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Journal ArticleDOI

Distinctive nuclear organisation of centromeres and regions involved in pluripotency in human embryonic stem cells

01 Sep 2005-Journal of Cell Science (The Company of Biologists Ltd)-Vol. 118, Iss: 17, pp 3861-3868
TL;DR: It is concluded that hES cell nuclei have a distinct nuclear architecture, especially at loci involved in maintaining pluripotency, which provides a framework within which other large-scale chromatin changes that may accompany differentiation can be considered.
Abstract: Nuclear organisation is thought to be important in regulating gene expression. Here we investigate whether human embryonic stem cells (hES) have a particular nuclear organisation, which could be important for maintaining their pluripotent state. We found that whereas the nuclei of hES cells have a general gene-density-related radial organisation of chromosomes, as is seen in differentiated cells, there are also distinctive localisations for chromosome regions and gene loci with a role in pluripotency. Chromosome 12p, a region of the human genome that contains clustered pluripotency genes including NANOG, has a more central nuclear localisation in ES cells than in differentiated cells. On chromosome 6p we find no overall change in nuclear chromosome position, but instead we detect a relocalisation of the OCT4 locus, to a position outside its chromosome territory. There is also a smaller proportion of centromeres located close to the nuclear periphery in hES cells compared to differentiated cells. We conclude that hES cell nuclei have a distinct nuclear architecture, especially at loci involved in maintaining pluripotency. Understanding this level of hES cell biology provides a framework within which other large-scale chromatin changes that may accompany differentiation can be considered.

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Citations
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Journal ArticleDOI
12 Jun 2008-Nature
TL;DR: A high-resolution map of the interaction sites of the entire genome with NL components in human fibroblasts is constructed and demonstrates that the human genome is divided into large, discrete domains that are units of chromosome organization within the nucleus.
Abstract: The architecture of human chromosomes in interphase nuclei is still largely unknown. Microscopy studies have indicated that specific regions of chromosomes are located in close proximity to the nuclear lamina (NL). This has led to the idea that certain genomic elements may be attached to the NL, which may contribute to the spatial organization of chromosomes inside the nucleus. However, sequences in the human genome that interact with the NL in vivo have not been identified. Here we construct a high-resolution map of the interaction sites of the entire genome with NL components in human fibroblasts. This map shows that genome-lamina interactions occur through more than 1,300 sharply defined large domains 0.1-10 megabases in size. These lamina-associated domains (LADs) are typified by low gene-expression levels, indicating that LADs represent a repressive chromatin environment. The borders of LADs are demarcated by the insulator protein CTCF, by promoters that are oriented away from LADs, or by CpG islands, suggesting possible mechanisms of LAD confinement. Taken together, these results demonstrate that the human genome is divided into large, discrete domains that are units of chromosome organization within the nucleus.

1,762 citations

Journal ArticleDOI
TL;DR: It is suggested that hyperdynamic binding of structural chromatin proteins is a functionally important hallmark of pluripotent ES cells that contributes to the maintenance of plasticity in undifferentiated ES cells and to establishing higher-order chromatin structure.

1,003 citations


Cites background from "Distinctive nuclear organisation of..."

  • ...In another system, no significant differences in the extent of centromere clustering were observed between undifferentiated human ES cells and two diploid differentiated cell types, including a lymphoblastoid cell line (FATO LCL) and primary fibroblasts (Wiblin et al., 2005)....

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  • ...However, centromeres in ES cells were mainly found within the nuclear interior, whereas, in differentiated cells, centromeres tend to localize at the nuclear periphery (Wiblin et al., 2005)....

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Journal ArticleDOI
TL;DR: This work discusses how unique properties of chromatin in ES cells contribute to the maintenance of pluripotency and the determination of differentiation properties.
Abstract: What makes a stem cell is still poorly understood. Recent studies have uncovered that chromatin might hold some of the keys to how embryonic stem cells maintain their pluripotency, their ability to self-renew and induce lineage specification. Embryonic stem (ES) cells are unique in that they are pluripotent and have the ability to self-renew. The molecular mechanisms that underlie these two fundamental properties are largely unknown. We discuss how unique properties of chromatin in ES cells contribute to the maintenance of pluripotency and the determination of differentiation properties.

707 citations


Cites background from "Distinctive nuclear organisation of..."

  • ...For example, comparable distributions of centromeres and of promyelocytic leukaemia (PML) bodies (which are implicated in transcription, apoptosis and cellular stress processes) are found in both human ES cells and differentiated cell...

