Molecular Circuits in the Hematopoietic Stem Cell Niche

造血干细胞生态位中的分子回路

基本信息

批准号：
10410454
负责人：
JAMES J COLLINS
金额：
$ 163.53万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-15 至 2025-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10410454
关键词：
ATAC-seq Adipocytes Adult Algorithms Anatomy Aorta Atlases Blood Bone Marrow Bone Marrow Transplantation Cataloging Catalogs Cell Adhesion Molecules Cell Communication Cell Count Cell Differentiation process Cell Line Cell Maintenance Cell surface Cells Chromatin Clustered Regularly Interspaced Short Palindromic Repeats Coculture Techniques Code Collaborations Communities Computational algorithm Computer software DNA Binding DNA cassette Data Data Set Derivation procedure Development Ecosystem Elements Embryo Embryonic Development Endothelial Cells Endothelium Engineering Equilibrium Fetal Liver Fishes Gene Expression Gene Expression Profile Gene Transfer Generations Genes Genetic Transcription Glean Goals Gonadal structure Hematology Hematopoiesis Hematopoietic Hematopoietic stem cells Homing Human In Situ Hybridization In Vitro Investigational Therapies Life Location Mammals Mesenchymal Mesonephric structure Mining Modeling Molecular Mouse Strains Mus Neural Crest Placenta Pluripotent Stem Cells Population Pregnancy Recreation Reporter Reporting Research Research Personnel Resolution Resources Shapes Signal Transduction Site Somites Source Specific qualifier value System Techniques Testing Therapeutic Transplantation Umbilical Cord Blood Work Zebrafish cell type computational pipelines critical period data resource directed differentiation fetal gene discovery gene regulatory network genetic manipulation hematopoietic differentiation hematopoietic stem cell differentiation hematopoietic stem cell emergence hematopoietic stem cell expansion hematopoietic stem cell niche hemogenic endothelium in vivo innovation insight intercellular communication knock-down macrophage novel open source programs recruit self-renewal single-cell RNA sequencing stem cell migration stem cells synthetic biology synthetic construct transcriptome sequencing translational applications user-friendly web site

项目摘要

In mammals, hematopoietic stem cells (HSCs) first arise from a specialized hemogenic endothelium that lines the developing embryonic aorta, migrate to and expand in the fetal liver, and ultimately colonize the bone marrow, which supports hematopoiesis throughout adult life. These distinct anatomic locations harbor specialized microenvironments that support the developmental maturation, expansion, and ultimately the balance of self-renewal and differentiation of HSCs. The transcriptional programs that promote formation and differentiation of hematopoietic stem and progenitor cells (HSPCs) have been widely interrogated, but much remains to be learned about the supportive niche cells of the hematopoietic microenvironment and the mechanisms of cell-cell interaction that specify HSC emergence during development, HSC migration, lodging, and expansion in fetal niches, and the ultimate quiescence, self-renewal, and differentiation in the bone marrow. In our preliminary data, we have gathered evidence for number of cell types, including endothelial cells, mesenchymal cells, macrophages, neural crest derivatives, and somites as components of the hematopoietic niche. We will gather comprehensive “omics” data to catalogue the gene expression programs within the distinct hematopoietic niche cells that occur during development in the aorta-gonad-mesonephros (AGM), fetal liver, bone marrow, and placenta (aim 1). Our approach begins with tomo-seq, which enables us to discover gene expression patterns unique to cell populations like endothelium that have region-specific specialization. We will validate cell-specific expression in FACS purified cells by single cell RNA-seq and in situ hybridization, and will document functionality using morpholino and CRISPR knock-down in the experimentally tractable zebrafish model. We then use ATAC-seq to define functional open chromatin around these genes, and motif-finding software to identify DNA-binding regulatory factors that are candidate drivers of hematopoietic cell fate. We will employ a computational pipeline and develop novel algorithms to analyze these data (aim 2). Hypotheses emerging from aims 1 and 2 will be tested by constructing novel reporter strains of zebrafish and mice, as well as engineered pluripotent stem cells carrying synthetic reporters and drivers (aim 3). Our goal is to define the molecular circuitry that specifies niche cells during the critical periods of HSC emergence and expansion, and to probe cross-talk between niche elements and HSPCs. We hope to glean unique insights into the molecular mechanisms that drive hematopoietic formation and maturation during embryonic development, and to enhance our understanding of HSC maintenance, quiescence, self-renewal and differentiation.

在哺乳动物中，造血干细胞 (HSC) 首先产生于特殊的造血内皮细胞，该内皮细胞排列在发育中的胚胎主动脉，迁移到胎儿肝脏并在其中扩张，最终定植于骨骼骨髓，在整个成年生命中支持造血功能。支持发育成熟、扩展以及最终发育的专门微环境 HSC 的自我更新和分化的平衡，促进形成和分化的转录程序。造血干细胞和祖细胞（HSPC）的分化已被广泛质疑，但很多关于造血微环境的支持性生态位细胞和细胞间相互作用的机制，指定发育过程中 HSC 的出现、HSC 迁移、倒伏、胎儿生态位的扩张和最终的静止、自我更新和骨骼的分化在我们的初步数据中，我们收集了多种细胞类型的证据，包括内皮细胞。细胞、间充质细胞、巨噬细胞、神经嵴衍生物和体节作为组成部分我们将收集全面的“组学”数据来对基因表达程序进行分类。在主动脉-性腺-中肾发育过程中出现的独特造血生态位细胞内 (AGM)、胎儿肝脏、骨髓和胎盘（目标 1）。我们的方法始于 tomo-seq，它使我们能够实现。发现具有区域特异性的细胞群（如内皮细胞）特有的基因表达模式我们将通过单细胞 RNA-seq 和原位验证 FACS 纯化细胞中的细胞特异性表达。杂交，并将在实验中使用吗啉代和 CRISPR 敲除来记录功能然后我们使用 ATAC-seq 来定义这些基因周围的功能性开放染色质，和基序寻找软件来识别 DNA 结合调节因子，这些因子是候选驱动因素我们将采用计算管道并开发新的算法来分析这些。数据（目标 2）将通过构建新的报告菌株来检验目标 1 和 2 中出现的假设。斑马鱼和小鼠，以及携带合成分子和驱动程序的工程多能干细胞（目标 3).我们的目标是定义HSC关键时期指定利基细胞的分子电路。的出现和扩展，并探讨利基元素和 HSPC 之间的串扰。对驱动造血形成和成熟的分子机制有独特的见解胚胎发育，并增强我们对 HSC 维持、静止、自我更新的理解和差异化。