided a catalog of CADM2 transcripts, including novel transcripts that retained RS exons. Intriguingly, ablating RS1 altered the gradient of RNA abundance across the first intron of CADM2, decreased the level of CADM2 expression, and impacted transcript usage. Decreased CADM2 expression was reflected in reduced exon usage downstream of the RS1 site and global alteration to genes involved in neuronal processes including synapse and axon development. Given the scale of our analyses and the widespread association of CADM2 with neurobehavioral traits, we sought to validate these findings using in vivo models and found that rodent models harboring Cadm2 RS1 deletions exhibited significant changes in relevant behaviors and functional brain connectivity. In summary, our analyses demonstrate a functional role for RS as a noncoding regulatory mechanism in a gene associated with a spectrum of neuropsychiatric and behavioral traits. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/666599v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@171156borg.highwire.dtl.DTLVardef@13553baorg.highwire.dtl.DTLVardef@bee7forg.highwire.dtl.DTLVardef@156f19e_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

genomics↗

bioRxiv2024

TSC2 loss in neural progenitor cells suppresses translation of ASD/NDD-associated transcripts in an mTORC1- and MNK1/2-reversible fashion

Tuberous sclerosis complex (TSC) is an inherited neurodevelopmental disorder (NDD) with frequent manifestations of epilepsy and autism spectrum disorder (ASD). TSC is caused by inactivating mutations in TSC1 or TSC2 tumor suppressor genes, with encoded proteins hamartin (TSC1) and tuberin (TSC2) forming a functional complex inhibiting mechanistic target of rapamycin complex 1 (mTORC1) signaling. This has led to treatment with allosteric mTORC1 inhibitor rapamycin analogs ("rapalogs") for TSC tumors; however, rapalogs are ineffective for treating neurodevelopmental manifestations. mTORC1 signaling controls protein synthesis by regulating formation of the eIF4F complex, with further modulation by MNK1/2 kinases via phosphorylation of the eIF4F subunit eIF4E. While both these pathways modulate translation, comparing their impact on transcriptome-wide mRNA translation, as well as effects of inhibiting these pathways in TSC has not been explored. Here, employing CRISPR-modified, isogenic TSC2 patient-derived neural progenitor cells (NPCs), we have examined transcriptome-wide changes in mRNA translation upon TSC2 loss. Our results reveal dysregulated translation in TSC2-Null NPCs, which significantly overlaps with the translatome from TSC1-Null NPCs. Interestingly, numerous non-monogenic ASD-, NDD-and epilepsy-associated genes identified in patients harboring putative loss-of-function mutations, were translationally suppressed in TSC2-Null NPCs. Importantly, translation of these ASD- and NDD-associated genes was reversed upon inhibition of either mTORC1 or MNK1/2 signaling using RMC-6272 or eFT-508, respectively. This study establishes the importance of mTORC1-eIF4F- and MNK-eIF4E-sensitive mRNA translation in TSC, ASD and other neurodevelopmental disorders laying the groundwork for evaluating drugs in clinical development that target these pathways as a treatment strategy for these disorders.

molecular biology↗

bioRxiv2023

Transcriptome analysis in a humanized mouse model of familial dysautonomia reveals tissue-specific gene expression disruption in the peripheral nervous system.

Familial dysautonomia (FD) is a rare recessive neurodevelopmental disease caused by a splice mutation in the Elongator acetyltransferase complex subunit 1 (ELP1) gene. This mutation results in a tissue-specific reduction of ELP1 protein, with the lowest levels in the central and peripheral nervous systems (CNS and PNS, respectively). FD patients exhibit complex neurological phenotypes due to the loss of sensory and autonomic neurons. Disease symptoms include decreased pain and temperature perception, impaired or absent myotatic reflexes, proprioceptive ataxia, and progressive retinal degeneration. While the involvement of the PNS in FD pathogenesis has been clearly recognized, the underlying mechanisms responsible for the preferential neuronal loss remain unknown. In this study, we aimed to elucidate the molecular mechanisms underlying FD by conducting a comprehensive transcriptome analysis of neuronal tissues from the phenotypic mouse model TgFD9; Elp1{Delta}20/flox. This mouse recapitulates the same tissue-specific ELP1 mis-splicing observed in patients while modeling many of the disease manifestations. Comparison of FD and control transcriptomes from dorsal root ganglion (DRG), trigeminal ganglion (TG), medulla (MED), cortex, and spinal cord (SC) showed significantly more differentially expressed genes (DEGs) in the PNS than the CNS. We then identified genes that were tightly co-expressed and functionally dependent on the level of full-length ELP1 transcript. These genes, defined as ELP1 dose-responsive genes, were combined with the DEGs to generate tissue-specific dysregulated FD signature genes and networks. Within the PNS networks, we observed direct connections between Elp1 and genes involved in tRNA synthesis and genes related to amine metabolism and synaptic signaling. Importantly, transcriptomic dysregulation in PNS tissues exhibited enrichment for neuronal subtype markers associated with peptidergic nociceptors and myelinated sensory neurons, which are known to be affected in FD. In summary, this study has identified critical tissue-specific gene networks underlying the etiology of FD and provides new insights into the molecular basis of the disease.

