Biologic characterization of leukemia-associated transcription factor AML1 (PEBP2αB) in hematopoiesis

白血病相关转录因子 AML1 (PEBP2αB) 在造血过程中的生物学特征

基本信息

批准号：
11138251
负责人：
OKUDA Tsukasa
金额：
$ 4.8万
依托单位：
Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas (A)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至无数据
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11138251/
关键词：
AML1 PEBP2 leukemia hematopoiesis transcription embryonic stem (ES) cell oncogene gene targeting

项目摘要

AML1 (PEBP2αB) is one of the most frequently mutated genes associated with human acute leukemia, and it encodes the DNA-binding subunit of the heterodimering transcriptional factor complex, core binding factor : CBF (or polyoma enhancer binding protein 2 : PEBP2). Disruption of either AML1 or its dimerizing partner, CBFβ (PEBP2β), results in the embryonic-lethality secondary to a complete block in fetal liver hematopoiesis, indicating an essential role of this transcription complex in the development of definitive hematopoiesis. The hematopoietic phenotype that results from the loss of AML1 could be replicated in vitro using a two-step culture system of murine embryonic stem (ES) cells. Using this experimental system, we demonstrated that this hematopoietic defect could be rescued by expressing the PEBP2αB1 (AML1b) isoform under the endogenous AML1-regulatory sequences through a knock-in (targeted-insertion) approach. Moreover, we found that these rescued AML1-/-ES cells could contribute to lympho-hematopoiesis within the context of chimeric animals. These results provide compelling evidence that the AML1 knockout phenotype is due solely to the lack of this gene, and that the PEBP2αB1 (AML1b) isoform is biologically active. Interestingly, hematopoietic rescue was not observed when AML1b was expressed under the control of a heterologous promoter, suggesting that the transcriptional control of AML1 is important for its biological activity. Using this assay, we also demonstrated that the transactivation domain of AML1b is required for its biological activity, whereas the conserved VWRPY-motif at the carboxyl terminus is not. The ES cell experimental system developed through the present study should serve as a unique tool to further define the biological properties of AML1 in normal and leukemic hematopoiesis.

AML1 (PEBP2αB) 是与人类急性白血病相关的最常见突变基因之一，它编码异二聚体转录因子复合物、核心结合因子：CBF（或多瘤增强子结合蛋白2：PEBP2）的DNA结合亚基。 AML1 或其二聚化伴侣 CBFβ (PEBP2β) 的缺失会导致继发于胎儿肝脏完全阻断的胚胎致死性造血作用，表明该转录复合物在最终造血作用的发展中发挥着重要作用。使用小鼠胚胎干（ES）细胞的两步培养系统可以在体外复制因 AML1 缺失而产生的造血表型。实验系统中，我们证明了这种造血缺陷可以通过在内源性 AML1 调节序列下通过敲入表达 PEBP2αB1 (AML1b) 同工型来挽救此外，我们发现这些被拯救的 AML1-/-ES 细胞可以在嵌合动物的背景下促进淋巴造血。这些结果提供了令人信服的证据，表明 AML1 敲除表型仅仅是由于缺乏。该基因，并且 PEBP2αB1 (AML1b) 亚型具有生物活性。启动子，表明 AML1 的转录控制对其生物活性很重要，我们还证明了 AML1b 的反式激活结构域是其生物活性所必需的，而羧基末端的保守 VWRPY 基序则不是。通过本研究开发的细胞实验系统应作为进一步确定 AML1 在正常和白血病造血中的生物学特性的独特工具。