Nuclear and territorial topography of chromosome telomeres in human lymphocytes

Amrichová, Jana; Lukášová, Emı́lie; Kozubek, Stanislav; Kozubek, Michal

doi:10.1016/s0014-4827(03)00208-8

Cited by 47 publications

(49 citation statements)

References 67 publications

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“…3B, mock-treated). This observation confirms previous studies done on fixed cells, showing a more central localization of centromeres as compared to telomeres in interphase nuclei in various cell types (29,30). The CDF plot revealed that few telomeres are localized at the nuclear periphery in untreated hMSCs (Fig.…”

Section: Changes In Spatial Distribution During Activation Of Apoptosissupporting

confidence: 93%

Segmentation and analysis of the three‐dimensional redistribution of nuclear components in human mesenchymal stem cells

et al. 2008

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To better understand the impact of changes in nuclear architecture on nuclear functions, it is essential to quantitatively elucidate the three-dimensional organization of nuclear components using image processing tools. We have developed a novel image segmentation method, which involves a contrast enhancement and a subsequent thresholding step. In addition, we have developed a new segmentation method of the nuclear volume using the fluorescent background signal of a probe. After segmentation of the nucleus, a first-order normalization is performed on the signal positions of the component of interest to correct for the shape of the nucleus. This method allowed us to compare various signal positions within a single nucleus, and also on pooled data obtained from multiple nuclei, which may vary in size and shape. The algorithms have been tested by analyzing the spatial localization of nuclear bodies in relation to the nuclear center. Next, we used this new tool to study the change in the spatial distribution of nuclear components in cells before and after caspase-8 activation, which leads to cell death. Compared to the morphological TopHat method, this method gives similar but significantly faster results. A clear shift in the radial distribution of centromeres has been found, while the radial distribution of telomeres was changed much less. In addition, we have used this new tool to follow changes in the spatial distribution of two nuclear components in the same nucleus during activation of apoptosis. We show that after caspase-8 activation, when centromeres shift to a peripheral localization, the spatial distribution of PML-NBs does not change while that of centromeres did. We propose that the use of this new image segmentation method will contribute to a better understanding of the 3D spatial organization of the cell nucleus. ' 2008 International Society for Advancement of CytometryKey terms confocal microscopy; image processing; caspase-8; apoptosis; telomeres; centromeres THE major function of the cell nucleus is to regulate gene activity, which depends on well-studied molecular mechanisms such as transcription, pre-mRNA splicing, and ribosome assembly. In contrast to what is known about the molecular regulation of these mechanisms, far less is understood about the extent that the dynamics of nuclear components and the three-dimensional (3D) structural organization of the nucleus contribute to the regulation of nuclear functions. Recent models of high-order genome organization suggest a nonrandom spatial localization of chromosome territories in the interphase nucleus (1). Also subchromosomal domains are suggested to be nonrandomly positioned. Centromeres containing pericentric satellite repeats show a preferential peripheral orientation in G0-arrested human cells (2) as well as in differentiated cells (2-4). Telomeres, which are satellite repeats at the ends of chromosomes, reveal a cell cycle-dependent localization in B-lymphocytes. Throughout the cell cycle, telomeres exhibit a spherical organizatio...

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Section: Changes In Spatial Distribution During Activation Of Apoptosissupporting

confidence: 93%

Segmentation and analysis of the three‐dimensional redistribution of nuclear components in human mesenchymal stem cells

et al. 2008

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“…An increasing body of evidence shows that telomeres are organized in a nonrandom, cell-type, and cell cycle-dependent manner (16,20,21,44,45). Although yeast cells are known to form few large clusters at the heterochromatic nuclear border (9,10), most human telomeres have been shown to avoid the nuclear periphery (16,21).…”

Section: Discussionmentioning

confidence: 99%

Controlled light exposure microscopy reveals dynamic telomere microterritories throughout the cell cycle

