Aug 2026· International Journal of Molecular Sciences· Vol 27, pp. 7612· 0 citations· 45 references
TL;DR
This work investigated the molecular basis of excessive neuronal process formation induced by depletion of NLGN2 using the N1E-115 cell line, an established model of neuronal differentiation, and identified the CCDC88A-G protein-ELMO signaling pathway as a key mediator of excessive neuronal morphogenesis following NLGN2 knockdown.
Abstract
Neuroligin-2 (NLGN2) is a cell adhesion molecule implicated in neurodevelopmental disorders (NDDs), including autism spectrum disorder (ASD) and intellectual disability (ID). While NLGN2 is well known as a postsynaptic organizer of predominantly inhibitory synapses, accumulating evidence suggests that NLGN family proteins are also involved in neuronal morphogenesis during early developmental stages. However, a gap remains in our understanding of how loss of function in NLGN2 potentially leads to abnormal neuronal morphogenesis. Here, we investigated the molecular basis of excessive neuronal process formation induced by depletion of NLGN2 using the N1E-115 cell line, an established model of neuronal differentiation. Clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13-mediated knockdown of NLGN2 promoted neuronal process elongation and neuronal differentiation marker expression. Mechanistically, NLGN2 knockdown resulted in activation of coiled-coil and hook domain-containing protein 88A (CCDC88A, also known as Girdin or GIV), a non-receptor guanine nucleotide exchange factor for heterotrimeric G proteins. Transfection of the regulator of G protein signaling (RGS) domain of RGS3, a negative regulator of G proteins, or the G protein-binding domain of engulfment and cell motility 1 (ELMO1) effectively decreased the excessive process elongation phenotype. Similar effects were observed in primary cortical neurons. Furthermore, these interventions normalized elevated Rac1 activity induced by NLGN2 knockdown. Collectively, our findings identify the CCDC88A-G protein-ELMO signaling pathway as a key mediator of excessive neuronal morphogenesis following NLGN2 knockdown. These results provide valuable insight into the mechanisms by which NLGN2 dysfunction may contribute to abnormal neuronal morphogenesis and suggest potential recovery strategies.
During development, neurexin-2 (NRXN2) is a cell adhesion molecule localized to presynaptic terminals as well as axonal shafts and immature neurites, where it participates in the regulation of neuronal cell morphogenesis. Given its critical role in early neuronal development, NRXN2 is considered a susceptibility gene product for neurodevelopmental disorders (NDDs) such as autism spectrum disorder (ASD) and intellectual disability (ID). However, the intracellular signaling mechanisms linking NRXN2 deficiency to abnormal neuronal cell morphology remain unclear. Herein, we investigated the molecular basis of excessive cell morphogenesis induced by the knockdown of NRXN2 using the N1E-115 cell line, a model of neuronal morphogenesis characterized by neurite outgrowth. Silencing NRXN2 using the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13 system resulted in a marked enhancement of process elongation. Mechanistically, we found that Girdin (also called GIV or CCDC88A), a non-receptor guanine nucleotide exchange factor for heterotrimeric G proteins, can mediate the excessive process length phenotype. Transfection of either the regulator of G protein signaling (RGS) domain of RGS3, a GTPase-activating protein for G proteins, or the G protein-binding domain of engulfment and cell motility 1 (ELMO1) rescued the excessive process formation. Similar results were obtained in primary cortical neurons. In addition, these interventions normalized downstream Rac1 activity in cells. Together, our findings elucidate Girdin signaling as a mediator of excessive neuronal process formation following NRXN2 knockdown, providing mechanistic insight into how the loss of function of NRXN2 leads to aberrant cell morphogenesis at least at the molecular and cellular levels. These results suggest that signaling through Girdin may contribute to the morphological abnormalities associated with NRXN2-related neurodevelopmental disorders.
Hideji Yako, Mikito Takahashi, Mami Akiyama et al.· International Journal of Mol...· 1 citation
Findings indicate that NRXN1α deletion disrupts neural lineage commitment through a multi-layered disruption involving spliceosome dysregulation of chromatin regulatory genes, H3K27me3 redistribution at developmental promoters, and chromatin-level priming into non-neural fates.
A. Ghahramani, Dania Winn, S. Shafiq et al.· bioRxiv· 0 citations
Neurodevelopmental disorders are frequently caused by mutations in pleiotropic kinases, yet downstream effectors driving neuronal pathology remain undefined. Here, we identify the G3BP1-dependent stress granule pathway as the dominant effector of casein kinase 2 (CK2α) in developing neurons, implying that its dysregulation underlies the neurodevelopmental deficits of Okur-Chung neurodevelopmental syndrome (OCNDS). OCNDS-associated CK2α mutations reduce phosphorylation of G3BP1 at serine 149, promoting aberrant phase separation and persistent granules that sequester neuronal mRNAs and suppress local protein synthesis across axonal and dendritic compartments. These phenotypes produce allele-specific deficits in neuronal morphogenesis, synaptic abundance, and network excitability, which are conserved in a knock-in mouse model and in patient-derived iPSC neurons. G3bp1 knockdown rescues translational and morphological phenotypes across all OCNDS alleles, demonstrating that restoring granule homeostasis reverses neuronal pathology. Together, these findings establish OCNDS as a disorder of compartment-specific translational dysregulation driven by impaired CK2α–G3BP1 control of RNA granule homeostasis. Summary OCNDS mutations disrupt CK2α–G3BP1 signaling, causing persistent granules and defective neuronal translation and development.
