Mechanisms of Flow-driven Transcriptional Control of Hematopoietic Stem Cell Development by YAP

YAP 流驱动转录控制造血干细胞发育的机制

基本信息

批准号：
10596610
负责人：
Wade William Sugden
金额：
$ 15.22万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10596610
关键词：
Achievement Address Adult Advisory Committees Algorithms Animals Aorta Arteries Award Behavior Blood Blood Cells Blood Vessels Blood flow Boston Candidate Disease Gene Cell Nucleus Cells Cellular biology Chemicals Circulation Clinical Complex Coupled Coupling Cues DNA DNA Binding DNA-Protein Interaction Data Dedications Development Distant Dorsal Embryo Embryonic Development Endothelial Cells Endothelium Enhancers Environment Erythroid Event Exhibits Fetal Liver Fishes Future Gene Expression Generations Genes Genetic Genetic Transcription Goals Growth Growth and Development function Hematological Disease Hematology Hematopoiesis Hematopoietic Hematopoietic Cell Production Hematopoietic Neoplasms Hematopoietic Stem Cell Specification Hematopoietic Stem Cell Transplantation Hematopoietic System Hematopoietic stem cells Home Human Image Immune In Vitro Individual Integrins Laboratories Learning Link Logic Lymphoid Mammals Mechanical Stimulation Mechanics Mediating Mediator Membrane Membrane Proteins Mentors Mentorship Methods Molecular Morphogenesis Morphology Myelogenous Nuclear Organism Patients Pediatric Hospitals Periodicity Phase Phenotype Piezo 1 ion channel Piezo ion channels Postdoctoral Fellow Production Protein Activation Pathway Proteins Protocols documentation RUNX1 gene Regulation Regulator Genes Research Research Training Scientist Signal Transduction Signaling Protein Solid Sorting Specific qualifier value Stem Cell Development Stretching System Therapeutic Therapeutic Uses Tissues Training Transcriptional Regulation Translational Research Translations Transplantation Viscosity Work Zebrafish blood treatment career development cell type cofactor developmental genetics directed differentiation endothelial stem cell genetic manipulation genome-wide hematopoietic differentiation hematopoietic gene hematopoietic stem cell expansion hematopoietic stem cell fate hematopoietic stem cell formation hematopoietic tissue hemodynamics hemogenic endothelium improved in vivo in vivo evaluation induced pluripotent stem cell interest knock-down large scale production leukemia/lymphoma link protein mechanical force mechanical signal medical schools migration new technology non-genetic novel programs protein activation response self-renewal shear stress stem cell biology success transcription factor transcriptomic profiling transcriptomics vertebrate embryos

