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

• 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

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.

AML 1（PEBP 2 αB）是人类急性白血病最常发生突变的基因之一，编码异源二聚体转录因子复合物的DNA结合亚基，核心结合因子CBF（或多瘤增强子结合蛋白2 PEBP 2）。AML 1或其二聚化伴侣CBFβ（PEBP 2 β）的破坏导致继发于胎肝造血完全阻断的胚胎致死性，表明该转录复合物在永久性造血发育中的重要作用。由AML 1缺失导致的造血表型可以使用小鼠胚胎干细胞（ES）的两步培养系统在体外复制。使用这个实验系统，我们证明了这种造血缺陷可以通过敲入（靶向插入）方法在内源性AML 1调控序列下表达PEBP 2 αB1（AML 1b）同种型来挽救。此外，我们发现这些拯救的AML 1-/-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，即AML 1敲除表型仅仅是由于缺乏该基因，并且PEBP 2 αB1（AML 1b）亚型具有生物活性。有趣的是，当AML 1b在异源启动子的控制下表达时，没有观察到造血拯救，这表明AML 1的转录控制对其生物活性很重要。使用这种方法，我们还证明了AML 1b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ES细胞实验系统应作为一个独特的工具，以进一步确定AML 1在正常和白血病造血的生物学特性。

项目成果

Okuda, T.: "Role of AML1 in normal and leukemic hematopoieses. In "Moleculau Target for Hematological Malignancies and Cancer" (Ed. by Niho Y.)"Kyushu University Press (Fukuoka, Japan) (in press). (2000)

Okuda, T.：“AML1 在正常和白血病造血中的作用。见“血液恶性肿瘤和癌症的分子靶点”（Niho Y. 编）”九州大学出版社（日本福冈）（正在出版）。

奥田司: "AML1遺伝子の生物学的意義."現代医療. 31. 1125-1134 (1999)

Tsukasa Okuda：“AML1 基因的生物学意义。” 现代医学 31. 1125-1134 (1999)

Okuda, T.: "Biological characteristics of the leukemia-associated transcriptional factor AML1 disclosed by hematopoietic rescue of AML1-deficient embryonic stem cells by using a knock-in strategy"Molecular and Cellular Biology. 20. 319-328 (2000)

Okuda, T.：“通过使用敲入策略对 AML1 缺陷型胚胎干细胞进行造血拯救，揭示了白血病相关转录因子 AML1 的生物学特征”《分子和细胞生物学》。

奥田司: "造血初期発生制御における転写因子AML1の役割"実験医学. 17・9. 1136-1143 (1999)

奥田司：“转录因子AML1在调节早期造血发育中的作用”实验医学17・9（1999）。