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Journal ArticleDOI
TL;DR: It is found that differentiated tissues show surprisingly large K9-modified regions (up to 4.9 Mb), which are large organized chromatin K9 modifications (LOCKs) and may provide a cell type–heritable mechanism for phenotypic plasticity in development and disease.
Abstract: Higher eukaryotes must adapt a totipotent genome to specialized cell types with stable but limited functions. One potential mechanism for lineage restriction is changes in chromatin, and differentiation-related chromatin changes have been observed for individual genes. We have taken a genome-wide view of histone H3 lysine 9 dimethylation (H3K9Me2) and find that differentiated tissues show surprisingly large K9-modified regions (up to 4.9 Mb). These regions are highly conserved between human and mouse and are differentiation specific, covering only approximately 4% of the genome in undifferentiated mouse embryonic stem (ES) cells, compared to 31% in differentiated ES cells, approximately 46% in liver and approximately 10% in brain. These modifications require histone methyltransferase G9a and are inversely related to expression of genes within the regions. We term these regions large organized chromatin K9 modifications (LOCKs). LOCKs are substantially lost in cancer cell lines, and they may provide a cell type-heritable mechanism for phenotypic plasticity in development and disease.

582 citations

Journal ArticleDOI
TL;DR: It is demonstrated that oxygen concentrations affected many aspects of stem-cell physiology, including growth and in vitro development, and may be a critical parameter during expansion and differentiation.

509 citations

References
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Journal ArticleDOI
TL;DR: The data demonstrate that transcription of both ubiquitous and tissue-restricted genes is not confined to the periphery of chromosome territories, suggesting that the basal transcription machinery and transcription factors can readily gain access to the chromosome interior.
Abstract: The position of genes within the nucleus has been correlated with their transcriptional activity. The interchromosome domain model of nuclear organization suggests that genes preferentially locate at the surface of chromosome territories. Conversely, high resolution analysis of chromatin fibers suggests that chromosome territories do not present accessibility barriers to transcription machinery. To clarify the relationship between the organization of chromosome territories and gene expression, we have used fluorescence in situ hybridization to analyze the spatial organization of a contiguous approximately 1 Mb stretch of the Wilms' tumor, aniridia, genitourinary anomalies, mental retardation syndrome region of the human genome and the syntenic region in the mouse. These regions contain constitutively expressed genes, genes with tissue-restricted patterns of expression, and substantial regions of intergenic DNA. We find that there is a spatial organization within territories that is conserved between mouse and humans: certain sequences do preferentially locate at the periphery of the chromosome territories in both species. However, we do not detect genes necessarily at the periphery of chromosome territories or at the surface of subchromosomal domains. Intraterritory organization is not different among cell types that express different combinations of the genes under study. Our data demonstrate that transcription of both ubiquitous and tissue-restricted genes is not confined to the periphery of chromosome territories, suggesting that the basal transcription machinery and transcription factors can readily gain access to the chromosome interior.

245 citations


"Distinctive nuclear organisation of..." refers background or methods in this paper

  • ...In differentiated cells, gene-rich domains and regions of coordinately regulated gene expression, loop out from CTs (Volpi et al., 2000; Mahy et al., 2002a)....

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  • ...5 (Mahy et al., 2002a)....

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  • ...Gene clusters, and individual chromosomal domains also have distinctive localisations within respect to their CTs (Volpi et al., 2000; Williams et al., 2002; Mahy et al., 2002a)....

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  • ...Analysis of probe position relative to the surface of CTs, and interphase separation (d) were as previously described (Mahy et al., 2002a; Mahy et al., 2002b; Chambeyron and Bickmore, 2004)....

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  • ...We measured the distances between hybridisation signals for BACs for the specific loci, and the visible edge of the hybridisation signal for the corresponding CT (Mahy et al., 2002)....

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Journal ArticleDOI
TL;DR: The distance between a locus and its nearest PML body correlates with the transcriptional activity and gene density around the locus, andGenes on the active X chromosome are more significantly associated with PML bodies than their silenced homologues on the inactive X chromosome.
Abstract: The promyelocytic leukemia (PML) protein is aggregated into nuclear bodies that are associated with diverse nuclear processes. Here, we report that the distance between a locus and its nearest PML body correlates with the transcriptional activity and gene density around the locus. Genes on the active X chromosome are more significantly associated with PML bodies than their silenced homologues on the inactive X chromosome. We also found that a histone-encoding gene cluster, which is transcribed only in S-phase, is more strongly associated with PML bodies in S-phase than in G0/G1 phase of the cell cycle. However, visualization of specific RNA transcripts for several genes showed that PML bodies were not themselves sites of transcription for these genes. Furthermore, knock-down of PML bodies by RNA interference did not preferentially change the expression of genes closely associated with PML bodies. We propose that PML bodies form in nuclear compartments of high transcriptional activity, but they do not directly regulate transcription of genes in these compartments.

229 citations


"Distinctive nuclear organisation of..." refers background in this paper

  • ...Their nuclear distribution has not been extensively studied, but many transcriptionally active genomic regions, including parts of the major histocompatibility complex (MHC) at 6p, are reported to be associated with them (Wang et al., 2004)....