Fragile X-associated tremor/ataxia syndrome (FXTAS) is an adult-onset neurodegenerative disease associated with carriers of premutation (PM) alleles of the fragile X messenger ribonucleoprotein 1 (FMR1) gene. Expanded CGG repeats in the PM alleles are sufficient to cause cellular stress and toxicity in animal and cellular models of FXTAS. Here, we show that in a Drosophila transgenic model of FXTAS, expanded CGG repeats result in the overexpression of heat shock RNA omega-nuclear (Hsrω-n), a stress-related long non-coding RNA (lncRNA) that dominantly enhances expanded CGG-induced neurotoxicity, whereas its reduction suppresses CGG-induced neurotoxicity. Furthermore, overexpression of Hsrω-n lncRNA concomitantly resulted in a significant enhancement in its association with Hrb87F, the fly ortholog of one of the previously identified CGG repeat RNA-binding proteins, hnRNP A2/B1, and with the imitation switch (ISWI) protein, a chromatin remodeling factor. We extended the findings of Hsrω-n to show that the mammalian stress-related lncRNAs SatIII and Neat1 are elevated in cells expressing expanded CGG repeats. Together, these findings highlight the critical roles of stress-related lncRNAs in CGG repeat-mediated toxicity and support a model in which the total steady-state levels of stress-related lncRNAs increase in the presence of expanded CGG repeats as part of the normal regulatory response to combat stress. However, the effects of chronic expanded CGG repeat expression in post-mitotic neurons may result in altered distribution and/or sequestration of specific RNA-binding proteins or chromatin remodeling factors associated with stress-related lncRNAs. This could contribute to the altered cellular homeostasis and genomic instability associated with FXTAS. Significance Individuals carrying the FMR1 allele with expanded CGG repeats (55–200) in the 5′ untranslated region are predisposed to fragile X–associated tremor/ataxia syndrome (FXTAS), a late-onset neurodegenerative disorder. FXTAS demonstrates incomplete penetrance, suggesting the possible involvement of genetic modifiers in its pathogenesis. We used a transgenic Drosophila FXTAS model to conduct genetic, molecular, and biochemical analyses. Our findings revealed that the expression of expanded CGG repeats aberrantly activates stress-related Hsrω-n lncRNA in Drosophila melanogaster. Overexpression of hsrω-n lncRNA exacerbates CGG-mediated neurodegeneration, whereas its repression alleviates this effect. Extending these findings to mammalian systems, we observed the upregulation of SATIII and Neat I, functional homologs of Hsrω-n lncRNA, in cells expressing expanded CGG repeats. Collectively, these findings identify stress-related lncRNAs as key mediators of CGG-mediated toxicity and underscore their potential as therapeutic targets for FXTAS.
Nadeem Ahmed, M. M. Reshi, A. K. Singh et al.· bioRxiv· 0 citations
It is demonstrated that SYNGAP1 haploinsufficiency disrupts early human brain development and accelerates intrinsic neuronal maturation, with pathogenic mechanisms emerging before synaptogenesis and extending beyond SYNGAP1’s established synaptic role.
Montanna Waters, Lucas Teasdale, Sean Byars et al.· bioRxiv· 0 citations
Neurexins and neuroligins are evolutionarily conserved synaptic adhesion molecules that play essential roles in synapse formation and neural circuit function, with mutations linked to neurodevelopmental disorders such as autism. Here, we combined whole-transcriptome sequencing with phenotypic characterization to define the molecular consequences of neurexin and neuroligin deficiency in Caenorhabditis elegans. Young adult worms carrying allele-specific loss-of-function mutations in nrx-1 (ok1649 and tm1961) or nlg-1 (ok259 and tm474), orthologues for human NRXNs or NLGNs, respectively, were subjected to RNA sequencing and compared with wild-type animals. Mutant strains exhibited impaired growth, altered locomotor activity, increased social aggregation, and reduced ventral nerve cord neuronal integrity. Transcriptomic analysis revealed extensive gene-expression changes, particularly in the nrx-1 (tm1961) allele, with dysregulation of genes involved in cuticle development, neuronal signaling, protein homeostasis, innate immunity, mitochondrial organization, and transcriptional regulation. Gene Ontology and KEGG enrichment analyses identified significant perturbations in developmental, metabolic, stress-response, translational, and synaptic pathways. Together, these findings demonstrate that disruption of neurexin–neuroligin signaling drives transcriptional reprogramming that extends beyond synaptic dysfunction, linking molecular alterations to developmental, behavioral, and neuromorphological abnormalities.
Omamuyouwi M Ijomone, Victor E. Anadu, Toheeb O. Oyerinde et al.· Research Square· 0 citations