It is demonstrated that CHAMP1 mutations disrupt multiple aspects of neuronal development, including homologous recombination (HR) dysfunction in NPCs, membrane excitability, ion channel function, and synaptic connectivity.
Abstract
Mutations in chromosome alignment maintaining phosphoprotein 1 (CHAMP1) have been linked to neurodevelopmental disorders characterized by intellectual disability, developmental delay, and autism spectrum disorder. However, the cellular and electrophysiological mechanisms by which CHAMP1 mutations disrupt human neuronal development remain poorly understood. In the present study, we used patient-derived induced pluripotent stem cells (iPSCs) carrying two pathogenic CHAMP1 mutations and generated neural progenitor cells (NPCs) and excitatory neurons to investigate the effects of each mutation on neuronal maturation and function, DNA repair, and gene expression. Proliferative capacity declines with CHAMP1 dosage, while DNA repair dysfunction is allele-specific. Whole-cell patch-clamp electrophysiology revealed that CHAMP1 mutant neurons exhibit significant alterations in intrinsic membrane properties during early developmental stages, including depolarized resting membrane potential, reduced action potential firing, and impaired waveform kinetics. These functional deficits were accompanied by reduced sodium and potassium current densities, suggesting impaired ion channel accumulation during neuronal maturation. Furthermore, recordings of spontaneous excitatory postsynaptic currents indicated altered synaptic activity and reduced proportions of synaptically active neurons. Morphological analyses showed that CHAMP1-deficient neurons exhibit impaired neurite outgrowth and branching, supporting a defect in neuronal maturation. Single-nucleus transcriptomic profiling further revealed delayed developmental trajectories and mutation-specific dysregulation of synaptic gene programs enriched for autism, ADHD, and epilepsy risk genes. Together, these findings demonstrate that CHAMP1 mutations disrupt multiple aspects of neuronal development, including homologous recombination (HR) dysfunction in NPCs, membrane excitability, ion channel function, and synaptic connectivity. Our results provide insights into the neurobiological consequences of CHAMP1 mutations and establish patient-derived neurons as a platform to investigate cellular pathophysiology and potential therapeutic strategies for CHAMP1-associated neurodevelopmental disorders.
Ataxia with oculomotor apraxia type 1 (AOA1), caused by mutations in the DNA repair protein aprataxin (APTX), leads to progressive neurodegeneration. In this study, we established an AOA1 patient-derived induced pluripotent stem cell (iPSC) and a neuronal differentiation model. We demonstrated that AOA1-derived neurons exhibit neurite morphology and maturation defects correlated with the accumulation of DNA single-strand break (SSB) signals. AOA1-derived neurons showed greater DNA-damage and PAR signals together with lower protein-normalized NAD(H) and ATP after genotoxic exposure. These parallel changes are consistent with metabolic stress but do not establish a PARP1-dependent causal pathway. Bulk transcriptomic profiling and alternative splicing (AS) analysis further revealed widespread transcriptomic dysregulation and altered AS events, particularly enriched in neuronal genes essential for neurite development and synaptic function. Collectively, our findings identify neuronal differentiation, DNA-damage, metabolic, and transcriptomic differences in AOA1 patient-derived cultures and motivate composition-controlled and rescue-based studies of APTX function.
Zirui Chen, Yihua Huang, Xinyue Hu et al.· Neurobiology of Disease· 0 citations
These findings establish DLG4 haploinsufficiency as a shared consequence of pathogenic DLG4 variants, while revealing additional variant-associated effects on neuronal structure and activity, rescued by AAV9-mediated neuronal restoration.
