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Inhibition of enhancer-promoter interaction involved in the developmental-stage specific gene regulation

抑制参与发育阶段特异性基因调控的增强子-启动子相互作用

基本信息

批准号：
13460149
负责人：
TANIMOTO Keiji
金额：
$ 7.17万
依托单位：
University of Tsukuba
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13460149/
关键词：
globin insulator YAC transgenic mouse

项目摘要

The human (3-globin locus contains five developmentally regulated (3-type globin genes. All five genes depend on the locus control region (LCR), located at the 5' end of the locus, for abundant globin gene transcription. The LCR is composed of five DNase I hypersensitive sites (HSs), at least a subset of which appear to cooperate to form a holocomplex in activating genes within the locus. We previously tested the requirement for proper LCR polarity by inverting it in human β-globin YAC transgenic mice (TgM) and observed reduced expression of all the β-type globin genes, regardless of developmental stage. This phenotype clearly demonstrated an orientation-dependent activity of the LCR, although the mechanistic basis for the observed activity was obscure. In this research project, we obtained genetic evidence demonstrating that human HSS includes enhancer-blocking (insulator) activity that was dependent on the presence of an intact CTCF-binding motif within LCR HSS. Curiously, this CTCF-dependent activity appeared to be both gene and developmental-stage specific. These observations demonstrated that the phenotype observed in the LCR-inverted locus was, in part, attributable to placing the HSS insulator between the LCR HS "enhancers (HS1-4)" and the promoter of the β-globin gene, but further that the HS5 sequence also beared a primitive erythroid silencing activity that is CTCF-independent.

人类β-珠蛋白基因座含有五个发育调节的β-型珠蛋白基因。所有五个基因都依赖于位于基因座5'端的基因座控制区（LCR），用于丰富的珠蛋白基因转录。LCR由5个DNA酶I超敏位点（HS）组成，其中至少一个子集似乎在基因座内的激活基因中合作形成全复合物。我们先前通过在人β-珠蛋白YAC转基因小鼠（TgM）中反转LCR极性来测试对适当LCR极性的要求，并观察到所有β-型珠蛋白基因的表达降低，与发育阶段无关。这种表型清楚地表明了LCR的取向依赖性活性，尽管所观察到的活性的机制基础尚不清楚。在这个研究项目中，我们获得的遗传证据表明，人类HSS包括增强子阻断（绝缘子）活性，这取决于LCR HSS内完整的CTCF结合基序的存在。奇怪的是，这种CTCF依赖的活性似乎是基因和发育阶段特异性的。这些观察结果表明，在LCR倒置基因座中观察到的表型部分归因于将HSS绝缘子置于LCR HS“增强子（HS 1 -4）”和β-珠蛋白基因的启动子之间，但进一步地，HS 5序列还具有CTCF非依赖性的原始红细胞沉默活性。