genomics↗

bioRxiv2022

Tissue and cell-type specific molecular and functional signatures of 16p11.2 reciprocal genomic disorder across mouse brain and human neuronal models

Recurrent deletion and duplication of [~]743 kilobases of unique genomic sequence and segmental duplications at chromosome 16p11.2 underlie a reciprocal genomic disorder (RGD; OMIM 611913 and 614671) associated with neurodevelopmental and psychiatric phenotypes, including intellectual disability, autism spectrum disorder (ASD), and schizophrenia (SCZ). To define molecular alterations associated with the 16p11.2 RGD, we performed transcriptome analyses of mice with reciprocal copy number variants (CNVs) of the syntenic chromosome 7qF3 region and human neuronal models derived from isogenic human induced pluripotent stem cells (hiPSCs) carrying CRISPR-engineered CNVs at 16p11.2. Analysis of differentially expressed genes (DEGs) in mouse cortex, striatum, cerebellum and three non-brain tissues, as well as in human neural stem cells and induced glutamatergic neurons revealed that the strongest and most consistent effects occurred within the CNV sequence, with notable instances of differential expression of genes in the immediate vicinity that could reflect position effect. While differential expression of genes outside of chromosome 16p11.2 was largely region, tissue, and cell type-specific, a small but significant minority of such DEGs was shared between brain regions or human cell types. Gene Ontology (GO) enrichment analyses to identify cellular processes dysregulated due to these CNVs found support in select circumstances for terms related to energy metabolism, RNA metabolism, and translation but did not reveal a single universally affected process. Weighted gene co-expression network analysis identified modules that showed significant correlation with reciprocal or individual CNV genotype and better captured shared effects, indicating that energy metabolism, RNA metabolism, translation and protein targeting were disrupted across all three brain regions. The first two of these processes also emerged in the human neural stem cell (NSC) data. A subset of co-expression modules that correlated with CNV genotype revealed significant enrichments for known neurodevelopmental disorder genes, loss-of-function constrained genes, FMRP targets, and chromatin modifiers. Intriguingly, neuronal differentiation of the hiPSCs revealed that both the deletion and duplication CNV resulted in similar deficits in neurite extension and branching and alterations in electrical activity. Finally, generation of cerebral organoid derivatives indicated that the CNVs reciprocally altered the ratio of excitatory and inhibitory GABAergic neurons generated during in vitro neurodevelopment, consistent with a major mechanistic hypothesis for ASD. Collectively, our data suggest that the 16p11.2 RGD involves disruption of multiple biological processes, with a relative impact that is context-specific. Perturbation of individual and multiple genes within the CNV region will be required to dissect single-gene effects, uncover regulatory interactions, and define how each contributes to abnormal neurodevelopment.

genomics↗

bioRxiv2022

Transcriptional and functional consequences of alterations to MEF2C and its topological organization in neuronal models

Point mutations and structural variants directly disrupting the coding sequence of MEF2C have been associated with a spectrum of neurodevelopmental disorders (NDDs), while recent studies have also implicated altered noncoding regulation of MEF2C expression in NDDs. However, the impact of haploinsufficiency of MEF2C on neurodevelopmental pathways and synaptic processes is not well understood, nor are the complex mechanisms that govern regulation of MEF2C. To explore the transcriptional and functional changes associated with coding and noncoding structural variants, we generated an allelic series of 204 isogenic iPSC-derived neuronal cell lines harboring CRISPR-engineered mutations that directly delete predominant isoforms of MEF2C, as well as deletions to the boundaries of topologically associating domains (TADs) and chromatin loops encompassing MEF2C. We then performed systematic profiling of mutation-specific alterations to transcriptional signatures, regulatory interactions, chromatin contacts, and electrophysiological effects. Our analyses reveal that direct deletion of MEF2C causes differential expression of genes enriched for neurodevelopmental and synaptic-associated pathways, accompanied by a significant reduction in synaptic firing and synchrony in neurons. By contrast, we observe robust buffering against MEF2C regulatory disruption upon deletion of a distal 5q14.3 TAD and loop boundary; however, homozygous loss of proximal loop boundary resulted in significant down-regulation of MEF2C expression and significantly reduced electrophysiological activity that was comparable to direct MEF2C disruption. Collectively, our findings demonstrate the functional impact of MEF2C haploinsufficiency in human-derived neural models and highlight the complex interactions of gene regulation and chromatin topology that challenge a priori regulatory predictions of structural variant disruption to three-dimensional genome organization.

