权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Defining Nfix's role as a novel key regulator of hematopoietic stem cells

定义 Nfix 作为造血干细胞新型关键调节因子的作用

基本信息

批准号：
9198983
负责人：
SHANNON L MCKINNEY-FREEMAN
金额：
$ 40.39万
依托单位：
ST. JUDE CHILDREN'S RESEARCH HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-01-01 至 2020-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9198983
关键词：
Adult Animal Testing Apoptosis Apoptotic B-Lymphocytes Binding Bioinformatics Blood Bone Marrow Bone Marrow Transplantation Cancer Patient Candidate Disease Gene Cell Death Cell Line Cell Survival Cell physiology Cells Cessation of life ChIP-seq Chimera organism Clinical Data Defect Embryo Engraftment Erythroid Exhibits Family member Frequencies Gene Expression Gene Family Genes Goals Growth Hematological Disease Hematopoiesis Hematopoietic Hematopoietic Neoplasms Hematopoietic Stem Cell Transplantation Hematopoietic stem cells Hippocampus (Brain)Impairment Laboratories Life Marrow Mediating Molecular Molecular Target Mus Mutation Myeloid Cells NFIX gene Natural regeneration Pathway interactions Patients Pattern Play Population Postdoctoral Fellow Proteins Regulation Regulator Genes Research Personnel Role Stem cells System TNFRSF5 gene Tamoxifen Testing Time Transplantation United States Work gene discovery granulocyte improved in vivo knock-down leukemia novel nuclear factor 1 overexpression reconstitution relating to nervous system screening small hairpin RNA stem stem cell population transcriptome

项目摘要

Project Summary Hematopoietic stem cell (HSC) transplantation is currently used >16,000 times/year in the United States to treat hematologic disease, leukemia, and to restore hematopoiesis in cancer patients. HSC are thus one of the most clinically exploited stem cell populations. Multiple recent studies demonstrate that the hematopoietic stem and progenitor cells (HSPC) that restore hematopoiesis following bone marrow transplant (BMT) are distinct from those that sustain native hematopoiesis. These findings place an imperative on better understanding the selective regulation of HSPC repopulation. Thus, our laboratory seeks to dissect the cellular and molecular mechanisms that regulate HSC repopulation. In a functional screen for genes whose depletion perturbs HSC in vivo repopulation, we identified the Nuclear Factor I (NFI) gene family member, Nfix, as a novel regulator of HSPC function post-BMT. Loss of Nfix severely curtailed the ability of HSPC to reconstitute ablated mice. Further, Nfix-deficient HSPC display increased apoptosis post-BMT, lack CFU potential, and are reduced in number in recipient bone marrow. As the expression of multiple apoptotic regulators is perturbed in the presence and absence of Nfix in HSPC, we propose that Nfix is a key intrinsic regulator of HSPC survival post-BMT. Here, we will test this hypothesis and also assess a role for Nfix in native hematopoiesis, according to the following specific aims: 1) to test the hypothesis that Nfix regulates HSPC survival post-BMT, 2) to identify the molecular targets of Nfix in HSPC, and 3) to test if Nfix is required during native hematopoiesis. For Aim 1, we will employ mice deficient in key cell death pathway regulator genes to test if apoptosis is critical to the loss of Nfix-deficient HSPC post-BMT. As Bcl-xL is downregulated in Nfix-deficient HSPC, we will also test if restored Bcl-xL expression rescues the repopulating defect of Nfix-deficient HSPC. Finally, Nfix overexpression greatly prolongs the ex vivo culture of primary hematopoietic cells. Thus, we will also test if Nfix overexpression protects HSPC from apoptosis ex vivo. For Aim 2, we will leverage the transcriptomes of HSC lacking Nfix, HSC overexpressing Nfix, and NFIX ChIP-seq binding patterns in primitive hematopoietic cell lines to identify candidate downstream targets of Nfix. These candidates will be functionally validated via shRNA-mediated gene knockdown in HSPC overexpressing Nfix for an obligate role in promoting the Nfix- dependent survival of these cultures. We predict that genes necessary here will also play a key role down- stream of Nfix in HSPC post-BMT. Finally, in Aim 3, we will analyze Nfix+/+, Nfixfl/+, and Nfixfl/flROSA26Cre-ERT2 bone marrow chimeras for perturbations in blood lineages, HSPC bone marrow compartments, and HSC quiescence following Nfix deletion. Treatment with tamoxifen efficiently deletes Nfix from the hematopoietic compartment in these mice. By defining Nfix's role as an HSC regulator, we will improve our understanding of HSC and identify new pathways that can be targeted to enhance marrow engraftment after BMT.

项目总结