et al. 2008

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Telomeres are complex end structures that confer functional integrity and positional stability to human chromosomes. Despite their critical importance, there is no clear view on telomere organization in cycling human cells and their dynamic behavior throughout the cell cycle. We investigated spatiotemporal organization of telomeres in living human ECV-304 cells stably expressing telomere binding proteins TRF1 and TRF2 fused to mCitrine using four dimensional microscopy. We thereby made use of controlled light exposure microscopy (CLEM), a novel technology that strongly reduces photodamage by limiting excitation in parts of the image where full exposure is not needed. We found that telomeres share small territories where they dynamically associate. These territories are preferentially positioned at the interface of chromatin domains. TRF1 and TRF2 are abundantly present in these territories but not firmly bound. At the onset of mitosis, the bulk of TRF protein dissociates from telomere regions, territories disintegrate and individual telomeres become faintly visible. The combination of stable cell lines, CLEM and cytometry proved essential in providing novel insights in compartment-based nuclear organization and may serve as a model approach for investigating telomere-driven genome-instability and studying long-term nuclear dynamics. ' 2008 International Society for Advancement of Cytometry Key termstelomere; TRF1; TRF2; nuclear organization; chromatin dynamics; CLEM; live cell imaging TELOMERES are the natural ends of linear chromosomes. A mammalian telomere consists of a double stranded array of simple TTAGGG repeats ending in a single stranded overhang that folds back to form a T-loop structure (1). In combination with sufficient telomere repeats, a complex of indirect and direct telomere binding proteins, dubbed shelterin, assures proper telomere structure and function. The specificity of shelterin for telomeric DNA is due to the recognition of TTAGGG repeats by three of its components: the homodimers telomeric repeat binding factor 1 and 2 (TRF1 and TRF2) that bind the duplex part of telomeres, and protection of telomeres 1 that binds to the single stranded TTAGGG repeats present at the three-overhang and in the D loop of the T-loop configuration (2-5).The nucleus has a meticulous organization with functional relevance to the cell assuring proper gene expression, replication and genome stability (6-8). Telomeres are considered to play an important role in spatial genome organization. Whereas yeast telomeres are known to form few large silencing clusters at the heterochromatic nuclear periphery (9,10), there is no consensus on how telomeres are organized in the mammalian nucleus. From an architectural point of view, human telomeres have been proposed to be anchored to a nuclear scaffold, thereby possibly providing struc-

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“…Studies in various organisms suggest that the nuclear periphery is a zone for transcription repression, although active genes are also found in association with nuclear pores (reviewed by Sexton et al, 2007). Heterochromatic regions, such as telomeres and centromeres are enriched at the nuclear periphery in a cell-cycle-dependent manner (Weierich et al, 2003;Amrichova et al, 2003;Taddei et al, 2004), and recent models suggest that these regions are directly involved in gene repression (Taddei et al, 2004). …”

Section: Introductionmentioning

confidence: 99%

The nuclear lamina promotes telomere aggregation and centromere peripheral localization during senescence of human mesenchymal stem cells

Raz

Vermolen

Garini

et al. 2008

Journal of Cell Science

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Ex vivo, human mesenchymal stem cells (hMSCs) undergo spontaneous cellular senescence after a limited number of cell divisions. Intranuclear structures of the nuclear lamina were formed in senescent hMSCs, which are identified by the presence of Hayflick-senescence-associated factors. Notably, spatial changes in lamina shape were observed before the Hayflick senescence-associated factors, suggesting that the lamina morphology can be used as an early marker to identify senescent cells. Here, we applied quantitative image-processing tools to study the changes in nuclear architecture during cell senescence. We found that centromeres and telomeres colocalised with lamina intranuclear structures, which resulted in a preferred peripheral distribution in senescent cells. In addition, telomere aggregates were progressively formed during cell senescence. Once formed, telomere aggregates showed colocalization with γ-H2AX but not with TERT, suggesting that telomere aggregates are sites of DNA damage. We also show that telomere aggregation is associated with lamina intranuclear structures, and increased telomere binding to lamina proteins is found in cells expressing lamina mutants that lead to increases in lamina intranuclear structures. Moreover, three-dimensional image processing revealed spatial overlap between telomere aggregates and lamina intranuclear structures. Altogether, our data suggest a mechanical link between changes in lamina spatial organization and the formation of telomere aggregates during senescence of hMSCs, which can possibly contribute to changes in nuclear activity during cell senescence.

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Nuclear and territorial topography of chromosome telomeres in human lymphocytes

Cited by 47 publications

References 67 publications

Segmentation and analysis of the three‐dimensional redistribution of nuclear components in human mesenchymal stem cells

Segmentation and analysis of the three‐dimensional redistribution of nuclear components in human mesenchymal stem cells

Controlled light exposure microscopy reveals dynamic telomere microterritories throughout the cell cycle

The nuclear lamina promotes telomere aggregation and centromere peripheral localization during senescence of human mesenchymal stem cells

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