Comprehensive single-cell atlas of the developing mouse brain

项目摘要 发育中的小鼠脑是研究脑内细胞类型起源的基础实验模型。 哺乳动物的大脑对小鼠大脑发育的全面了解对于比较研究至关重要 神经发育过程，这是理解显着的进化创新的关键， 将人类与其他物种区分开来。此外，发育信息使得能够在细胞分类中细化细胞分类。 通过将细胞类型和谱系起源的知识结合到成人细胞分类中来研究成人大脑。尽管 最近创建的成年小鼠大脑分子图谱所带来的变革性见解，我们目前 缺乏对发育中小鼠大脑细胞类型的全面普查，以及将其联系起来的谱系关系。 他们是成年人中的佼佼者。 在这里，我们试图生成一个全面的，空间和时间分辨率，细胞分辨率的地图集， 整个发育中的小鼠大脑，在整个胚胎期和出生后以高分辨率采样 大脑发育（从E8.0到P28）。我们将采用三种互补的方法来生成 全面的多组单细胞图谱：10 x Genomics单细胞RNA-seq（scRNA-seq），10 x Genomics Multiome（同步单核RNA-seq和ATAC-seq，用于联合转录组和表观基因组 分析）和Smart-seq 3（用于全长深度RNA测序）。同时，我们将使用空间分辨的 转录组学方法MERFISH在相同的密集采样时间轴上，以确定空间分布 所有细胞类型和整个小鼠大脑细胞状态的动态变化。我们将应用计算 从这些空间和时间分辨的数据集预测发育谱系关系的方法， 并通过基于条形码的体内谱系追踪和通过 使用多重子宫内CRISPR筛选（Perturb-seq）功能性测试候选分子效应物。 最后，我们将试验跨物种发育数据集的整合，绘制单细胞组学数据集 从发育中的人类和非人类灵长类动物的大脑到全面的小鼠大脑发育细胞， 类型图集建立在这里，以创建一个计算对齐的发展时间， 了解物种间特定发育事件的差异调节。总的来说，这个项目 汇集了一组在大脑发育，电路， 单细胞基因组学，以及大脑图谱的组装，以产生一个全面的发育脑细胞图谱， 旨在作为神经科学界研究的第一个基础资源， 哺乳动物大脑发育和神经发育障碍的机制。我们的计划将 大大有助于BICAN的总体目标，以产生不同细胞的基础知识， 类型和它们在大脑中的三维组织原则跨越寿命和进化。

PROJECT SUMMARY The developing mouse brain is a foundational experimental model for investigation of the origins of cell types in the mammalian brain. Comprehensive knowledge of mouse brain development is critical for comparative studies of neurodevelopmental processes, which are key to understanding the remarkable evolutionary innovations that distinguish humans from other species. In addition, developmental information enables refining cell taxonomy in the adult brain by incorporating knowledge of cell type and lineage origins into adult cell classification. Despite the transformative insights enabled by the recently created molecular atlas of the adult mouse brain, we currently lack a comprehensive census of cell types of the developing mouse brain, and the lineage relationships that link them to their adult counterparts. Here we seek to generate a comprehensive, spatially- and temporally-resolved, cellular-resolution atlas of the whole developing mouse brain, sampled at high resolution through the entire period of embryonic and postnatal brain development (from E8.0 to P28). We will employ three complementary approaches to generate comprehensive multi-omic single-cell profiles: 10x Genomics single-cell RNA-seq (scRNA-seq), 10x Genomics Multiome (simultaneous single-nucleus RNA-seq and ATAC-seq, for combined transcriptomic and epigenomic profiling), and Smart-seq3 (for full-length deep RNA-sequencing). In parallel, we will use the spatially resolved transcriptomic method MERFISH across the same densely-sampled timeline, to identify the spatial distribution of all cell types and dynamic changes in cell states across the entire mouse brain. We will apply computational methods to predict developmental lineage relationships from these spatially and temporally resolved datasets, and experimentally validate lineage relationships through both barcode-based in vivo lineage tracing and by functionally testing candidate molecular effectors using multiplexed in utero CRISPR screening (Perturb-seq). Finally, we will pilot integration of developmental datasets across species, mapping single-cell omics datasets from the developing human and non-human primate brains onto the comprehensive mouse brain developing cell type atlas established here, to create a computational alignment of developmental time that will enable understanding of differential regulation of specific developmental events across species. Overall, this project brings together a team of investigators with extensive, demonstrated expertise in brain development, circuitry, single-cell genomics, and assembly of brain atlases to produce a comprehensive developmental brain cell atlas, intended to serve as a first-of-its-kind foundational resource to the neuroscience community for the study of mechanisms of mammalian brain development and neurodevelopmental disorders. Our proposed project will contribute substantially to the overarching goal of BICAN to generate fundamental knowledge on diverse cell types and their three-dimensional organizational principles in the brain across lifespan and evolution.