Spatial organization of chromosome territories in the interphase nucleus of trisomy 21 cells

In the interphase cell nucleus, chromosomes adopt a conserved and non-random arrangement in subnuclear domains called chromosome territories (CTs). Whereas chromosome translocation can affect CT organization in tumor cell nuclei, little is known about how aneuploidies can impact CT organization. Here, we performed 3D-FISH on control and trisomic 21 nuclei to track the patterning of chromosome territories, focusing on the radial distribution of trisomic HSA21 as well as 11 disomic chromosomes. We have established an experimental design based on cultured chorionic villus cells which keep their original mesenchymal features including a characteristic ellipsoid nuclear morphology and a radial CT distribution that correlates with chromosome size. Our study suggests that in trisomy 21 nuclei, the extra HSA21 induces a shift of HSA1 and HSA3 CTs out toward a more peripheral position in nuclear space and a higher compaction of HSA1 and HSA17 CTs. We posit that the presence of a supernumerary chromosome 21 alters chromosome compaction and results in displacement of other chromosome territories from their usual nuclear position.     

Chromosome territory positioning of conserved homologous chromosomes in different primate species

Interphase chromosomes form distinct spatial domains called chromosome territories (CTs). The position of CTs is known not to be at random and is related to chromosome size and gene density. To elucidate how CTs are arranged in primate proliferating fibroblasts and whether the radial position of CTs has been conserved during primate evolution, several specific CTs corresponding to conserved chromosomes since the Simiiformes (human 6, 12, 13, and 17 homologous CTs) have been studied in 3D preserved interphase nuclei from proliferant cells of two New World monkey species (Lagothrix lagothricha, Saimiri sciureus) and in human by three-dimensional fluorescent in situ hybridization (3D-FISH). Our results indicate that both gene-density and chromosome size influence chromosome territory arrangement in the nucleus. This influence is greater for chromosome-size than for gene-density in the three species studied. A comparison of the radial position of a given CT and its homolog in the species analyzed suggests similar CT distributions for homologous chromosomes. Our statistical analysis using the logit model shows that such homologous positionings cannot, however, be considered identical.Electronic Supplementary Material Supplementary material is available for this article at This paper is dedicated to the memory of Prof. Josep Egozcue, our enthusiast teacher and a good friend.     

Non-random radial arrangements of interphase chromosome territories: evolutionary considerations and functional implications

In the nucleus of animal and plant cells individual chromosomes maintain a compartmentalized structure. Chromosome territories (CTs), as these structures were named by Theodor Boveri, are essential components of the higher-order chromatin architecture. Recent studies in mammals and non-mammalian vertebrates indicate that the radial position of a given CT (or segments thereof) is correlated with its size, its gene-density and its replication timing. As a representative case, chicken cell nuclei show highly consistent radial chromatin arrangements: gene-rich, early replicating microchromosomes are clustered within the nuclear interior, while gene-poor, later replicating macrochromosomes are preferentially located at the nuclear periphery. In humans, chromosomes 18 and 19 (HSA18 and 19) territories that are of similar size show a distinctly different position in the cell nuclei of lymphocytes and lymphoblastoid cells: the gene-rich and early replicating HSA19 CTs are typically found close to the nuclear center, while the gene-poor and later replicating HSA18 CTs are preferentially located at the nuclear periphery. Recent comparative maps between human and chicken chromosomes revealed that the chicken macrochromosomes 2 and Z contain the genes homologous to HSA18, while the genes on HSA19 are located onto the chicken microchromosomes. These data lend tentative support to the hypothesis that differences in the radial nuclear positions of gene-rich, early replicating and gene-poor, later replicating chromatin have been evolutionarily conserved during a period of more than 300 million years irrespective of the evolution of highly divergent karyotypes between humans and chicken.     

