High-throughput in vivo and in vitro functional and multi-omics screens of neuropsychiatric and neurodevelopmental disorder risk genes

项目总结 迫切需要开发高通量、可伸缩的分析方法来识别潜在的生物学机制 神经发育和神经精神障碍(NPD)的危险基因。在这项提案中，我们的目标是利用 人类诱导多能干细胞和斑马鱼两种可扩展系统的独特优势 -在体外和体内进行NPD基因的平行功能分析，并试点开发 可跨系统应用的创新空间多组学技术。我们建议建立一项化验和 数据生成中心(ADGC)是SSPsygene联盟的一部分，该联盟利用独特的 我们实验室在大规模HiPSC CRISPR屏幕(Brennand)、高通量 斑马鱼屏幕(Hoffman)和尖端多组学工具开发(Fan)。我们的目标是获得新奇的东西 对不同NPD基因功能丧失的趋同和分化机制的洞察 在分子、细胞、结构、回路和行为水平上影响神经发育。我们建议 在HiPSCs和斑马鱼中采用分级策略通过汇集和排列筛选250个NPD基因 HIPSCs来源的神经元和神经胶质细胞的转录和表型筛选(目标1)，CRISPR筛选 斑马鱼评估基因功能丧失对全脑结构、活动和基本行为的影响 (目标2)，以及空间转录和多染色体CRISPR筛选，以研究 两个系统中NPD基因的破坏(目标3)。我们将通过识别生物相关的生物来推进这一领域 NPD基因在多个尺度上功能丧失所导致的表型，告知基因优先顺序 模式，并建立新的空间多组学平台用于NPD基因的功能分析。这些 研究将产生前所未有的匹配的分子、细胞、结构、电路和 HiPSC和斑马鱼的行为数据，将提供给更广泛的社区公开分发 为NPD提供新的见解。

PROJECT SUMMARY There is a critical need to develop high-throughput scalable assays to identify biological mechanisms underlying risk genes in neurodevelopmental and neuropsychiatric disorders (NPD). In this proposal, we aim to leverage the unique advantages of two scalable systems – human induced pluripotent stem cells (hiPSCs) and zebrafish – to perform parallel functional assays of NPD genes in vitro and in vivo, and to pilot the development of innovative spatial multi-omics technologies applicable across systems. We propose to establish an Assay and Data Generation Center (ADGC) as part of the SSPsyGene Consortium that capitalizes on the unique and complementary expertise of our labs in large-scale hiPSC CRISPR screens (Brennand), high-throughput zebrafish screens (Hoffman), and cutting-edge multi-omics tool development (Fan). Our goal is to gain novel insights into the convergent and divergent mechanisms by which diverse NPD gene loss of function affects neurodevelopment at the molecular, cellular, structural, circuit, and behavioral levels. We propose to screen 250 NPD genes using a tiered strategy in hiPSCs and zebrafish by conducting pooled and arrayed transcriptomic and phenotypic screens in hiPSCs-derived neurons and glia (Aim 1), CRISPR screens in zebrafish to assess the effects of gene loss of function on whole-brain structure, activity, and basic behaviors (Aim 2), and spatial transcriptomic and multi-omic CRISPR screens to investigate the transcriptional effects of NPD gene disruption in both systems (Aim 3). We will advance the field by identifying biologically relevant phenotypes resulting from NPD gene loss of function across multiple scales, informing gene prioritization schema, and establishing new spatial multi-omics platforms for the functional analysis of NPD genes. These studies will generate an unprecedented resource of matched molecular, cellular, structural, circuit, and behavioral data in hiPSCs and zebrafish, which will be provided for open distribution to the broader community to yield new insights into NPD.