Child Health Institute of New Jersey and Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Rutgers University, New Brunswick, NJ, 08901, USA.
Center for Medical Genetics & Hunan Key Laboratory of Medical Genetics, School of Life Sciences, and Department of Psychiatry, The Second Xiangya Hospital, Central South University, 410008, Changsha, China.
Mol Psychiatry. 2024 Jun;29(6):1620-1635. doi: 10.1038/s41380-022-01834-x. Epub 2022 Oct 24.
Mutations in many synaptic genes are associated with autism spectrum disorders (ASD), suggesting that synaptic dysfunction is a key driver of ASD pathogenesis. Among these mutations, the R451C substitution in the NLGN3 gene that encodes the postsynaptic adhesion molecule Neuroligin-3 is noteworthy because it was the first specific mutation linked to ASDs. In mice, the corresponding Nlgn3 R451C-knockin mutation recapitulates social interaction deficits of ASD patients and produces synaptic abnormalities, but the impact of the NLGN3 R451C mutation on human neurons has not been investigated. Here, we generated human knockin neurons with the NLGN3 R451C and NLGN3 null mutations. Strikingly, analyses of NLGN3 R451C-mutant neurons revealed that the R451C mutation decreased NLGN3 protein levels but enhanced the strength of excitatory synapses without affecting inhibitory synapses; meanwhile NLGN3 knockout neurons showed reduction in excitatory synaptic strengths. Moreover, overexpression of NLGN3 R451C recapitulated the synaptic enhancement in human neurons. Notably, the augmentation of excitatory transmission was confirmed in vivo with human neurons transplanted into mouse forebrain. Using single-cell RNA-seq experiments with co-cultured excitatory and inhibitory NLGN3 R451C-mutant neurons, we identified differentially expressed genes in relatively mature human neurons corresponding to synaptic gene expression networks. Moreover, gene ontology and enrichment analyses revealed convergent gene networks associated with ASDs and other mental disorders. Our findings suggest that the NLGN3 R451C mutation induces a gain-of-function enhancement in excitatory synaptic transmission that may contribute to the pathophysiology of ASD.
许多突触基因的突变与自闭症谱系障碍（ASD）有关，这表明突触功能障碍是 ASD 发病机制的关键驱动因素。在这些突变中，编码突触后粘附分子 Neuroligin-3 的 NLGN3 基因中的 R451C 取代引人注目，因为它是第一个与 ASDs 相关的特定突变。在小鼠中，相应的 Nlgn3 R451C 敲入突变再现了 ASD 患者的社交互动缺陷，并产生了突触异常，但尚未研究 NLGN3 R451C 突变对人类神经元的影响。在这里，我们生成了具有 NLGN3 R451C 和 NLGN3 缺失突变的人类敲入神经元。引人注目的是，对 NLGN3 R451C 突变神经元的分析表明，R451C 突变降低了 NLGN3 蛋白水平，但增强了兴奋性突触的强度而不影响抑制性突触；同时，NLGN3 敲除神经元显示兴奋性突触强度降低。此外，NLGN3 R451C 的过表达在人类神经元中再现了突触增强。值得注意的是，用移植到小鼠前脑中的人类神经元进行体内研究证实了兴奋性传递的增强。使用共培养的兴奋性和抑制性 NLGN3 R451C 突变神经元的单细胞 RNA-seq 实验，我们鉴定了与突触基因表达网络相对应的相对成熟的人类神经元中的差异表达基因。此外，基因本体论和富集分析揭示了与 ASDs 和其他精神障碍相关的趋同基因网络。我们的研究结果表明，NLGN3 R451C 突变诱导兴奋性突触传递的功能获得性增强，这可能有助于 ASD 的病理生理学。
