Role of the zinc finger transcription factor ZBP89 in blood & vascular developmen

锌指转录因子 ZBP89 在血液中的作用

• 批准号: 7796572
• 负责人: M. AMIN ARNAOUT
• 金额: $ 41.58万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7796572
• 关键词: Adult; Alleles; Anemia; B-Lymphocytes; Biochemical; Biological Assay; Biotinylation; Blast Cell; Blood; Blood Cells; Blood Vessels; Bone Marrow; Bone Marrow Cells; Bone Marrow Transplantation; C2H2 Zinc Finger; CCAAT-Enhancer-Binding Protein-alpha; Candidate Disease Gene; Cardiac; Cell Culture Techniques; Cell Line; Cell Lineage; Cells; Chronic; Commit; Complementary DNA; Data; Defect; Development; EMSA; ES Cell Line; Ectopic Expression; Embryo; Endothelial Cells; Erythrocytes; Erythroid; Exhibits; Failure; Fishes; Gene Expression; Generations; Genes; Genetic; Genomics; Hand; Helix-Turn-Helix Motifs; Hematopoiesis; Hematopoietic; Hematopoietic System; Hematopoietic stem cells; Human; Hybrids; ITGAM gene; In Vitro; Inflammation; Injection of therapeutic agent; Integrins; KDR gene; Knock-out; Knockout Mice; L-Plastin; Leukocytosis; Link; Lymphoid; MYB gene; Mediating; Mesoderm; Methods; Modeling; Molecular; Mus; Mutagenesis; Myelogenous; Myeloid Cells; Myocardial Infarction; Nature; Orthologous Gene; PECAM1 gene; PTPRC gene; Perinatal mortality demographics; Phenotype; Play; Population; Process; Program Development; Proteins; Proteomics; Proto-Oncogene Proteins c-myb; RNA Caps; Radiation; Reporter; Role; Series; Specific qualifier value; Staging; Stem cells; Transfusion; Transgenic Organisms; Transplantation; Undifferentiated; Yeasts; Zebrafish; Zinc Fingers; angiogenesis; cadherin 5; cell type; chemotherapy; developmental plasticity; embryonic stem cell; fetal; in vivo; macrophage; monocyte; mutant; neutrophil; novel strategies; overexpression; progenitor; programs; promoter; protein complex; public health relevance; reconstitution; repaired; self-renewal; small hairpin RNA; transcription factor; tumor

DESCRIPTION (provided by applicant): The hematopoietic system originates from a small population of self-renewing hematopoietic stem cells (HSCs) that in turn derive from hemangioblasts, common precursors of blood and blood vessels. Extensive studies have identified key transcription factors, such as the basic helix loop helix (bLH) SCL/TAL1 and GATA factors, which play critical roles in the successive steps of differentiation of HSCs into the erythroid, myeloid and lymphoid lineages. Despite these advances, the nature of the factors that commit the hemangioblast to blood and endothelial cells are obscure and the transcriptional networks operative in HSC self-renewal and lineage commitment remain incomplete. In preliminary studies, we have identified the Kruppel-like zinc finger transcription factor ZBP-89 as a master regulator of early development of FLK1+ mesoderm into blood and blood vessels. Forced expression of ZBP-89 increased expression of primitive and definitive hematopoiesis in zebrafish and in mouse embryonic stem cells (ESCs)/embryonic body (EB) cultures, but reduced axial and intersomitic blood vessel formation in zebrafish embryos and the endothelial replating potential and sprouting angiogenesis in mouse EB cultures. Knockdown of ZBP-89 in zebrafish or in mouse ESCs resulted in a dramatic reduction in primitive and definitive hematopoietic markers (e.g. SCL and GATA factors), but an increase in endothelial lineage markers, distinguishing the mechanism of action of ZBP-89 from all other known transcription factors regulating hematovascular development. Further, we find that mouse pubs homozygous for a hypomorphic ZBP-89 mutant allele die perinatally and exhibit a marked reduction in circulating red blood cells, but an increase in mature myeloid cells, reflecting an additional critical role for ZBP-89 in fetal hematopoiesis, perhaps at the level of the bipotential Common Myeloid Progenitor (CMP) stem cell, which gives rise to both the erythroid and myeloid lineages. The effects of ZBP-89 modulation on adult hematopoiesis and the underlying transcriptional networks involved in its action at the level of hemangioblasts and CMPs are unexplored. In this application, we propose to assess the consequences of loss of ZBP-89 on fetal and adult hematopoiesis and vascular development in vivo (Aim 1), determine the effects of its ectopic expression on blood and vessel lineage development in adult mice and zebrafish (Aim 2), and elucidate the mechanism(s) underlying ZBP-89- mediated hematopoietic lineage commitment. (Aim 3). Genetic, biochemical, proteomics, genomics, stem cell cultures and bone marrow transplantation in mouse and zebrafish will be utilized. PUBLIC HEALTH RELEVANCE: Formation of all blood cells is closely linked developmentally to formation of blood vessels and the two processes are regulated in large part by transcription factors, proteins that control the decision of a common undifferentiated stem cell precursor to become a blood cell or a vascular cell. The instructional network that underlies this cell fate decision remains ill defined, despite its potential impact on development of new therapies. For example, blood stem cells are being used to reconstitute bone marrow damaged by radiation or chemotherapy, but they are very few in numbers, a major challenge in cellular therapy. Increasing their number by enhancing their development from their stem cell precursors will serve a critical need. Methods for enhancing or suppressing new blood vessel formation (angiogenesis) may also be useful respectively, in cardiac repair following a heart attack or in reducing harmful angiogenesis (induced by tumors or chronic inflammation). We have discovered a transcription factor that lies at the cross road of blood and blood vessel development. We are proposing a series of studies to pinpoint the role of this factor, and define the developmental networks it coordinates.

