| Objective: Most research on neural developmental disorders,including autism spectrum disorders(ASD),intellectual disabilities,bipolar disorder,schizophrenia,etc,were studied in animal models such as mice.However,the mice brain lacks the same cell types and structures as the human brain.In addition,due to the limited access of human embryos,it is hard to obtain enough specimens for studies in vitro,and it is impossible to complete the dynamic observation and research on the early stage of human embryos development.The establishment of human brain organoid model has solved this problem and become another research model for early neurodevelopmental disorders.ASD is a genetically heterogeneous disorder linked with rare,inherited and de novo mutations occurring in two main functional gene categories: gene expression regulation and synaptic function.SYNGAP1 is one of the top risk genes of ASD.Accumulating evidence points to dysregulation in cortical neurogenesis as a convergent mechanism in ASD pathophysiology.This study aimed to investigate the pathological roles of SYNGAP1 haploinsufficiency in the neurodevelopment with the human brain organoid model.Methods: To explore the effect of SYNGAP1 on early neural development in human,we searched for evidence of SYNGAP1 expression at the transcriptomic level,using the existing single-cell sequencing datasets as well as Brainspan database of human early embryos to study the main cell types that SYNGAP1 expressed in.Then,we confirmed the distribution of SYNGAP1 protein in the brain tissue of early human embryo as well as the induced pluripotent stem cells(i PSCs)derived brain organoids.Finally,we investigated the regulatory role of SYNGAP1 mutation in early neurodevelopment by comparing the organoids from patients with SYNGAP1 haploinsufficiency and the organoids from the control group after gene editing.To explore the effects of SYNGAP1 on the neural development,we studied the formation and structure of the cytoskeleton of human radial glial cells(RGCs),the development of cortical plate and cells stratification,the developmental trajectory of radial glial cells,and the development and maturation of cortical projection neurons.Results: Here we show for the first time the expression of the synaptic Ras GTP-ase activating protein 1(SYNGAP1),one of the top ASD risk genes,in human cortical progenitors(h CPs).Interestingly,we found that multiple components of the postsynaptic density(PSD)of excitatory synapses,of which SYNGAP1 is one of the most abundant components,are enriched in the proteome of h CPs.Specifically,we discover that SYNGAP1 is expressed within the apical domain of human radial glia cells(RGCs)where it lines the wall of the developing cortical ventricular zone colocalizing with the tight junction-associated protein and MAGUK family member TJP1.In a cortical organoid model of SYNGAP1 haploinsufficiency,we show dysregulated cytoskeletal dynamics that impair the scaffolding and division plane of RGCs,resulting in disrupted lamination of the cortical plate and accelerated maturation of cortical projection neurons.These effects may rely on its RASGAP domain.With single-cell transcriptomic analysis of the cerebral cortical organoids of the SYNGAP1 mutant line and the control line,combined with the analysis of the morphology and function of neurons in organoids,we found that SYNGAP1 could regulate the timeline of neural differentiation and maturation.With SYNGAP1 mutantation,the cerebral organoids showed accelerated maturation of cortical projection neurons.Conclusion: Overall,the dual function of SYNGAP1 in neuronal synapses and progenitor cells reframes our understanding of the pathophysiology of SYNGAP1-related disorders.SYNGAP1 haploinsufficiency could dysregulate cytoskeletal dynamics of h RGCs and impair the adherens of ventricular zone(VZ),resulting in disrupted lamination of the cortical plate and the migration of early born neurons.SYNGAP1 haploinsufficiency could lead to more asymmetric division of RGCs and accelerate the maturation of cortical projection neurons.More broadly,this study underscores the importance of dissecting the role of synaptic genes associated with neurodevelopmental disorders in distinct cell types across developmental stages. |
