5-AZACYTIDINE-INDUCED REACTIVATION OF THE HUMAN X-CHROMOSOME-LINKED PGK1 GENE IS ASSOCIATED WITH A LARGE REGION OF CYTOSINE DEMETHYLATION IN THE 5' CPG ISLAND
5-AZACYTIDINE-INDUCED REACTIVATION OF THE HUMAN X-CHROMOSOME-LINKED PGK1 GENE IS ASSOCIATED WITH A LARGE REGION OF CYTOSINE DEMETHYLATION IN THE 5' CPG ISLAND
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DOI：
10.1073/pnas.87.11.4174
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发表时间：
1990-06-01
影响因子：
11.1
通讯作者：
GARTLER, SM
中科院分区：
文献类型：
--
作者：
HANSEN, RS;GARTLER, SM
用5-氮杂胞苷处理含有失活的人X染色体的仓鼠-人细胞杂交体，并检查衍生克隆的人PGK 1（X染色体连锁磷酸甘油酸激酶）基因中的磷酸甘油酸激酶活性和胞嘧啶甲基化。表达和非表达克隆之间的比较表明，5“CpG岛以外的几个甲基化敏感性限制性位点的去甲基化对于表达是不必要的。和Tha I位点显示，所有表达PGK 1的克隆在包括转录起始位点和400个碱基对的CpG岛的大区域中未甲基化上游许多非表达克隆具有不连续的去甲基化模式。在非表达杂交体的亚克隆中经常观察到再甲基化。这些数据表明，PGK 1启动子内的甲基化自由DNA的特定区域是转录所必需的。此外，邻近甲基胞嘧啶的存在似乎降低了该区域中未甲基化CpG的遗传稳定性。
Hamster-human cell hybrids containing an inactive human X chromosome were treated with 5-azacytidine and derived clones were examined for phosphoglycerate kinase activity and cytosine methylation in the human PGK1 (X chromosome-linked phosphoglycerate kinase) gene. Comparisons between expressing and nonexpressing clones indicated that demethylation of several methylation-sensitive restriction sites outside of the 5'' CpG island were unnecessary for expression High-resolution polyacrylamide gel analysis of 25 Hpa II, Hha I, and Tha I sites revealed that all clones expressing PGK1 were unmethylated in a large region of the CpG island that includes the transcription start site and 400 base pairs upstream. Many nonexpressing clones had discontinuous patterns of demethylation. Remethylation was often observed in subclones of nonexpression hybrids. These data suggest that a specific zone of methylation-free DNA within the PGK1 promoter is required for transcription. In addition, the presence of neighboring methylcytosines appears to decrease the heritable stability of unmethylated CpGs in this region.