genomics↗

bioRxiv2021

Dystonia-specific mutations in THAP1 alter transcription of genes associated with neurodevelopment and myelin

Dystonia is a neurologic disorder associated with an increasingly large number of variants in many genes, resulting in characteristic disturbances in volitional movement. Dissecting the relationships between these mutations and their functional outcomes is a critical step in understanding the key pathways that drive dystonia pathogenesis. Here we established a pipeline for characterizing an allelic series of dystonia-specific mutations in isogenic induced pluripotent stem cells (iPSCs). We used this strategy to investigate the molecular consequences of variation in THAP1, which encodes a transcription factor that has been linked to neural differentiation. Multiple pathogenic mutations that have been associated with dystonia cluster within distinct THAP1 functional domains and are predicted to alter its DNA binding properties and/or protein interactions differently, yet the relative impact of these varied changes on molecular signatures and neural deficits is unclear. To determine the effects of these mutations on THAP1 transcriptional activity, we engineered an allelic series of eight mutations in a common iPSC background and differentiated these lines into a panel of near-isogenic neural stem cells (n = 94 lines). Transcriptome profiling of these neural derivatives followed by joint analysis of the most robust individual signatures across mutations identified a convergent pattern of dysregulated genes functionally related to neurodevelopment, lysosomal lipid metabolism, and myelin. Based on these observations, we examined mice bearing Thap1-disruptive alleles and detected significant changes in myelin gene expression and reduction of myelin structural integrity relative to tissue from control mice. These results suggest that deficits in neurodevelopment and myelination are common consequences of dystonia-associated THAP1 mutations and highlight the potential role of neuron-glial interactions in the pathogenesis of dystonia.

genetics↗

bioRxiv2021

Developmental regulation of neuronal gene expression by Elongator complex protein 1 dosage.

Elongator is a highly conserved protein complex required for transcriptional elongation, intracellular transport and translation. Elongator complex protein 1 (ELP1) is the scaffolding protein of Elongator and is essential for its assembly and stability. Familial dysautonomia (FD), a hereditary sensory and autonomic neuropathy, is caused by a mutation in ELP1 that lead to a tissue-specific reduction of ELP1 protein. Our work to generate a phenotypic mouse model for FD led to the discovery that homozygous deletion of the mouse Elp1 gene leads to embryonic lethality prior to mid-gestation. Given that FD is caused by a reduction, not loss, of ELP1, we generated two new mouse models by introducing different copy numbers of the human FD ELP1 transgene into the Elp1 knockout mouse (Elp1-/-) and observed that human ELP1 expression rescues embryonic development in a dose dependent manner. We then conducted a comprehensive transcriptome analysis in mouse embryos to identify genes and pathways whose expression correlates with the amount of ELP1. We found that ELP1 is essential for the expression of genes responsible for the formation and development of the nervous system. Further, gene length analysis of the differentially expressed genes showed that the loss of Elp1 mainly impacts the expression of long genes and that by gradually restoring Elongator their expression is progressively rescued. Finally, through evaluation of co-expression modules, we identified gene sets with unique expression patterns that depended on ELP1 expression. Overall, this study highlights the crucial role of ELP1 during early embryonic neuronal development and reveals gene networks and biological pathways that are regulated by Elongator.

developmental biology↗

bioRxiv2020

CHD8 Suppression Impacts on Histone H3 Lysine 36 Trimethylation and Alters RNA Alternative Splicing.

