Homeodomain proteins MEIS1 and PBXs regulate the lineage-specific transcription of the platelet factor 4 gene
Homeodomain proteins MEIS1 and PBXs regulate the lineage-specific transcription of the platelet factor 4 gene
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DOI：
10.1182/blood-2002-02-0380
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发表时间：
2003-06-15
期刊：
影响因子：
20.3
通讯作者：
Doi, T
中科院分区：
文献类型：
--
作者：
Okada, Y;Nagai, R;Doi, T
血小板因子4（PF 4）在巨核细胞分化过程中表达。我们以前报道过加塔-1和ETS-1调节大鼠PF 4启动子并反式激活PF 4基因。在本研究中，我们研究了负责PF 4基因谱系特异性表达的调控元件及其转录因子。对缺失构建体的启动子活性进行了评价，并定义了一个新的调控元件TME（tandem repeat of MEIS 1 binding element）（-219 to -182）。电泳迁移率变动分析（EMSA）在HEL细胞核提取物中检测到与TME结合的蛋白，并通过DNA亲和层析纯化。通过进行蛋白质印迹和supershift测定，结合蛋白被鉴定为同源结构域蛋白，MEIS 1，PBX 1B和PBX 2。这些因子在人脐带血中由CD 34（+）细胞分化而来的巨核细胞中表达。MEIS 1和PBX作为MEIS 1/PBX复合物与TME结合并激活PF 4启动子。在非巨核细胞HepG 2细胞中，加塔-1和ETS-1激活PF 4启动子约10倍。令人惊讶的是，我们发现MEIS 1和PBX 2两者的额外表达使这种主要激活增加了2倍。当TME中的MEIS 1结合位点被破坏时，没有观察到这种激活。此外，在巨核细胞HEL细胞中，内源性MEIS 1/PBX复合物与TME的结合的抑制使启动子活性降低了几乎一半。因此，这些研究表明同源结构域蛋白MEIS 1、PBX 1B和PBX 2在巨核细胞基因表达中起重要作用。(C)2003年，美国血液学会。
Platelet factor 4 (PF4) is expressed during megakaryocytic differentiation. We previously reported that GATA-1 and ETS-1 regulate the rat PF4 promoter and transactivate the PF4 gene. For the present study, we investigated the regulatory elements and their transcription factors responsible for the lineage-specific expression of the PF4 gene. The promoter activities of deletion constructs were evaluated, and a novel regulatory element termed TME (tandem repeat of MEIS1 binding element) (-219 to -182) was defined. Binding proteins to TME were strongly detected in HEL nuclear extracts by electrophoresis mobility shift assay (EMSA), and they were purified by DNA affinity chromatography. By performing Western blottings and supershift assays, the binding proteins were identified as homeodomain proteins, MEIS1, PBX1B, and PBX2. These factors are expressed in megakaryocytes differentiated from CD34(+) cells in human cord blood. MEIS1 and PBXs bind to the TME as MEIS1/PBX complexes and activate the PF4 promoter. In non-megakaryocytic HepG2 cells, GATA-1 and ETS-1 activate the PF4 promoter approximately 10-fold. Surprisingly, we found that additional expression of both MEIS1 and PBX2 multiplied this major activation another 2-fold. This activation was not observed when MEIS1 binding sites in the TME were disrupted. Furthermore, inhibition of the binding of endogenous MEIS1/PBX complexes to the TME decreased the promoter activity by almost one half, in megakaryocytic HEL cells. Thus, these studies demonstrate that the homeodomain proteins, MEIS1, PBX1B, and PBX2, play an important role in megakaryocytic gene expression. (C) 2003 by The American Society of Hematology.