UCLA High-Throughput Neuropsychiatric Disorder Phenotyping Center (UCLA HT-NPC)

加州大学洛杉矶分校高通量神经精神疾病表型中心 (UCLA HT-NPC)

• 批准号: 10643541
• 负责人: DANIEL H GESCHWIND
• 金额: $ 165.22万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10643541
• 关键词: 3-Dimensional; Address; Alleles; Anatomy; Architecture; Astrocytes; Biological; Biological Assay; Biological Testing; Brain; CRISPR/Cas technology; Calcium; Cell Line; Cell Survival; Cells; Cerebrum; Clustered Regularly Interspaced Short Palindromic Repeats; Coculture Techniques; Communities; Core Facility; Coupled; Custom; Data; Defect; Development; Developmental Delay Disorders; Dideoxy Chain Termination DNA Sequencing; Disease; Disease Pathway; Essential Genes; Female; Gene Expression; Genes; Genetic Heterogeneity; Genetic study; Genome engineering; Human; Human Genetics; Image; In Vitro; Intellectual functioning disability; Knock-out; Link; Measures; Methods; Modeling; Molecular; Morphology; Mutation; Nervous System; Neurobiology; Neurodevelopmental Disorder; Neurons; Neurophysiology - biologic function; Nuclear RNA; Optics; Organoids; Pathway interactions; Phenotype; Physiological; Process; Production; Proliferating; Proteins; Reproducibility; Research Personnel; Resolution; Resource Sharing; Role; Schizophrenia; Small Nuclear RNA; Synapses; System; Testing; Time; autism spectrum disorder; cell type; data sharing; density; disease phenotype; disorder risk; engineered stem cells; excitatory neuron; gene function; genetic manipulation; genetic risk factor; high dimensionality; human disease; human embryonic stem cell; human model; human pluripotent stem cell; human stem cells; in vivo; induced pluripotent stem cell; inhibitory neuron; knockout gene; male; model organism; molecular phenotype; multimodality; nerve stem cell; neural; neurite growth; neurodevelopment; neurogenesis; neuronal cell body; neuropsychiatric disorder; novel; null mutation; phenotypic data; risk variant; screening; sensor; sex; single nucleus RNA-sequencing; single-cell RNA sequencing; small hairpin RNA; stem cell model; stem cells; three dimensional cell culture; three-dimensional modeling; transcriptome sequencing; voltage

Project Summary/Abstract Human genetic studies have identified hundreds of genes contributing to Neuropsychiatric and Neurodevelop- mental Disease (NPD) risk. But for most genes, their normal function or the consequences of their absence or reduction on neurodevelopment and neural function are not known. Here, we propose to address the substantial challenges of discerning potential functions of hundreds of NPD genes through the development of a High Throughput Neuropsychiatric Disease Phenotyping Center (UCLA HT-NPC), driven by the activity of 9 highly collaborative investigators (Aharoni, Bhaduri, Damoiseaux, Geschwind, Golshani, Kitai, Luo, Novich, and Wells) and two substantial core facilities (UCLA Molecular Screening Shared Resource and the Human Stem Cell and Genome Engineering Center). Through a tiered approach, we combine high throughput and high value, quantitative phenotyping with stem cell engineering to characterize the functional consequences of NPD gene knockouts (null alleles), a key initial step that will inform our understanding of disease pathways. In the first step, we will rapidly generate null alleles for 250 genes chosen by the Consortium using a rapid, high throughput lentiviral based system in hESCs. Viability and neural induction potential will be assessed, and quantitative phenotyping conducted using RNA-seq on all lines. Those genes passing viability and neural induction tests will be used in the production of clonal null hiPSC lines (male and female) for downstream phenotyping and wider distribution to the community. Subsequently, we will perform high throughput, quantitative, multi-scale phenotyping at the molecular, morphological, and physiological levels in both 2D and 3D hiPSC-based models of human cortical development. We leverage the relative strengths and scalability of each model to enable us to perform both snRNA and bulk RNA-seq, measure the maturation, morphology, and synaptic density of neural cells using automated imaging, including the multiplexed, protein-based CODEX (Phenocycler) platform, and characterize neuronal activity and synchronization through optical recordings using custom-built mini-scope arrays (STIMscope). By using multiple systems (e.g. hESC/hiPSC; gene editing, 2D and 3D cultures), we test biological reproducibility across systems and technical reproducibility through replication. The use of experimentally validated, quantitative phenotypes across multiple scales of analysis facilitates data sharing and comparisons with other SSPsyGene investigators and provides a template for the field more broadly.

项目总结/摘要 人类遗传学研究已经确定了数百个基因有助于神经精神和神经发育- 精神疾病（NPD）风险。但对于大多数基因来说，它们的正常功能或缺失的后果， 对神经发育和神经功能的影响尚不清楚。在这里，我们建议解决实质性的问题， 通过开发一个高水平的NPD基因来识别数百个NPD基因的潜在功能的挑战 美国加州大学洛杉矶分校神经精神疾病表型中心（UCLA HT-NPC），由9个高度活跃的 合作研究者（Aharoni、Bhaduri、Damoiseaux、Geschwind、Golshani、Kitai、Luo、Novich和威尔斯） 和两个实质性的核心设施（加州大学洛杉矶分校分子筛选共享资源和人类干细胞， 基因组工程中心）。通过分层方法，我们将联合收割机的高吞吐量和高价值， 定量表型与干细胞工程，以表征NPD基因的功能后果 基因敲除（无效等位基因），这是一个关键的初始步骤，将告知我们对疾病途径的理解。第一步， 我们将使用快速、高通量的方法， 在hESC中的基于慢病毒的系统。将评估活力和神经诱导潜力，并定量 使用RNA-seq对所有品系进行表型分型。那些通过活力和神经诱导测试的基因将 用于生产克隆无效hiPSC系（雄性和雌性），用于下游表型分析和更广泛的 分发给社区。随后，我们将进行高通量，定量，多尺度 在基于2D和3D hiPSC的模型中，在分子、形态和生理水平上进行表型分析 人类大脑皮层发育的一部分我们利用每个模型的相对优势和可扩展性， 进行snRNA和批量RNA-seq，测量神经细胞的成熟、形态和突触密度， 使用自动成像的细胞，包括多路复用的基于蛋白质的CODEX（Phenocycler）平台，以及 使用定制的微型显微镜通过光学记录表征神经元活动和同步 阵列（STIMscope）。通过使用多个系统（例如hESC/hiPSC;基因编辑，2D和3D培养），我们测试了 系统间的生物再现性和通过复制的技术再现性。使用 实验验证，跨多个分析尺度的定量表型促进了数据共享， 与其他SSPsyGene研究人员进行比较，并为更广泛的领域提供了模板。