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：
10231033
负责人：
JAMES J COLLINS
金额：
$ 166.57万
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/10231033
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 Culture Techniques 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 Regulator Genes 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 fetal gene discovery 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 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）首先是由专门的血液生成内皮产生的发育中的胚胎主动脉，迁移到胎儿肝脏并扩展，并最终定居骨骼骨髓，在成年生活中支持造血。这些独特的解剖位置藏有支持发展成熟，扩展，最终的专门微环境 HSC的自我更新和分化的平衡。促进形成和的转录程序造血茎和祖细胞（HSPC）的分化已被广泛询问，但很多关于造血微环境和在发育，HSC迁移，住宿，指定HSC出现的细胞 - 细胞相互作用机制和胎儿壁ni的扩张，以及骨骼的最终静止，自我更新和分化骨髓。在我们的初步数据中，我们收集了有关细胞类型数量的证据，包括内皮细胞，间充质细胞，巨噬细胞，神经rest衍生物和部分作为组成部分造血利基。我们将收集全面的“ OMIC”数据以对基因表达程序进行分类在主动脉 - 贡纳德 - 肾小球发育过程中发生的不同造血小众细胞中（AGM），胎儿肝，骨髓和plapeta（AIM 1）。我们的方法始于Tomo-Seq，这使我们能够发现具有特定区域特异性的细胞群体（例如内皮）所特有的基因表达模式专业化。我们将通过单细胞RNA-seq和原位验证FACS纯化细胞中细胞特异性表达杂交，并将在实验中使用Morpholino和CRISPR敲击记录功能可处理的斑马鱼模型。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，和图案找到软件，以识别DNA结合调节因素，这些因素是候选驱动因素造血细胞命运。我们将采用计算管道并开发新算法来分析这些数据（AIM 2）。 AIM 1和2出现的假设将通过构建新的记者菌株来测试斑马鱼和小鼠，以及携带合成记者和驱动因素的工程多能干细胞（AIM 3）。我们的目标是定义在HSC的关键时期指定利基细胞的分子电路出现和扩展，并探测利基元素和HSPC之间的串扰。我们希望能收集对驱动造血形成和成熟过程的分子机制的独特见解胚胎开发，并增强我们对HSC维护，静止，自我更新的理解和分化。