描述（由申请人提供）：造血系统起源于一小群自我更新的造血干细胞（HSC），而这些细胞又源自成血管细胞，即血液和血管的常见前体。广泛的研究已经确定了关键的转录因子，例如基本螺旋环螺旋 (bLH) SCL/TAL1 和 GATA 因子，它们在 HSC 分化为红系、髓系和淋巴系的连续步骤中发挥着关键作用。尽管取得了这些进展，但将成血管细胞定型为血液和内皮细胞的因素的性质尚不清楚，并且在 HSC 自我更新和谱系定型中起作用的转录网络仍然不完整。在初步研究中，我们已经确定 Kruppel 样锌指转录因子 ZBP-89 是 FLK1+ 中胚层早期发育成血管的主要调节因子。 ZBP-89的强制表达增加了斑马鱼和小鼠胚胎干细胞（ESC）/胚胎体（EB）培养物中原始和确定性造血的表达，但减少了斑马鱼胚胎中的轴向和体间血管形成以及小鼠EB培养物中的内皮再增殖潜力和萌芽血管生成。在斑马鱼或小鼠ESC中敲低ZBP-89会导致原始和决定性造血标记物（例如SCL和GATA因子）显着减少，但内皮谱系标记物增加，这将ZBP-89的作用机制与所有其他已知的调节血管发育的转录因子区分开来。此外，我们发现，ZBP-89 亚型突变等位基因纯合的小鼠 pubs 在围产期死亡，并表现出循环红细胞显着减少，但成熟骨髓细胞增加，反映出 ZBP-89 在胎儿造血中的额外关键作用，可能是在双能共同骨髓祖细胞 (CMP) 干细胞的水平上，该干细胞产生 红系和髓系谱系。 ZBP-89 调节对成体造血作用的影响以及其在成血管细胞和 CMP 水平上的作用所涉及的潜在转录网络尚未被探索。在本申请中，我们建议评估 ZBP-89 缺失对胎儿和成人造血和血管发育的体内影响（目标 1），确定其异位表达对成年小鼠和斑马鱼的血液和血管谱系发育的影响（目标 2），并阐明 ZBP-89 介导的造血谱系定向的潜在机制。 （目标 3）。将利用小鼠和斑马鱼的遗传、生物化学、蛋白质组学、基因组学、干细胞培养和骨髓移植。公共健康相关性：所有血细胞的形成在发育上都与血管的形成密切相关，这两个过程在很大程度上受到转录因子和蛋白质的调节，转录因子控制着共同的未分化干细胞前体成为血细胞或血管细胞的决定。尽管其对新疗法的开发具有潜在影响，但作为细胞命运决定基础的指导网络仍然不明确。例如，造血干细胞被用来重建因放疗或化疗而受损的骨髓，但它们的数量很少，这是细胞治疗的一大挑战。通过增强干细胞前体的发育来增加它们的数量将满足迫切的需求。用于增强或抑制新血管形成（血管生成）的方法也可分别用于心脏病发作后的心脏修复或减少有害的血管生成（由肿瘤或慢性炎症诱导）。我们发现了一种位于血液和血管发育十字路口的转录因子。我们正在提出一系列研究来查明这个因素的作用，并定义它协调的发展网络。