Functional analysis of AML1 gene in normal hematopoietic cells and leukemia cells

正常造血细胞和白血病细胞中AML1基因的功能分析

基本信息

批准号：
07457229
负责人：
HIRAI Hisamaru
金额：
$ 4.48万
依托单位：
University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

The AML1 gene on chromosome 21 is disrupted in the (8 ; 21) (q22 ; q22) and (3 ; 21) (q26 ; q22) translocations associated with myelogenous leukemias and encodes a transcription factor. From AML1 gene, two representative forms of proteins, AML1a and AML1b, are produced by an alternative splicing. Overexpressed AML1a totally suppresses granulocytic differentiation and stimulates cell proliferation in 32Dc13 murine myeloid cells treated with granulocyte colony-stimulating factor. These effects by AML1a were canceled by the concomitant overexpression of AML1b. Such biological phenomena could be explained by our observations that AML1a, which solely has no effects as a transcriptional regulator, dominantly suppresses transcriptional activation by AML1b, and that AML1a exhibits the higher affinity for DNA-binding than AML1b. These antagonistic actions could be important for leukemogenesis and/or myeloid cell differentiation.We also investigate the regulatory mechanisms of AML1 functions through signal transduction pathways. AML1 is phosphorylated in vivo on two serine residues within the proline-, serine- and threonine-rich region, with dependence on the activation of extracellular signal-regulated kinase (ERK). These in vivo phosphorylation sites of AML1 were directly phosphorylated in vitro by ERK.Although alterations in the DNA-binding affinity were not observed between wild AML1 and non-phosphorylated mutants, we have shown that ERK-dependent phosphorylation potentiates the transactivation ability of AML1. Furthermore the phosphorylation site-mutations reduced the transforming capacity of AML1 in fibroblast cells. These data suggest that AML1 functions are regulated by ERK which is activated by cytokine or growth facotr-stimuli. This study would give important clues to clarify unidentified facets of regulatory mechanisms of AML1 function.

在（8; 21）（q22; q22）和（3; 21）（q26; q22）易位与骨髓性白血病相关的易位（Q22; Q22）和（3; 21）（Q22）和（3; Q22）（Q22; Q22）和（3; Q22）（Q22; Q22）中的AML1基因受到破坏并编码转录因子。从AML1基因中，蛋白质的两种代表性形式AML1A和AML1B由替代剪接产生。过表达的AML1A完全抑制粒细胞分化并刺激32DC13用粒细胞菌落刺激因子处理的32DC13鼠髓样细胞中的细胞增殖。 AML1A的这些影响通过AML1B的过表达取消。这种生物学现象可以通过我们的观察结果来解释，即AML1a仅作为转录调节剂没有效果，主要抑制AML1B的转录激活，而AML1A比AML1B表现出对DNA结合的亲和力更高。这些拮抗作用对于白血病和/或髓样细胞分化可能很重要。我们还通过信号转导途径研究了AML1功能的调节机制。 AML1在脯氨酸，丝氨酸和苏氨酸富含区域内的两个丝氨酸残基上在体内磷酸化，并依赖于细胞外信号调节激酶（ERK）的激活。这些AML1的体内磷酸化位点在体外被ERK直接磷酸化。尽管在野生AML1和非磷酸化突变体之间未观察到DNA结合亲和力的改变，我们已经表明ERK依赖性的磷酸化磷酸化能力增强了AML1的过渡能力。此外，磷酸化位点 - 突变降低了成纤维细胞中AML1的转化能力。这些数据表明AML1函数受ERK的调节，ERK被细胞因子或生长FACOTR-stimuli激活。这项研究将提供重要的线索，以阐明AML1功能的调节机制的身份不明的方面。