Thrombopoietin initiates demethylation-based transcription of GP6 during megakaryocyte differentiation
Thrombopoietin initiates demethylation-based transcription of GP6 during megakaryocyte differentiation
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DOI：
10.1182/blood-2004-08-3109
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发表时间：
2005-05-15
期刊：
影响因子：
20.3
通讯作者：
Kunicki, TJ
中科院分区：
文献类型：
--
作者：
Kanaji, S;Kanaji, T;Kunicki, TJ
糖蛋白VI(GPVI)是一种必需的胶原蛋白受体，仅在巨核系中表达。人类基因GP6的转录主要由GATA结合蛋白1(GATA-1)、特异性蛋白1(Sp1)和Friend白血病整合蛋白1(Fli-1)驱动。在这篇报道中，我们发现巨核细胞分化过程中GPVI的表达依赖于胞嘧啶-磷酸-鸟苷(CpG)去甲基化，而胞嘧啶-磷酸-鸟苷(CpG)去甲基化可由促血小板生成素(TPO)启动。亚硫酸氢钠基因组测序证实，在GPVI不表达的细胞系，如UT-7/EPO和C8161中，GP6启动子区域的一个富含CpG的岛在10个CpG位完全甲基化，但在GIRVI表达的细胞系，包括UT-7/TPO和CHRF288-11，完全没有甲基化。为了进一步证实CpG去甲基化与原代细胞中GPVI表达的关系，我们用TPO处理人脐血细胞。GP6启动子在脐带血单个核细胞(祖细胞)中高度甲基化，但在TPO分化后获得的CD41(+)富集性细胞中不存在。此外，当TPO处理稳定表达人C-髓增殖性白血病病毒配体(c-MPI)的UT-7/EPO-MPI细胞时，可诱导GP6启动子去甲基化。在每一种情况下，GP6启动子的去甲基化都与mRNA水平的增加相关。因此，GP6基因的巨核细胞特异性表达在一定程度上受到CpG去甲基化的调节，而CpG去甲基化可以直接由TPO启动。
Glycoprotein VI (GPVI) is an essential platelet receptor for collagens that is exclusively expressed in the megakaryocytic lineage. Transcription of the human gene GP6 is driven largely by GATA-binding protein 1 (GATA-1), specificity protein 1 (Sp1), and Friend leukemia integration 1 (Fli-1). In this report, we show that GPVI expression during megakaryocytic differentiation is dependent on cytosine-phosphate-guanosine (CpG) demethylation that can be initiated by thrombopoietin (TPO). Sodium bisulfite genomic sequencing established that a CpG-rich island within the GP6 promoter region is fully methylated at 10 CpG sites in GPVI-nonexpressive cell lines, such as UT-7/EPO and C8161, but completely unmethylated in GIRVI-expressive cell lines, including UT-7/TPO and CHRF288-11. To further confirm the relationship between CpG demethylation and expression of GPVI in primary cells, we treated human cord blood cells with TPO. The GP6 promoter is highly methylated in cord blood mononuclear cells (progenitors) but not in CD41(+)-enriched cells obtained after TPO differentiation. Furthermore, when UT-7/EPO-MpI cells, which stably express human C-myeloproliferative leukemia virus ligand (c-MpI), were treated with TPO, demethylation of the GP6 promoter was induced. In every case, demethylation of the GP6 promoter correlated with an increase in mRNA level. Thus, megakaryocyte-specific expression of the GP6 gene is regulated, in part, by CpG demethylation, which can be directly initiated by TPO.