Single-cell transcriptomic analysis of sibling progenitors with differential Notch activity

具有差异Notch活性的兄弟祖细胞的单细胞转录组分析

• 批准号: 10372848
• 负责人: Su Guo
• 金额: $ 44.41万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10372848
• 关键词: Address; Bar Codes; Biological; Brain; Brain Diseases; Brain Neoplasms; Cell Culture System; Cell division; Cells; Cerebral Ventricles; Clone Cells; Communities; Computer Analysis; DNA; DNA delivery; Data; Data Set; Daughter; Development; Diagnosis; Electroporation; Embryo; Exploratory/Developmental Grant; Foundations; Future; Gene Expression; Genes; Genetic; Human; Image; In Situ; In Vitro; Knowledge; Label; Methods; Molecular; Mus; Nature; Neurodevelopmental Disorder; Neuroglia; Outcome; Pathway interactions; Population; Process; Prosencephalon; Public Health; RNA; Radial; Recovery; Reporter; Research; Resolution; Sampling; Siblings; Technology; Time; Transcript; Viral; Work; Zebrafish; base; daughter cell; differential expression; embryonic stem cell; experimental study; genetic signature; high reward; high risk; in vivo; nerve stem cell; neurogenesis; notch protein; novel; novel therapeutics; progenitor; programs; self-renewal; single-cell RNA sequencing; stem cell self renewal; transcriptomics

PROJECT SUMMARY It has been observed across species that embryonic radial glia neural progenitors undergo asymmetric cell division (ACD) to generate daughter cells with different Notch activity. The Notchhi daughter undergoes self- renewal, whereas the Notchlo daughter embarks on differentiation. Being able to molecularly define such different states will significantly advance our understanding of how self-renewal and differentiation are regulated. Although the relative Notch activity levels between embryonic daughters are correlated with their self- renewing vs. differentiation potential, the absolute Notch activity is heterogeneous across the progenitor population. This makes it impossible to simply sort single-cell RNA-seq (scRNA-seq) data based on the expression levels of Notch effectors (e.g. hes/her transcript levels). It is therefore important to track the lineage relationships among progenitors precisely at the level of sibling cells in scRNA-seq experiments. This exploratory R21 application, motivated by an important biological problem, aims to establish high resolution sib lineage-tracing and combine it with scRNA-seq. This represents a technological breakthrough that will enable comparison of gene expression profiles between sibling cells. Expected outcomes and impact: If successful, this project will establish a new and broadly applicable method in which sib cell states can be compared at the transcriptomic level both in vivo and in vitro. By applying this method to embryonic radial glia progenitors that are undergoing asymmetric cell division during active neurogenesis, we expect to uncover evolutionarily conserved core genes and pathways distinguishing Notchhi and Notchlo sib states that are shared across sib-lineages. Since Notchhi and Notchlo sib states are associated with self-renewal and differentiation respectively in embryonic progenitors, we expect to gain a glimpse into whether and how self-renewal and differentiation as distinct cellular states can be depicted at the transcriptomic level that are uncoupled from specific lineage outcomes. We will make this novel dataset with precise clonal tracking widely available to the broad research community. This project will lay foundation for a future R01, which aims to dissect the function of signature genes and pathways that define Notchhi vs. Notchlo cell states. In the long run, new basic knowledge about the underlying genetic programs will aid in developing new therapeutic ideas. The high risk and high reward nature of the proposed work makes this application well suited for the R21 mechanism.

项目概要 跨物种观察到胚胎放射状胶质神经祖细胞经历不对称细胞 分裂（ACD）以产生具有不同Notch活性的子细胞。 Notchhi 女儿经历了自我 更新，而 Notchlo 女儿则开始分化。能够从分子上定义这种不同的 各国将极大地增进我们对自我更新和分化如何监管的理解。 尽管胚胎女儿之间的相对Notch活动水平与其自我相关 更新与分化潜力，绝对Notch活性在整个祖细胞中是异质的 人口。这使得不可能简单地基于单细胞 RNA-seq (scRNA-seq) 数据进行排序 Notch 效应子的表达水平（例如他/她的转录水平）。因此追踪血统很重要 在 scRNA-seq 实验中，祖细胞之间的关系精确地在兄弟细胞水平上进行。 这一探索性 R21 应用受到一个重要的生物学问题的推动，旨在建立高 解析同胞谱系追踪并将其与 scRNA-seq 相结合。这代表着一项技术突破 将能够比较兄弟细胞之间的基因表达谱。 预期成果和影响：如果成功，该项目将建立一个新的、广泛的 可以在体内和转录组水平上比较同胞细胞状态的适用方法 体外。通过将此方法应用于正在经历不对称细胞的胚胎径向神经胶质祖细胞 在活跃的神经发生过程中进行分裂，我们期望发现进化上保守的核心基因和通路 区分 Notchhi 和 Notchlo 同胞状态，这些状态在同胞谱系之间共享。由于 Notchhi 和 Notchlo 同胞 状态分别与胚胎祖细胞的自我更新和分化相关，我们期望 了解是否以及如何将自我更新和分化描述为不同的细胞状态 在转录组水平上，与特定的谱系结果无关。我们将制作这本小说 具有精确克隆跟踪的数据集，广泛可供广大研究界使用。 该项目将为未来的 R01 奠定基础，旨在剖析特征基因的功能 以及定义 Notchhi 与 Notchlo 细胞状态的途径。从长远来看，新的基础知识 潜在的基因计划将有助于开发新的治疗理念。高风险和高回报的本质 所提出的工作使该应用程序非常适合 R21 机制。