Assessing the impact of mono- and bi-allelic deletions in NRXN1 on synaptic function

Massrali, A.; Paul, A.; Gatford, N. J.; Dutan-Polit, L.; Kedia, S.; Rahman, S.; Srivastava, D. P.; Kotter, M.; Adhya, D.; Baron-Cohen, S.

doi:10.64898/2026.02.20.707024

bioRxiv · 10.64898/2026.02.20.707024

Assessing the impact of mono- and bi-allelic deletions in NRXN1 on synaptic function

Massrali, A.·Paul, A.·Gatford, N. J.·Dutan-Polit, L.·Kedia, S.·Rahman, S.·Srivastava, D. P.·Kotter, M.·Adhya, D.·Baron-Cohen, S.

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Abstract

Neurexin 1 (NRXN1) is an adhesion protein involved in synapse development and function. Mutations in NRXN1 are strongly linked with neurodevelopmental and psychiatric conditions. Mono-allelic NRXN1 mutations are associated with autistic traits, with increased likelihood of co-occurring intellectual disability. However, mono-allelic mutations have variable penetrance and occur in individuals without neurodevelopmental phenotypes. Conversely, bi-allelic mutations, though rarer, are associated with more stable penetrance and severe neurodevelopmental phenotypes. Human induced pluripotent stem cells (iPSC) have been used to study how mutations in NRXN1 impacts its function, with most studies focusing on monoallelic mutations. In this study, we systematically compared monoallelic and biallelic mutations in NRXN1, characterising their effects on molecular, synaptic, and functional phenotypes. Using CRISPR-Cas9, we introduced indels in NRXN1 exon 19, in an iPSC line containing inducible NGN2. These edits caused either mono-allelic or compound bi-allelic frameshift mutations. iPSCs containing either mutation robustly generated glutamatergic neurons, but these neurons displayed reduced expression of major NRXN1 isoforms. Transcriptomic profiling revealed modest gene expression changes in mono-allelic mutant neurons, whereas bi-allelic mutants exhibited extensive dysregulation of gene networks associated with neuronal maturation and synaptic function. Furthermore, synaptic phenotypes were mild in mono-allelic mutants but pronounced in bi-allelic mutant neurons. Both mono-allelic and bi-allelic mutant neurons displayed alterations in neuronal network activity and reduced peak depolarisation responses to KCl stimulation. Together, these data demonstrate that NRXN1 exhibits gene-dosage sensitivity, with bi-allelic disruption of exon 19 unmasking molecular, synaptic, and functional phenotypes that are only modest in mono-allelic mutant neurons.

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Massrali, A., Paul, A., Gatford, N. J., Dutan-Polit, L., Kedia, S., Rahman, S., Srivastava, D. P., Kotter, M., Adhya, D., Baron-Cohen, S.. 2026-02-21. Assessing the impact of mono- and bi-allelic deletions in NRXN1 on synaptic function. https://doi.org/10.64898/2026.02.20.707024

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