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Regulation of the human epsilon-globin gene transcription in the switching mechanism

开关机制中人ε-珠蛋白基因转录的调控

基本信息

批准号：
05680598
负责人：
WADA Yuko
金额：
$ 1.22万
依托单位：
Nippon Medical School
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1995
项目状态：
已结题

项目摘要

The human epsilon-globin gene contains not only silencer but also several important cis-acting factors in the upstream flanking region, which regulate stage-specific switching of globin genes. Previously we characterized the human epsilon-globin gene silencer by in vitro transcription assay. Here we first examined the S1 nuclease hypersensitive sites (S1 HS) in the upstream flanking region and found that two regions were S1 HS.The unusual feature in the region, triplex DNA or slipped DNA,was most likely to be the structure that causes S1 nuclease sensitivity. We also examined bent DNA which is one of the unusual structures intrinsically adopted by the sequence. The circular permutation experiment revealed that the DNA bend sites appeared every 680 bp on average in the entire region. In order to know the periodicity of the bend sites in other globin genes, we next mapped the sites in the beta-globin gene region. The relative positions of the sites to the cap site were identical to those in the epsilon-globin gene region, suggesting that the bend sites have been conserved during molecular evolution of the genomic sequence among two genes. The result also strengthens their significance in the genomic organization such as a signal for nucleosome phasing. Finally, we mapped the bend sites in the region spanning 13.3kb containing the Ggamma-Agamma-psibeta-globin genes. The bend sites were present at an interval of roughly 700bp and they divide the region into units. Furthermore, they were conserved in the promoter region of nearly all beta-family globin genes, although the periodicity of the sites was locally disturbed at the junctions of the duplicated Ggamma-and Agamma-globin genes and their second introns. This suggested that the periodicity of the bend sites is ranked lower in the hierarchy of genomic DNA organization than genome rearrangement and gene expression.

人类ε珠蛋白基因不仅含有沉默子，而且在上游侧翼区域还含有几个重要的顺式作用因子，这些因子调节珠蛋白基因的阶段特异性转换。之前我们通过体外转录测定对人类ε-珠蛋白基因沉默子进行了表征。在这里，我们首先检查了上游侧翼区域的S1核酸酶超敏位点（S1 HS），发现两个区域是S1 HS。该区域的不寻常特征，三链DNA或滑动DNA，很可能是导致S1核酸酶敏感性的结构。我们还检查了弯曲 DNA，这是该序列本质上采用的不寻常结构之一。循环排列实验表明，整个区域平均每680 bp出现一个DNA弯曲位点。为了了解其他珠蛋白基因中弯曲位点的周期性，我们接下来绘制了 β-珠蛋白基因区域中的位点。这些位点与帽位点的相对位置与ε-珠蛋白基因区域中的位置相同，表明弯曲位点在两个基因的基因组序列的分子进化过程中是保守的。该结果还增强了它们在基因组组织中的重要性，例如核小体定相的信号。最后，我们绘制了包含 Ggamma-Agamma-psibeta-globin 基因的 13.3kb 区域中的弯曲位点。弯曲位点以大约 700bp 的间隔存在，并将该区域划分为单元。此外，它们在几乎所有β家族珠蛋白基因的启动子区域中都是保守的，尽管这些位点的周期性在重复的Ggamma-和Agamma-珠蛋白基因及其第二内含子的连接处局部受到干扰。这表明弯曲位点的周期性在基因组 DNA 组织的层次结构中排列得比基因组重排和基因表达低。