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Journal ArticleDOI
TL;DR: This study of the epidermal differentiation complex at 1q21 provides a further example of a gene-dense complex capable of assuming extraterritorial positioning in relation to cell type/transcription status.

206 citations


"Distinctive nuclear organisation of..." refers background in this paper

  • ...Gene clusters, and individual chromosomal domains also have distinctive localisations within respect to their CTs (Volpi et al., 2000; Williams et al., 2002; Mahy et al., 2002a)....

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  • ...Within CTs themselves, the position of gene clusters is altered in different differentiated human cell types (Volpi et al., 2000; Williams et al., 2002)....

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Journal ArticleDOI
TL;DR: The location of centromeres and telomeres was studied in human and mouse lymphocyte nuclei employing 3D-FISH, confocal microscopy, and quantitative image analysis and found a peripheral location of both centromres and CTs for 1, 11, 12, 18, X.
Abstract: The location of centromeres and telomeres was studied in human and mouse lymphocyte nuclei (G0) employing 3D-FISH, confocal microscopy, and quantitative image analysis. In both human and murine lymphocytes, most centromeres were found in clusters at the nuclear periphery. The distribution of telomere clusters, however, differed: in mouse nuclei, most clusters were detected at the nuclear periphery, while, in human nuclei, most clusters were located in the nuclear interior. In human cell nuclei we further studied the nuclear location of individual centromeres and their respective chromosome territories (CTs) for chromosomes 1, 11, 12, 15, 17, 18, 20, and X. We found a peripheral location of both centromeres and CTs for 1, 11, 12, 18, X. A mostly interior nuclear location was observed for CTs 17 and 20 and the CTs of the NOR-bearing acrocentric 15 but the corresponding centromeres were still positioned in the nuclear periphery. Autosomal centromeres, as well as the centromere of the active X, were typically located at the periphery of the respective CTs. In contrast, in about half of the inactive X-CTs, the centromere was located in the territory interior. While the centromere of the active X often participated in the formation of centromere clusters, such a participation was never observed for the centromere of the inactive X.

190 citations


"Distinctive nuclear organisation of..." refers background in this paper

  • ...Centromeres are also generally found at the nuclear periphery, or around nucleoli (Carvalho et al., 2001; Weierich et al., 2003; Gilchrist et al., 2004), whereas telomeres are mainly found in the nuclear interior (Weierich et al....

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  • ...Centromeres are also generally found at the nuclear periphery, or around nucleoli (Carvalho et al., 2001; Weierich et al., 2003; Gilchrist et al., 2004), whereas telomeres are mainly found in the nuclear interior (Weierich et al., 2003)....

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  • ...Telomeres are dispersed throughout the nucleoplasm of differentiated cells (Weierich et al., 2003)....

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  • ...Human centromeres are localised close to either the nuclear periphery or the nucleolus (Carvalho et al., 2001; Weierich et al., 2003)....

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  • ..., 2004), whereas telomeres are mainly found in the nuclear interior (Weierich et al., 2003)....

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Journal ArticleDOI
TL;DR: The observations indicate that alteration of relative chromosome positioning might play a key role in the tumorigenesis of human liposarcomas, and demonstrate the potential impact of higher order chromatin organization on the epigenetic mechanisms that control gene expression and gene silencing during cell differentiation.
Abstract: Chromosomes are highly restricted to specific chromosome territories within the interphase nucleus The arrangement of chromosome territories is non-random, exhibiting a defined radial distribution as well as a preferential association with specific nuclear compartments, which indicates a functional role for chromosome-territory organization in the regulation of gene expression In this report, we focus on changes in adipocyte differentiation that are related to a specific chromosomal translocation associated with liposarcoma tumorigenesis, t(12;16) We have examined the relative and radial positioning of the chromosome territories of human chromosomes 12 and 16 during adipocyte differentiation, and detected a close association between the territories of chromosomes 12 and 16 in differentiated adipocytes, an association not observed in preadipocytes Although further studies are required to elucidate the underlying reasons for the adipocyte-specific translocation of chromosomes 12 and 16, our observations indicate that alteration of relative chromosome positioning might play a key role in the tumorigenesis of human liposarcomas In addition, these results demonstrate the potential impact of higher order chromatin organization on the epigenetic mechanisms that control gene expression and gene silencing during cell differentiation

122 citations


"Distinctive nuclear organisation of..." refers background in this paper

  • ...However, to date no significant change in radial position of a human chromosome within the nucleus had been documented during differentiation, although there may be changes in chromosome associations (Kuroda et al., 2004)....

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  • ...changes in chromosome associations (Kuroda et al., 2004)....

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  • ...However, to date no significant change in radial position of a human chromosome within the nucleus has been documented during differentiation, although there may be changes in chromosome associations (Kuroda et al., 2004)....

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