Dania Abdellatif, Mustafa Obeid, Rami I. Aqeilan· bioRxiv· 0 citations
Heterozygous loss-of-function variants in Neurabin I (PPP1R9A), responsible for encoding a cytoskeletal scaffolding protein essential for synaptic plasticity, are recurrently associated with neurodevelopmental and neuropsychiatric disorders, yet their direct effects on human neuronal maturation remain unclear. Here, we establish the first comprehensive human mechanistic model of PPP1R9A haploinsufficiency using an isogenic CRISPR/Cas9-engineered iPSC system differentiated into cortical neurons to define dosage-dependent functional consequences. PPP1R9A+/- neurons exhibited pronounced hyperspinogenesis and increased neuritic complexity, indicative of aberrant structural maturation; however, whole-cell patch-clamp recordings revealed impaired intrinsic excitability, including reduced action potential firing, altered waveform properties, and defective axo-somatic coupling, uncovering a striking dissociation between neuronal morphology and function. Long-read single-cell transcriptomics and quantitative proteomics identified coordinated downregulation of ion channel and synaptic transmission pathways, including genes essential for sodium channel function and glutamatergic signaling, together with disruption of synaptic vesicle cycling, axon guidance, and neurodevelopmental programs. Pseudotime trajectory analysis further demonstrated delayed neuronal differentiation, with mutant neurons accumulating at intermediate developmental states rather than acquiring mature cortical identities. Importantly, molecular rescue experiments confirmed causality, as restoration of full-length PPP1R9A expression robustly normalized transcriptional and synaptic signaling programs, whereas allele-specific antisense oligonucleotide-mediated suppression of the mutant transcript achieved only partial rescue. Taken together, these findings establish PPP1R9A haploinsufficiency as a driver of impaired molecular, electrophysiological, and developmental maturation in human cortical neurons, providing a human-specific mechanistic framework linking reduced Neurabin I dosage to neurodevelopmental and psychiatric disease risk.
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
OBJECTIVE
SCN2A pathogenic mutations, such as the recurrent heterozygous Nav1.2-L1342P, are monogenic causes of epilepsy. In this human-induced pluripotent stem cell-derived model system, we aim to investigate the molecular and cellular mechanisms underlying SCN2A-L1342P-associated pathology.
METHODS
Using a human male induced pluripotent stem cell (iPSC) reference line (KOLF) carrying the Nav1.2-L1342P mutation, we generated three-dimensional (3D) cortical organoids for functional studies. Patch-clamp, multi-electrode array (MEA) recordings, immunocytochemistry, and RNA sequencing were used to characterize the disease phenotypes.
RESULTS
Nav1.2-L1342P organoid neurons displayed increased intrinsic excitability and amplified excitatory post-synaptic currents, which are consistent with an increase in excitatory synapse formation revealed by SYN1/PSD95 immunostaining. Moreover, elevated network firing activity, as demonstrated by MEA, indicates a pronounced network hyperexcitability. Transcriptomic profiling of organoids carrying the Nav1.2-L1342P mutation further revealed significant alterations in synaptic, glutamatergic, developmental, and senescence/apoptotic pathways.
SIGNIFICANCE
Our findings demonstrate that the Nav1.2-L1342P mutation drives a multifaceted disease phenotype, including network hyperexcitability and disruption of pathways related to neuronal and synaptic functions. These results advance our understanding of SCN2A-related developmental and epileptic encephalopathy (DEE), laying a foundation for personalized interventions.
M. I. Olivero-Acosta, Morgan Robinson, Zhefu Que et al.· Epilepsia· 0 citations
Amyotrophic lateral sclerosis (ALS) is a progressive neuromuscular disorder characterized by motoneurons degeneration. Functional studies have linked ALS to hyperexcitability and excitotoxicity, but the cause of the disease is unknown, though familial ALS cases are linked to pathogenic variants in several genes, including SOD1, TARDBP and FUS. Here we focused on the effect of the severe FUS (P525L) mutation on the functional properties of human spinal neurons derived from induced pluripotent stem cells (hiPSCs). This mutation delayed functional maturation, as revealed by the observation that mutated neurons showed alterations of membrane potential, reduced spontaneous synaptic activity, and altered action potentials at early differentiation stages. FUS (P525L) mutation was associated with a significant alteration of inhibitory signalling transmission: mutated neurons showed a significantly lower current response to GABA and glycine compared to control isogenic WT neurons of the same age. Also, glutamatergic currents exhibited a different temporal evolution in control and mutated neurons, but at a lower extent in comparison to inhibitory neurotransmitters. The decrease in the glycine-evoked currents was confirmed by the reduction of the expression of the α1 subunit of glycine receptor, measured by immunofluorescence assay. Similar functional alterations were measured in spinal neurons differentiated form a second hiPSC line, confirming the causative role of the FUS (P525L) mutation. Our data indicate that the FUS (P525L) mutation reduces the maturation rates and the function of hiPSC-derived spinal neurons, with a strong decrease of inhibitory transmission, which may affect the excitatory/inhibitory balance, possibly predisposing to excitotoxicity and neurodegeneration.
T. D’Andrea, Maria Cristina Benedetti, Michela Mochi et al.· Cellular and molecular neuro...· 0 citations