Disruptive mutations in the chromodomain helicase DNA binding protein 8 (CHD8) have been recurrently associated with Autism Spectrum Disorders (ASD). In normal cellular physiology, CHD8 co-purifies with MLL1 and MOF transcriptional activation complex, with elongating RNAPII and directly binds to DNA promoters and enhancers regions, thus a regulatory role in transcriptional initiation and elongation could be postulated. Here we investigated how chromatin landscape reacts to CHD8 suppression by analyzing a panel of histone modifications in induced pluripotent stem cell-derived neural progenitors. We interrogated transcriptionally active and repressed regions, as well as active and poised enhancers. CHD8 suppression led to significant reduction (47.82%) in histone H3K36me3 peaks at gene bodies, particularly impacting on transcriptional elongation chromatin states. H3K36me3 reduction specifically affects highly expressed, CHD8-bound genes. Histone H3K36me3 reduction associated to CHD8-suppression does not functionally impact on global transcriptional levels, but correlated with altered alternative splicing patterns of [~] 2000 protein coding genes implicated in "RNA splicing", "mitotic cell cycle phase transition" and "mRNA processing", especially affecting alternative first exon and exon skipping events. In summary, our results point toward broad molecular consequences of CHD8 suppression, implicating altered histone deposition/maintenance and RNA processing regulation as important regulatory processes in ASD.

genetics↗

bioRxiv2020

A deep learning approach to identify new gene targets of a novel therapeutic for human splicing disorders

Pre-mRNA splicing is a key control point in human gene expression. Disturbances in splicing due to mutation or aberrant splicing regulatory networks lead to dysregulated protein expression and contribute to a substantial fraction of human disease. Several classes of active and selective splicing modulator compounds have been recently identified, thus proving that pre-mRNA splicing is a viable target for therapy. We describe herein the identification of BPN-15477, a novel splicing modulator compound, that restores correct splicing of exon 20 in the Elongator complex protein 1 (ELP1) gene carrying the major IVS20+6T>C mutation responsible for familial dysautonomia. We then developed a machine learning approach to evaluate the therapeutic potential of BPN-15477 to correct splicing in other human genetic diseases. Using transcriptome sequencing from compound-treated fibroblast cells, we identified treatment responsive sequence signatures, the majority of which center at the 5 splice site of exons whose inclusion or exclusion is modulated by SMC treatment. We then leveraged this model to identify 155 human disease genes that harbor ClinVar mutations predicted to alter pre-mRNA splicing as potential targets for BPN-15477 treatment. Using in vitro splicing assays, we validated representative predictions by demonstrating successful correction of splicing defects caused by mutations in genes responsible for cystic fibrosis (CFTR), cholesterol ester storage disease (LIPA), Lynch syndrome (MLH1) and familial frontotemporal dementia (MAPT). Our study shows that deep learning techniques can identify a complex set of sequence signatures and predict response to pharmacological modulation, strongly supporting the use of in silico approaches to expand the therapeutic potential of drugs that modulate splicing.

bioinformatics↗

bioRxiv2018

Hypomorphic mutation of the mouse Huntingtons disease gene orthologue

Rare individuals with hypomorphic inactivating mutations in the Huntingtons Disease (HD) gene (HTT), identified by CAG repeat expansion in the eponymous neurodegenerative disorder, exhibit variable abnormalities that imply HTT essential roles during organ development. Here we report phenotypes produced when increasingly severe hypomorphic mutations in Htt, the murine HTT orthologue (in HdhneoQ20, HdhneoQ50, HdhneoQ111 mice), were placed over a null allele (Hdhex4/5). The most severe hypomorphic allele failed to rescue null lethality at gastrulation, while the intermediate alleles yielded perinatal lethality and a variety of fetal abnormalities affecting body size, skin, skeletal and ear formation, and transient defects in hematopoiesis. Comparative molecular analysis of wild-type and Htt-null retinoic acid-differentiated cells revealed gene network dysregulation associated with organ development and proposed polycomb repressive complexes and miRNAs as molecular mediators. Together these findings demonstrate that the HD gene acts both pre- and post-gastrulation and possibly suggest pleiotropic consequences of HTT-lowering therapeutic strategies.\n\nAuthor SummaryThe HTT gene product mutated in Huntingtons Disease (HD) has essential roles during normal organism development, however, still not fully predictable are the functional consequences of its partial inactivation. Our genetic study provides a comprehensive effects description of progressively stronger suppression of Htt gene, the murine HTT counterpart. The most severe Htt reduction leads to embryo lethality, while intermediate Htt dosages yield a variety of developmental abnormalities affecting body size, skin, skeletal and ear formation, and hematopoiesis. Complementing molecular analysis in differentiating cells depleted of a functional Htt gene further elucidates genes networks dysregulated during organ development and proposes chromatin regulators and short non-coding RNAs as key molecular mediators. Together these findings demonstrate that the HD gene acts both at early and later stages of development, thus possibly suggesting long-term consequences associated to the newest HD therapeutic strategies aimed at lowering the HTT gene product.

genetics↗