Functional analysis of trans-acting factor binding to imprinting control region.

与印记控制区结合的反式作用因子的功能分析。

• 批准号: 15590290
• 负责人: OHTA Tohru
• 金额: $ 2.18万
• 依托单位: Health Sciences University of Hokkaido (2004)Nagasaki University (2003)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590290/
• 关键词: genomic imprinting; imprinting control center; insulator; CTCF; ゲノムインプリティング; インプリティング調節中枢; ゲノム刷り込み; Snrpn isoform; 刷り込み調節中枢

The mechanism of imprinting in chromosome 15q11-q13, AS/PWS domain has been analyzed continuously from several years ago. We have previously identified AS and PWS imprinting control center (IC) located at SNURF-SNRPN upstream region. The bipartite IC is supposed to have a function to establish maternal epigenotype on maternal chromosome in female germ line and early embryogenesis with AS-IC, and maintenance of the paternal epigenotype on paternal chromosome in somatic cells with PWS-IC. This function was predicted from inherited pattern of several familial AS or PWS patients with imprinting defect, AS/PWS model mouse analysis, and transgenic mouse analysis. Th elucidate the IC function, we analyzed the PWS-IC region with identification of some trans-acting factor binding sites. Since the human PWS-IC and mouse Snurf-Snrpn promoter are ortholog region based on several evidence of the knock out mouse, phylogenetic analysis was performed, and several conserved sequence were found in the PWS-IC region.In H19/Igf2 region, CTCF binding sites were found in the H19 promoter region. These sites are working as an insulator to prevent enhancer activity to the Igf2promoter.In the phylogenetic analysis, there are conserved sequences that have similar with the CTCF binding site, and some transcription binding sites in the PWS-IC as well as the H 19 promoter. To confirm the function of the transcription binding sites, transient transfection assay was performed with several kind of DNA construct including mutation at the binding site. In the assay, CTCF like binding site didn't show any insulator activity, and other transcription factor binding site show a positive activity for the expression. Therefore, the CTCF like binding site may not have a function of the insulator, and other transcription-binding site could have some function for imprinting maintenance. This research is currently ongoing. Furthermore, several imprinting genes were also analyzed.

染色体15q11-q13 AS/PWS结构域的印记机制从几年前就被不断地分析。我们之前已经确定 AS 和 PWS 印记控制中心（IC）位于 SNURF-SNRPN 上游区域。二分IC被认为具有在雌性种系中建立母本染色体上的母本表观基因型的功能，以及在AS-IC中建立早期胚胎发生的功能，以及在体细胞中维持父本染色体上的父本表观基因型的功能（在PWS-IC中）。该功能是根据几个具有印迹缺陷的家族性 AS 或 PWS 患者的遗传模式、AS/PWS 模型小鼠分析和转基因小鼠分析预测的。为了阐明 IC 功能，我们分析了 PWS-IC 区域并鉴定了一些反式作用因子结合位点。由于人PWS-IC和小鼠Snurf-Snrpn启动子是基于敲除小鼠的多个证据的直系同源区域，因此进行系统发育分析，在PWS-IC区域中发现了几个保守序列。在H19/Igf2区域中，CTCF在 H19 启动子区域发现了结合位点。这些位点作为绝缘体，阻止 Igf2 启动子的增强子活性。在系统发育分析中，存在与 CTCF 结合位点相似的保守序列，以及 PWS-IC 和 H 19 启动子中的一些转录结合位点。为了确认转录结合位点的功能，使用包括结合位点突变在内的几种DNA构建体进行瞬时转染测定。在测定中，CTCF样结合位点没有表现出任何绝缘子活性，而其他转录因子结合位点则表现出阳性表达活性。因此，CTCF样结合位点可能不具有绝缘体的功能，而其他转录结合位点可能具有一些印记维持功能。目前这项研究正在进行中。此外，还分析了几个印记基因。

项目成果

Yamada T. et al.: "Imprinting analysis of 10 genes and/or transcripts in a 1.5-Mb MEST-flanking region at human chromosome 7q32"Genomics. 83(3). 402-412 (2004)

Yamada T. 等人：“人类染色体 7q32 1.5-Mb MEST 侧翼区域中 10 个基因和/或转录物的印记分析”基因组学。

Kayashima T. et al.: "Atp10a, the mouse ortholog of the human imprinted ATP10A gene, escapes genomic imprinting"Genomics. 81(6). 644-647 (2003)

Kayashima T. 等人：“Atp10a，人类印记 ATP10A 基因的小鼠直系同源物，逃脱了基因组印记”Genomics。

Yamasaki K. et al.: "Neurons but not glial cells show reciprocal imprinting of sense and an tisense transcripts of Ube3a"Human Molecular Genetics. 12(8). 837-847 (2003)

Yamasaki K.等人：“神经元而非神经胶质细胞显示出Ube3a有义和反义转录本的相互印记”人类分子遗传学。

Imprinting analysis of 10 genes and/or transcripts in a 1.5-Mb MEST-flanking region at human chromosome 7q32.

• DOI: 10.1016/j.ygeno.2003.08.016
• 发表时间: 2004-03
• 期刊: Genomics
• 影响因子: 4.4
• 作者: Takahiro Yamada;K. Mitsuya;Tomohiko Kayashima;K. Yamasaki;T. Ohta;K. Yoshiura;N. Matsumoto;Hideto Yamada;H. Minakami;M. Oshimura;N. Niikawa;T. Kishino

Imprinting analysis of 10 genes and/or transcripts in a 1.5-Mb MEST-flanking region region at human chromosome 7q32.

对人类染色体 7q32 的 1.5 Mb MEST 侧翼区域中的 10 个基因和/或转录本进行印记分析。

• 发表时间: 2004
• 期刊: Genomics 83(3)
• 作者: Yamada T;Yamada T
• 通讯作者: Yamada T