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Studies on Intranuclear Arrangement of HumanChromosome 12 and Relative Locations ofChromosome 15 Centromeres and of SNRPNGenes in Interphase Nuclei of HL 60 Cells
https://ir.soken.ac.jp/records/941
https://ir.soken.ac.jp/records/941e216dd2b-b07b-49f7-9f60-4bfe38e19b13
| 名前 / ファイル | ライセンス | アクション |
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要旨・審査要旨 / Abstract, Screening Result (460.4 kB)
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本文 (7.0 MB)
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| Item type | 学位論文 / Thesis or Dissertation(1) | |||||
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| 公開日 | 2010-02-22 | |||||
| タイトル | ||||||
| タイトル | Studies on Intranuclear Arrangement of HumanChromosome 12 and Relative Locations ofChromosome 15 Centromeres and of SNRPNGenes in Interphase Nuclei of HL 60 Cells | |||||
| タイトル | ||||||
| タイトル | Studies on Intranuclear Arrangement of HumanChromosome 12 and Relative Locations ofChromosome 15 Centromeres and of SNRPNGenes in Interphase Nuclei of HL 60 Cells | |||||
| 言語 | en | |||||
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| 言語 | eng | |||||
| 資源タイプ | ||||||
| 資源タイプ識別子 | http://purl.org/coar/resource_type/c_46ec | |||||
| 資源タイプ | thesis | |||||
| 著者名 |
野上, 正弘
× 野上, 正弘 |
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| フリガナ |
ノガミ, マサヒロ
× ノガミ, マサヒロ |
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| 著者 |
NOGAMI, Masahiro
× NOGAMI, Masahiro |
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| 学位授与機関 | ||||||
| 学位授与機関名 | 総合研究大学院大学 | |||||
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| 学位名 | 博士（理学） | |||||
| 学位記番号 | ||||||
| 内容記述タイプ | Other | |||||
| 内容記述 | 総研大甲第406号 | |||||
| 研究科 | ||||||
| 値 | 生命科学研究科 | |||||
| 専攻 | ||||||
| 値 | 18 遺伝学専攻 | |||||
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| 学位授与年月日 | 1999-03-24 | |||||
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| 値 | 1998 | |||||
| 要旨 | ||||||
| 内容記述タイプ | Other | |||||
| 内容記述 | Two sets of approximately 3x109bp of human DNA are compacted within a small volume of the cell nucleus. A number of genes on the genome express timely and play each function under precise regulation. The whole genome duplicates in numerous numbers of human cells under the perfect order. To accomplish such sophisticated biological processes, the mammalian genome is organized by various levels of domain structures that essentially associate with biological functions. An understanding of chromosome organization and its relation to intranuclear arrangement is important for models of nuclear structure and function. Extensive efforts to obtain information about the primary structure of complex genomes. However, much less is understood about the higher order organization of the genome within the nucleus, in particular the chromosome organization and its relation to intranuclear arrangement.<br /> Application of the fluorescence in situ hybridization (FISH) technique using specific DNA probes lead to direct visualization of the genome organization in the nucleus as well as contribute to diagnostic analysis. Various types of probes for FISH are going to be available with the progress of the genome projects. Chromosome specific painting probes demonstrated territorial organization of chromosomes in the nucleus. Interphase chromosomes in mammalian cells are generally considered to be less condensed than their mitotic counterparts, but individual chromosomes appear to occupy restricted subcompartments in the interphase nucleus, i.e., chromosome territories. Many observations have been reported for temporally and spatially ordered organization of the genome within the nucleus. However, little is known about the relationship among specific gene locations, chromosome territories, and various intracellular processes.<br /> For understanding of the arrangement of specific genome sequences in the interphase nucleus of mammalian cells, I adopted two approaches. Firstly, I analyzed the relative positioning of specific DNA segments in the nucleus by multicolor FISH. In this approach, thousands of nuclei can be surveyed in a short time, and an intranuclear position of one specific DNA segment is inferred from comparison with those of other DNA segments from known genomic sites. The detailed information about each clone will be available, such as chromosomal localization, DNA replication timing, GC content, and gene or repetitive sequences. As an example, intranuclear arrangement of human chromosome 12 in G0(G1) nuclei of human myeloid leukemia HL60 cells was analyzed by using band-specific cosmid clones as probes. Each set of two cosmids was detected in different colors to the HL60 nuclei fixed by paraformaldehyde, and their relative positioning, internal or periphery, in the individual nucleus was scored. The results suggest that the intranuclear arrangement of human chromosome 12 is not random. Some chromosomal domains, including centromeric region, localized in the nuclear periphery, while other parts, including telomeric regions, positioned in the internal parts of the nucleus in G0(G1) cells. Based on the replication banding pattern of metaphase spreads, human chromosome 12 was divided roughly into five large domains. Interestingly, the clones in later replicating domains preferentially localized in the nuclear periphery, whereas those in earlier replicating domains were arranged in the internal position of the nuclei. DNA replication timing of each cosmid determined by FISH-based assay was consistent with the replication R-banding profile of chromosome 12. These results suggest that a topological arrangement of a human chromosome correlates to large-scale replication domains of the genome, and possibly to the chromosome bands, even before the DNA replication stage of cell cycle.<br /> Next, I adopted an approach to simultaneously visualize four targets including a specific gene, a centromere, a whole chromosome, and a nucleus, and to three-dimensionally analyze their relative positioning in the nucleus during cell cycle by using a deconvolution system. I selected an imprinted gene, SNRPN (small nuclear ribonucleoprotein polypeptide N), as an example, because this gene are well-analyzed on its imprinted expression, replication timing, etc., and also homologous association of this gene region at late S phase has been reported. The relative positions of chromosome 15 centromeres and SNRPN genes in the interphase nuclei of HL60 cells and their cell cycle dependency were analyzed with respect to the territories occupied by the whole chromosome 15 in which these DNAs are localized. Chromosome 15 telritory, its centromere, SNRPN gene, and the nucleus were simultaneously visualized in three-dimensionally preserved nuclei by multicolor FISH. Spatial distribution of DNAs analyzed by a cooled CCD camera deconvolution system revealed that the SNRPN gene relatively localizes on the periphery of the chromosome territories and that preferentially faces to the nuclear membrane in late S and G2 phases. The chromosome 15 centromere and SNRPN gene come close to each other in late S phase, suggesting that both DNA segments replicate together within the same intranuclear domains, since they replicate in later half of S phase and DNA replication occurs in large foci at this stage of cell cycle. Preferential association of SNRPN does not occur in HL60 cells through the cell cycle. This contrasts with a report of homologous association of this imprinted chromosomal domain found in lymphocytes and lymphoblasts with the imprinting expression. RNA-FISH using an intron probe within SNRPN gene and the methylation status of this imprinted domain demonstrate that SNRPN gene is also imprinted in HL60 cells and allelically expressed through the cell cycle. This gene region in HL60 cells exhibits asynchronous replication which is a general feature of imprinted genome domains, similar with lymphocytes. These results suggest that factors other than imprinted expression and replication timing may be important determinants for the homologous association of imprinted genes. | |||||
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| 値 | 有 | |||||
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| 内容記述タイプ | Other | |||||
| 内容記述 | application/pdf | |||||
