AML1 IN NORMAL AND LEUKEMIC CELLS

正常细胞和白血病细胞中的 AML1

• 批准号: 6103242
• 负责人: JAMES R DOWNING
• 金额: $ 19.77万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-06-01 至 2000-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6103242
• 关键词: carcinogenesis; developmental genetics; fusion gene; gene expression; gene targeting; hematopoiesis; laboratory mouse; neoplasm /cancer genetics; oncoproteins; pediatric neoplasm /cancer; tissue /cell culture; transcription factor

The AML-1/CBFbeta transcription factor complex is the most frequent target of translocations in human leukemia and is altered in one-third of acute myeloid and lymphoblastic leukemias. AML-1 binds to the enhancer core DNA sequence and is believed to play a critical role in the lineage-specific expression of a number of myeloid and T-cell specific genes. To investigate the normal function of AML-1, we generate AML-1- deficient mice by homologous recombination. Our preliminary results demonstrate that AML1-/- embryos have normal morphogenesis and yolk sac- derived hematopoiesis, but have a complete absence of fetal liver hematopoiesis, and die from hemorrhages during mid-embryonic development. These data suggest that AML-1/CBFbeta plays a pivotal role in regulating transcription of genes that are essential for definitive hematopoiesis. Our hypothesis is that leukemia-associated alterations of this complex lead to disruption of AML-1-mediated signals and result in abnormal hematopoietic development and eventual leukemia. In this project we have proposed a series of experiments to investigate the normal functions of AML-1, and to determine the effects of the t(8;21)-encoded AML-1/ETO products on these functions. First we will define where the defect resulting from the loss of AML-1 lies at a cellular level within the hematopoietic developmental hierarchy. This will involve an evaluation of the hematopoietic activity of yolk sacs progenitors from AML1-/- embryos, and determination of the differentiation potential of AML1-/- ES cells, both in vivo in chimeric mice, and in vitro using cultures of embryoid bodies and two-step hematopoietic colony assays. We will next investigate the role of AML-1 in adult hematopoiesis by use of Cre-loxP- mediated selective gene targeting of AML1 in postnatal development. Lastly, we will investigate the effects of the t(8;21)-encoded AML-1/ETO chimeric product on normal hematopoiesis. This will include an analysis of the ability of AML-1/ETO to rescue the defects resulting from the loss of AML-1, and of the consequences of germline transmission of AML-1/ETO. Together these studies will provide valuable insights into the normal functions of AML-1 and help to elucidate how disruption of these functions leads to leukemogenesis.

AML-1/CBFbeta 转录因子复合物是最常见的 人类白血病易位的目标，三分之一发生改变 急性髓系白血病和淋巴细胞白血病。 AML-1 结合 增强子核心DNA序列被认为在增强子核心DNA序列中发挥着关键作用 许多骨髓和 T 细胞特异性的谱系特异性表达 基因。 为了研究 AML-1 的正常功能，我们生成了 AML-1- 通过同源重组产生缺陷小鼠。 我们的初步结果 证明 AML1-/- 胚胎具有正常的形态发生和卵黄囊- 衍生造血，但完全没有胎儿肝脏 造血，并在胚胎中期发育过程中因出血而死亡。 这些数据表明 AML-1/CBFbeta 在调节中发挥着关键作用 对确定的造血作用至关重要的基因的转录。 我们的假设是，该复合体与白血病相关的改变 导致 AML-1 介导的信号破坏并导致异常 造血发育并最终导致白血病。 在这个项目中我们有 提出了一系列实验来研究正常功能 AML-1，并确定 t(8;21) 编码的 AML-1/ETO 的影响 具有这些功能的产品。 首先我们要定义缺陷在哪里 AML-1 缺失导致的结果是在细胞水平上 造血发育等级。 这将涉及评估 AML1-/- 卵黄囊祖细胞的造血活性 胚胎和AML1-/-分化潜能的测定 ES细胞，无论是在嵌合小鼠体内，还是在体外使用培养物 拟胚体和两步造血集落测定。我们接下来 使用 Cre-loxP- 研究 AML-1 在成人造血中的作用 介导出生后发育中 AML1 的选择性基因靶向。 最后，我们将研究 t(8;21) 编码的 AML-1/ETO 的影响 对正常造血作用的嵌合产物。 这将包括分析 AML-1/ETO 挽救因丢失而导致的缺陷的能力 AML-1，以及 AML-1/ETO 种系传播的后果。 这些研究将共同​​为正常情况提供有价值的见解。 AML-1 的功能并有助于阐明如何破坏这些 功能导致白血病发生。