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Expression of human histo-blood group ABO genes is dependent upon DNA methylation of the promoter region

人类组织血型 ABO 基因的表达依赖于启动子区域的 DNA 甲基化

基本信息

批准号：
11670410
负责人：
KOMINATO Yoshihiko
金额：
$ 2.37万
依托单位：
TOYAMA MEDICAL AND PHARMACEUTICAL UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670410/
关键词：
the human ABO blood group genes transcription promoter assay CpG island DNA methylation

项目摘要

We have studied the transcriptional regulatory mechanism of the human histo-blood group ABO genes. The promoter assay using transient transfection showed that the sequence just upstream of the transcription start site (cap site) and an enhancer element which is located further upstream (between -3899 and -3618 bp from the transcription initiation site) are responsible for the transcriptional activity of the ABO genes in gastric cancer cells. Electrophoretic mobility shift assays showed that a transcriptional factor, CBF/NF-Y binds the minisatellite, which is essential for the enhancer activity, on between -3899 and -3618 bp from the transcription initiation site. However, the promoter assay demonstrated that the proximal promoter is active in the cells not expressing the ABO genes. Next, mechanism of repression of the ABO genes in tumors was investigated. The promoter region of the ABO genes fulfills the criteria for CpG island, having a G+C content of greater than 60% and having a CpG … More /GpC ratio of at least 0.6. Methylated CpG islands are associated with long-range changes in chromatin, which mediate a closed, nucleosomal conformation, and transcriptional repression. Therefore, we have investigated the role of DNA methylation in the regulation of the expression of the human ABO blood group genes. Studies on the methylation of the ABO blood group gene promoter in cultured cells demonstrated that the promoter is methylated in vivo in the cells not expressing the ABO genes, and that hypomethylation of the promoter is correlated with constitutive gene expression. Demethylation of the promoter in vivo by treatment of the cells with 5-aza-2'-deoxycytidine led to the expression of A-antigens on gastric cancer cells and hypermethylation of the promoter in vitro suppressed the promoter activity. These results indicated that DNA methylation of the ABO gene promoter regulates gene expression, suggesting that alternations in DNA methylation may be one of the mechanisms regulating the loss of ABO antigens in the tumors. Less

我们研究了人类组织血型ABO基因的转录调控机制。瞬时转染的启动子实验表明，在胃癌细胞中，转录起始位点（cap位点）上游的序列和位于转录起始位点-3899和-3618 bp之间的上游增强子元件负责ABO基因的转录活性。电泳迁移位移分析表明，一个转录因子CBF/NF-Y在转录起始位点-3899至-3618 bp之间结合了对增强子活性至关重要的小卫星。然而，启动子实验表明，近端启动子在不表达ABO基因的细胞中是活跃的。接下来，研究ABO基因在肿瘤中的抑制机制。ABO基因的启动子区域符合CpG岛的标准，G+C含量大于60%，CpG…More /GpC比值至少为0.6。甲基化的CpG岛与染色质的长期变化有关，染色质介导封闭的核小体构象和转录抑制。因此，我们研究了DNA甲基化在人类ABO血型基因表达调控中的作用。对培养细胞中ABO血型基因启动子甲基化的研究表明，该启动子在体内不表达ABO基因的细胞中甲基化，并且启动子的低甲基化与组成基因的表达相关。在体内用5-aza-2'-脱氧胞苷处理细胞后，启动子去甲基化导致胃癌细胞上a抗原的表达，在体外启动子的高甲基化抑制了启动子的活性。这些结果表明，ABO基因启动子的DNA甲基化调节基因的表达，提示DNA甲基化的改变可能是调节肿瘤中ABO抗原丢失的机制之一。少