奥地利科学院分子生物技术研究所(IMBA)Juergen A. Knoblich，Chong Li和瑞士生物系统科学与工程系Barbara Treutlein共同合作，近期取得重要工作进展。他们研究开发了单细胞脑类器官筛查方法，可以用来识别自闭症的发育缺陷。相关研究成果2023年9月13日在线发表于《自然》杂志上。

据介绍，人类大脑的发育涉及独特的过程（在许多其他物种中没有观察到），这些过程可能导致神经发育障碍。大脑类器官能够在人类环境中研究神经发育障碍。

研究人员开发了CRISPR-人类类器官-单细胞RNA测序（CHOOSE）系统，该系统使用经过验证的成对引导RNA，基于诱导型CRISPR–Cas9的遗传破坏和单细胞转录组学来筛选嵌合类器官的功能丧失。研究人员发现了36个与转录调控相关的高危自闭症谱系障碍基因的扰动，揭示了它们对细胞命运决定的影响。研究人员发现，背侧中间祖细胞、腹侧祖细胞和上层兴奋性神经元是最脆弱的细胞类型。

研究人员从单细胞转录组和染色质模式构建了大脑类器官的发育基因调控网络，并鉴定了自闭症谱系障碍相关和干扰富集的调控模块。BRG1/BRM相关因子（BAF）染色质重塑复合体成员的扰动导致端脑腹侧祖细胞的富集。具体而言，BAF亚基ARID1B的突变影响祖细胞向少突胶质细胞和中间神经元前体细胞的命运转变，研究人员在患者特异性诱导多功能干细胞来源的类器官中证实了这一表型。

总之，这一研究为类器官模型中疾病易感基因的高通量表型表征（具有细胞状态、分子途径和基因调控网络读数）提供了策略方法。

附：英文原文

Title: Single-cell brain organoid screening identifies developmental defects in autism

Author: Li, Chong, Fleck, Jonas Simon, Martins-Costa, Catarina, Burkard, Thomas R., Themann, Jan, Stuempflen, Marlene, Peer, Angela Maria, Vertesy, bel, Littleboy, Jamie B., Esk, Christopher, Elling, Ulrich, Kasprian, Gregor, Corsini, Nina S., Treutlein, Barbara, Knoblich, Juergen A.

Issue&Volume: 2023-09-13

Abstract: The development of the human brain involves unique processes (not observed in many other species) that can contribute to neurodevelopmental disorders1–4. Cerebral organoids enable the study of neurodevelopmental disorders in a human context. We have developed the CRISPR–human organoids–single-cell RNA sequencing (CHOOSE) system, which uses verified pairs of guide RNAs, inducible CRISPR–Cas9-based genetic disruption and single-cell transcriptomics for pooled loss-of-function screening in mosaic organoids. Here we show that perturbation of 36 high-risk autism spectrum disorder genes related to transcriptional regulation uncovers their effects on cell fate determination. We find that dorsal intermediate progenitors, ventral progenitors and upper-layer excitatory neurons are among the most vulnerable cell types. We construct a developmental gene regulatory network of cerebral organoids from single-cell transcriptomes and chromatin modalities and identify autism spectrum disorder-associated and perturbation-enriched regulatory modules. Perturbing members of the BRG1/BRM-associated factor (BAF) chromatin remodelling complex leads to enrichment of ventral telencephalon progenitors. Specifically, mutating the BAF subunit ARID1B affects the fate transition of progenitors to oligodendrocyte and interneuron precursor cells, a phenotype that we confirmed in patient-specific induced pluripotent stem cell-derived organoids. Our study paves the way for high-throughput phenotypic characterization of disease susceptibility genes in organoid models with cell state, molecular pathway and gene regulatory network readouts. We develop a high-throughput CRISPR screening system in cerebral organoids and identify vulnerable cell types and gene regulatory networks associated with autism spectrum disorder from single-cell transcriptomes and chromatin modalities.

DOI: 10.1038/s41586-023-06473-y

Source: https://www.nature.com/articles/s41586-023-06473-y