Kanno Y, Kanno T, Sakakura C, Bae S C, Ito Y
Department of Viral Oncology, Institute for Virus Research, Kyoto University, Kyoto 606, Japan.
Mol Cell Biol. 1998 Jul;18(7):4252-61. doi: 10.1128/MCB.18.7.4252.
The polyomavirus enhancer binding protein 2 (PEBP2)/core binding factor (CBF) is a transcription factor composed of two subunits, alpha and beta. The gene encoding the beta subunit is disrupted by inv(16), resulting in the formation of a chimeric protein, beta-SMMHC, which is associated with acute myelogenous leukemia. To understand the effect of beta-SMMHC on PEBP2-mediated transactivation, we used a luciferase assay system in which contribution of both the alpha and beta subunits was absolutely required to activate transcription. Using this system, we found that the minimal region of the beta subunit required for transactivation resides between amino acid 1 and 135, which is known to dimerize with the alpha subunit. In contrast, beta-SMMHC, despite having this minimal region for dimerization and transactivation, failed to support transcription with the alpha subunit. Furthermore beta-SMMHC blocked the synergistic transcription achieved by PEBP2 and CCAAT/enhancer binding protein alpha. By using a construct in which the PEBP2 alpha subunit was fused to the glucocorticoid receptor ligand binding domain, we demonstrated that coexpressed beta-SMMHC tightly sequestered the alpha subunit in the cytoplasm and blocked dexamethasone-dependent nuclear translocation of the alpha subunit. Thus, the result suggess that beta-SMMHC inhibits PEBP2-mediated transcription via cytoplasmic sequestration of the alpha subunit. Lastly proliferation of ME-1 cells that harbor inv(16) was blocked by an antisense oligonucleotide complementary to the junction of the chimeric mRNA, suggesting that beta-SMMHC contributes to leukemogenesis by blocking the differentiation of myeloid cells.
多瘤病毒增强子结合蛋白2（PEBP2）/核心结合因子（CBF）是一种由α和β两个亚基组成的转录因子。编码β亚基的基因被inv(16)破坏，导致形成一种嵌合蛋白β-SMMHC，它与急性髓性白血病相关。为了了解β-SMMHC对PEBP2介导的反式激活的影响，我们使用了一种荧光素酶检测系统，其中α和β亚基对激活转录都是绝对必需的。利用这个系统，我们发现反式激活所需的β亚基的最小区域位于氨基酸1至135之间，已知该区域与α亚基二聚化。相比之下，β-SMMHC尽管具有这个用于二聚化和反式激活的最小区域，但却无法与α亚基一起支持转录。此外，β-SMMHC阻断了PEBP2和CCAAT/增强子结合蛋白α实现的协同转录。通过使用一种将PEBP2α亚基与糖皮质激素受体配体结合域融合的构建体，我们证明共表达的β-SMMHC将α亚基紧密隔离在细胞质中，并阻断了α亚基的地塞米松依赖性核转位。因此，结果表明β-SMMHC通过在细胞质中隔离α亚基来抑制PEBP2介导的转录。最后，携带inv(16)的ME-1细胞的增殖被与嵌合mRNA连接处互补的反义寡核苷酸阻断，这表明β-SMMHC通过阻断髓系细胞的分化促进白血病发生。
