The overwhelming success of exome- and genome-wide association studies in discovering thousands of disease-associated genes necessitates developing novel high-throughput functional genomics approaches to elucidate the molecular mechanisms of these genes. Here, we have coupled multiplexed repression of neurodevelopmental disease–associated genes to single-cell transcriptional profiling in differentiating human neurons to rapidly assay the functions of multiple genes in a disease-relevant context, assess potentially convergent mechanisms, and prioritize genes for specific functional assays. For a set of 13 autism spectrum disorder (ASD)–associated genes, we show that this approach generated important mechanistic insights, revealing two functionally convergent modules of ASD genes: one that delays neuron differentiation and one that accelerates it. Five genes that delay neuron differentiation (ADNP, ARID1B, ASH1L, CHD2, and DYRK1A) mechanistically converge, as they all dysregulate genes involved in cell-cycle control and progenitor cell proliferation. Live-cell imaging after individual ASD-gene repression validated this functional module, confirming that these genes reduce neural progenitor cell proliferation and neurite growth. Finally, these functionally convergent ASD gene modules predicted shared clinical phenotypes among individuals with mutations in these genes. Altogether, these results show the utility of a novel and simple approach for the rapid functional elucidation of neurodevelopmental disease-associated genes.

中文翻译：

---

神经发育疾病相关基因的高通量单细胞功能阐明揭示了改变神经元分化的收敛机制。

外显子组和全基因组关联研究在发现数千种疾病相关基因方面取得了压倒性的成功，因此需要开发新的高通量功能基因组学方法来阐明这些基因的分子机制。在这里，我们将神经发育疾病相关基因的多重抑制与区分人类神经元的单细胞转录谱相结合，以快速分析多个基因在疾病相关环境中的功能，评估潜在的收敛机制，并优先考虑特定功能分析的基因. 对于一组 13 个自闭症谱系障碍 (ASD) 相关基因，我们表明这种方法产生了重要的机制见解，揭示了 ASD 基因的两个功能收敛模块：一个延迟神经元分化，一个加速神经元分化。ADNP、ARID1B、ASH1L、CHD2和DYRK1A）在机制上趋于一致，因为它们都失调了参与细胞周期控制和祖细胞增殖的基因。个体 ASD 基因抑制后的活细胞成像验证了该功能模块，证实这些基因减少神经祖细胞增殖和神经突生长。最后，这些功能上收敛的 ASD 基因模块预测了这些基因突变的个体之间的共享临床表型。总而言之，这些结果显示了一种新颖且简单的方法可用于快速阐明神经发育疾病相关基因的功能。