Hematopoietic Commitment: Molecular Mechanisms

造血承诺：分子机制

• 批准号: 7002346
• 负责人: KYUNGHEE CHOI
• 金额: $ 29.88万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7002346
• 关键词: AP1 protein; NOD mouse; SCID mouse; activins; angiogenesis; biological signal transduction; bone morphogenetic proteins; cell differentiation; chromatin immunoprecipitation; developmental genetics; embryogenesis; embryonic stem cell; enzyme activity; fibroblast growth factor; gene expression; genetic regulation; genetically modified animals; growth factor receptors; hematopoiesis; hematopoietic stem cells; protein protein interaction; protein structure function; transcription factor; vascular endothelial growth factors

DESCRIPTION (provided by applicant): Our long-term goal is to understand how the hematopoietic system is established in the developing embryo. Our studies of in vitro differentiated embryonic stem (ES) cells indicate that the hematopoietic system is established via distinct, sequentially generated Flk-1 and SCL-expressing cells. Flk-I+SCL - cells first arise in developing embryoid bodies (EBs, in vitro differentiated progeny of ES cells). The Scl gene is turned on within Flk-1 + SCL- cells to give rise to Flk-1 + SCL + cells. The hemangioblast cell population, a common progenitor of hematopoietic and endothelial cells, was enriched within these cells. Within Flk-I+SCL + cells, Flk-1 is down regulated to finally generate Flk-I-SCL + hematopoietic progenitors. A serum free in vitro differentiation model of ES cells has identified that bone morphogenetic protein (BMP) -4 is critical in the generation of Flk-1 + and SCL + cells. Many studies implicate the importance of BMP-4 in hematopoietic development. However, none of the studies examined the precise developmental stage in which the BMP-4 functions. Our proposal is to further characterize molecular mechanisms controlling Flk-1 and Scl expression and to determine the precise developmental stage where BMP-4 functions to establish the hematopoietic system. Specific aim I: Further characterize molecular events involved in the generation of Flk-1 + and SCL + cells. Our studies indicate that the activation of both map kinase and Smadl pathways by BMP-4 is critical for the generation of Flk-1 + cells. Furthermore, recent studies suggest that the transcription factor GATA-2 is one component that binds Scl enhancer region. Thus, our aim is to further understand mechanisms controlling Flk-1 and Scl gene expression. Specific aim II. Examine the role of BMP-4 mediated signals in hemangioblast development. Our experiments indicate that BMP-4 is critical for the generation of Flk-1 + cells. What is not clear from our current studies is whether BMP-4 mediated signals are still required after the Flk-I + cell stage in establishing hematopoiesis. Our specific aim is to utilize Flk-l-cre mice and conditional Alk-3 (BMP-4 receptor) knockout mice to determine whether BMP-4 mediated signals are obligatory for the generation of the hemangioblast within the Flk-1 + cells. Specific aim III. Examine the role of BMP-4 mediated signals in hematopoietic and endothelial cell development. Subsequent to hemangioblast development, hematopoietic and endothelial cell lineage commitment commences. Our specific aim is to determine whether BMP-4 mediated signals are required for hematopoietic lineage commitment from the hemangioblast. Scl-cre mice and conditional Alk-3 knockout mice will be utilized.

描述（由申请人提供）：我们的长期目标是了解造血系统是如何在发育中的胚胎中建立的。我们对体外分化的胚胎干细胞（ES）的研究表明，造血系统是通过不同的，顺序产生的Flk-1和SCL表达细胞建立的。Flk-1 +SCL -细胞首先出现在发育中的胚状体（EB，ES细胞的体外分化后代）中。Scl基因在Flk-1 + SCL-细胞内被打开以产生Flk-1 + SCL +细胞。成血管细胞群是造血细胞和内皮细胞的共同祖细胞，在这些细胞中富集。在Flk-1 +SCL +细胞内，Flk-1下调以最终产生Flk-1-SCL +造血祖细胞。ES细胞的无血清体外分化模型已经确定骨形态发生蛋白（BMP）-4在Flk-1 +和SCL +细胞的产生中是关键的。许多研究暗示BMP-4在造血发育中的重要性。然而，没有一项研究检查了BMP-4发挥作用的精确发育阶段。我们的建议是进一步表征控制Flk-1和Scl表达的分子机制，并确定BMP-4功能建立造血系统的精确发育阶段。 具体目标I：进一步表征参与Flk-1 +和SCL +细胞生成的分子事件。我们的研究表明，BMP-4激活MAP激酶和Smad 1通路对于Flk-1 +细胞的产生至关重要。此外，最近的研究表明，转录因子加塔-2是结合Scl增强子区的一个组分。因此，我们的目标是进一步了解Flk-1和Scl基因表达的调控机制。 具体目标二。检查BMP-4介导的信号在成血管细胞发育中的作用。我们的实验表明BMP-4对于Flk-1 +细胞的产生是至关重要的。从我们目前的研究中尚不清楚的是，在Flk-I +细胞阶段之后，在建立造血中是否仍然需要BMP-4介导的信号。我们的具体目标是利用Flk-1-cre小鼠和条件性Alk-3（BMP-4受体）敲除小鼠来确定BMP-4介导的信号是否对于Flk-1 +细胞内的成血管细胞的产生是必需的。 具体目标三。检查BMP-4介导的信号在造血和内皮细胞发育中的作用。继成血管细胞发育之后，造血和内皮细胞谱系定型开始。我们的具体目标是确定BMP-4介导的信号是否需要从成血管细胞造血谱系承诺。将使用Scl-cre小鼠和条件性Alk-3敲除小鼠。