Analysis of molecular structure of nucleosomes consisting of repetitive DNA sequences and their function in chromatin

由重复 DNA 序列组成的核小体的分子结构及其在染色质中的功能分析

• 批准号: 10670114
• 负责人: MISHIMA Yukio
• 金额: $ 2.05万
• 依托单位: NIIGATA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10670114/
• 关键词: repetitive sequences; GGA repeats; chromosome; reconstituted chromatin; DNA-DNA complex; homopurinesequences; microsatellite; DNase I footprinting; DNaseIプットフリント法; トリフレックス; 原子間力顕微鏡; 再構成ヌクレオソーム; トリプレックス; DNAペアリング

We investigated the ability of mononucleosomes reconstituted on DNA fragments containing (GGA/TCC)n repeats for DNA-DNA complex formation and examined the functional aspect of GGA-repeats by isolating genomic DNA fragments containing GGA-repeats and by analyzing their chromosomal localization. The results obtained were as follows.1. We isolated 19 DNA fragments containing (GGA /TCC)n repeats from mouse genome by triplex affinity capture.2. 19 clones of them were mapped to different locations of 13 chromosomes.3. Almost half (9/19) of the isolated DNA fragments containing (GGA/TCC)n repeats were enriched to heterochromatin fraction and the remains were equally distributed to hetero- and eu-chromatin fractions. Neither of the clones examined was enriched to euchromatin fraction.4. The results from gel mobility shift assay and DNase I protection mapping indicated that genomic DNA fragments containing GGA-repeats formed DNA- DNA complexes in a sequence-dependent manner.5. The results from the analysis of DNA-DNA complex formation of mononucleosomes reconstituted on various fragments of plasmid DNAs which contained different repeat-number of GGA-repeats or different flanking sequences carrying at either or both sites of GGA-repeats indicated that their complexes formations though GGA-repeats were required for both GGA-repeat number and homologous flanking sequences at both sites of GGA-repeats.

我们研究了在含DNA-DNA复合物形成的DNA片段（GGA/TCC）N重复序列上重构的单核小体的能力，并通过分离含有GGA重新培养并分析其染色体定位的基因组DNA片段来检查GGA重复的功能方面。获得的结果如下。1。我们分离了19个DNA片段，其中包含（GGA /TCC）n重复通过三重相关性捕获。2。它们的19个克隆被映射到13个染色体的不同位置3。将几乎一半（9/19）的含有含有（GGA/TCC）N重复量的分离的DNA片段富集到异染色质分数中，并将残留物均等分布到异性和EU-染色质分数。所检查的克隆都没有富含斑塑素分数。4。凝胶迁移率分析和DNase I保护映射的结果表明，以序列依赖性方式形成了含有GGA重复的基因组DNA片段。5。分析在质粒DNA的各种片段上重现的单核小体的DNA-DNA复合物形成的结果，这些质粒DNA中包含不同的重复数量的GGA重复或不同的侧翼序列，或在GGA重新重复的位置或两个位点携带的侧翼序列，虽然GGA序列均需要GGA序列，但它们均需要GGA序列的序列，而GGA则是GGA数字的序列。 GGA重复。

项目成果

Takuya Watanabe: "Differential chromatin packaging of genomic imprinted regions between expressed and non-expressed alleles."Human Molecular Genetics. 9. 3029-3035 (2000)

Takuy​​a Watanabe：“表达和非表达等位基因之间基因组印记区域的差异染色质包装。”人类分子遗传学。

Y.Endo et al.: "Compact chromatin packaging of inactive X chromosome involves the actively transcribed Xist gene" Mammalian Genome. in press. (1999)

Y.Endo 等人：“非活性 X 染色体的紧凑染色质包装涉及活跃转录的 Xist 基因”哺乳动物基因组。

Y.Endo, T.Watanabe, K.Kuwabara, K.Tsunashima, Y.Mishima, M.Arakawa, N.Takagi and R.Kominami: "Difference in chromatin packing between active and inactive X chromosomes by fractionation and allele-specific detection."Biochem.Biophys.Res.Commun.. 244. 220-2

Y.Endo、T.Watanabe、K.Kuwabara、K.Tsunashima、Y.Mishima、M.Arakawa、N.Takagi 和 R.Kominami：“通过分级和等位基因特异性检测，活性和非活性 X 染色体之间染色质堆积的差异