项目摘要

Project Summary/Abstract Hematopoietic stem cells (HSCs) are capable of producing all erythroid, myeloid and lymphoid blood cells of an organism. Coupled with their unique capacity for self-renewal, successful transplantation of healthy HSCs is the only therapy currently available that can completely replace and restore the blood system in patients with leukemia and lymphoma. Despite this need, HSCs presently cannot be efficiently created or cultured in vitro, suggesting that extrinsic factors supporting their growth and development in vivo are lacking from existing protocols. Previous work from our lab demonstrated that blood flow is an essential non-genetic environmental cue required for HSC production in vertebrate embryos, mediated in part by stimulating mechanical activation of the Yes-associated protein (YAP) transcription factor (TF). This proposal intends to resolve the physical, genetic and molecular mechanisms underlying mechanically-activated, YAP-driven HSC production. YAP, while a potent co-activator of gene expression, lacks DNA-binding ability of its own. To understand the molecular logic behind flow/YAP-driven hematopoiesis, the goal of the first aim is to employ chemical, physical and genetic perturbation of shear stress and cyclic stretch in live zebrafish embryos to assess the impact of these individual components of hemodynamic force on HSC production from hemogenic endothelium (HE). To this will be added tissue- specific transcriptomic and genome-wide YAP/DNA interaction profiling from sorted HE from wildtype zebrafish, flow-deficient and yap-/- animals (with normal blood flow) in order to discriminate flow-dependent gene regulatory modules and transcriptional targets that rely on YAP. Hypothesis-driven candidate TFs will be tested in vivo and in vitro to evaluate YAP-interaction ability and uncover key partners required for normal YAP-dependent hematopoiesis. In the second aim, the zebrafish system will be used to investigate candidate membrane- localized proteins, Piezos and Integrins, as components linking hemodynamic forces with YAP activation. These studies stand to provide a comprehensive “membrane-to-nucleus” paradigm for how blood flow activates YAP to guide developmental hematopoiesis, which may improve current efforts to generate or expand HSCs. As a postdoctoral fellow, Dr. Sugden will conduct his research in the laboratory of Dr. Trista North at Boston Children's Hospital. Her expertise in extrinsic regulation of developmental hematopoiesis, together with dedicated co-mentorship by Dr. George Daley (an expert in stem cell biology and hematology) and a strong advisory team provide an exceptionally well-supported environment for career development and research training. Dr. Sugden will build on a solid background in developmental genetics and live-imaging, by adding new technologies in transcription factor/DNA interaction profiling, transcriptomics and in vitro methods to study protein interactions. A rigorous research and training plan lay the groundwork for success, both in the mentored and independent phases of the award. The environment at Boston Children's Hospital and Harvard Medical School will provide the ideal surroundings to support Dr. Sugden to become a successful independent scientist.

项目概要/摘要造血干细胞 (HSC) 能够产生所有红系、髓系和淋巴系血液加上有机体独特的自我更新能力，成功移植健康的细胞。 HSCs是目前唯一可以完全替代和恢复患者血液系统的疗法尽管有这种需要，但目前还无法在体外有效地产生或培养 HSC，这表明现有的技术缺乏支持其体内生长和发育的外在因素。我们实验室之前的工作表明，血流是一种重要的非遗传环境。脊椎动物胚胎中 HSC 产生所需的线索，部分通过刺激机械激活来介导 Yes相关蛋白（YAP）转录因子（TF）这一提议旨在解决物理、遗传问题。以及机械激活、YAP 驱动的 HSC 生产的分子机制。基因表达的共激活剂，缺乏其自身的 DNA 结合能力了解背后的分子逻辑。流/YAP驱动的造血，第一个目标是利用化学、物理和遗传扰动活体斑马鱼胚胎中的剪切应力和循环拉伸，以评估这些单独成分的影响血流动力学对造血内皮 (HE) 生成 HSC 的影响来自野生型斑马鱼的分选 HE 的特定转录组和全基因组 YAP/DNA 相互作用分析，血流缺陷和 yap-/- 动物（血流正常），以区分血流依赖性基因调控依赖于 YAP 的假设驱动的候选 TF 的模块和转录目标将在体内和体外进行测试。体外评估 YAP 相互作用能力并发现正常 YAP 依赖所需的关键伙伴在第二个目标中，斑马鱼系统将用于研究候选膜。局部蛋白、压电蛋白和整合素，作为将血流动力学与 YAP 激活联系起来的成分。研究将为血流如何激活 YAP 提供全面的“膜到核”范例指导发育性造血，这可能会改善目前生成或扩大 HSC 的努力。作为博士后研究员，Sugden 博士将在 Trista North 博士的实验室进行研究她在发育性造血的外在调节方面的专业知识以及 George Daley 博士（干细胞生物学和血液学专家）的专门共同指导和强大的顾问团队为职业发展和研究提供非常良好的支持环境 Sugden 博士将通过添加新的内容，建立在发育遗传学和实时成像方面的坚实背景之上。转录因子/DNA 相互作用分析、转录组学和体外蛋白质研究方法等技术严格的研究和培训计划为指导和培训的成功奠定了基础。该奖项的独立阶段。波士顿儿童医院和哈佛医学院的环境。将为萨格登博士成为一名成功的独立科学家提供理想的环境。