Inheritance of gene density-related higher order chromatin arrangements in normal and tumor cell nuclei

A gene density-related difference in the radial arrangement of chromosome territories (CTs) was previously described for human lymphocyte nuclei with gene-poor CT #18 located toward the nuclear periphery and gene-dense CT #19 in the nuclear interior (Croft, J.A., J.M. Bridger, S. Boyle, P. Perry, P. Teague, and W.A. Bickmore. 1999. J. Cell Biol. 145:1119-1131). Here, we analyzed the radial distribution of chromosome 18 and 19 chromatin in six normal cell types and in eight tumor cell lines, some of them with imbalances and rearrangements of the two chromosomes. Our findings demonstrate that a significant difference in the radial distribution of #18 and #19 chromatin is a common feature of higher order chromatin architecture in both normal and malignant cell types. However, in seven of eight tumor cell lines, the difference was less pronounced compared with normal cell nuclei due to a higher fraction of nuclei showing an inverted CT position, i.e., a CT #18 located more internally than a CT #19. This observation emphasizes a partial loss of radial chromatin order in tumor cell nuclei.     

Conservation of relative chromosome positioning in normal and cancer cells

Chromosomes exist in the interphase nucleus as individual chromosome territories. It is unclear to what extent chromosome territories occupy particular positions with respect to each other and how structural rearrangements, such as translocations, affect chromosome organization within the cell nucleus. Here we analyze the relative interphase positioning of chromosomes in mouse lymphoma cells compared to normal splenocytes. We show that in a lymphoma cell line derived from an ATM(-/-) mouse, two translocated chromosomes are preferentially positioned in close proximity to each other. The relative position of the chromosomes involved in these translocations is conserved in normal splenocytes. Relative positioning of chromosomes in normal splenocytes is not due to their random distribution in the interphase nucleus and persists during mitosis. These observations demonstrate that the relative arrangement of chromosomes in the interphase nucleus can be conserved between normal and cancer cells and our data support the notion that physical proximity facilitates rearrangements between chromosomes.     

Yeast importin-β is required for nuclear import of the Mig2 repressor

Background

In interphase nuclei of a wide range of species chromosomes are organised into their own specific locations termed territories. These chromosome territories are non-randomly positioned in nuclei which is believed to be related to a spatial aspect of regulatory control over gene expression. In this study we have adopted the pig as a model in which to study interphase chromosome positioning and follows on from other studies from our group of using pig cells and tissues to study interphase genome re-positioning during differentiation. The pig is an important model organism both economically and as a closely related species to study human disease models. This is why great efforts have been made to accomplish the full genome sequence in the last decade.

Results

This study has positioned most of the porcine chromosomes in in vitro cultured adult and embryonic fibroblasts, early passage stromal derived mesenchymal stem cells and lymphocytes. The study is further expanded to position four chromosomes in ex vivo tissue derived from pig kidney, lung and brain.

Conclusions

It was concluded that porcine chromosomes are also non-randomly positioned within interphase nuclei with few major differences in chromosome position in interphase nuclei between different cell and tissue types. There were also no differences between preferred nuclear location of chromosomes in in vitro cultured cells as compared to cells in tissue sections. Using a number of analyses to ascertain by what criteria porcine chromosomes were positioned in interphase nuclei; we found a correlation with DNA content.     

Lamin A/C and polymeric actin in genome organization

In this work, we have studied the structural and functional linkage between lamin A/C, nuclear actin, and organization of chromosome territories (CTs) in mammary carcinoma MCF-7 cells. Selective down-regulation of lamin A/C expression led to disruption of the lamin A/C perinuclear layer and disorganization of lamin-bound emerin complexes at the inner nuclear membrane. The silencing of lamin A/C expression resulted in a decrease in the volume and surface area of chromosome territories, especially in chromosomes with high heterochromatin content. Inhibition of actin polymerization led to relaxation of the structure of chromosome territories, and an increase in the volumes and surface areas of the chromosome territories of human chromosomes 1, 2 and 13. The results show an important role of polymeric actin in the organization of the nuclei and the chromosome territories.     

Chromosome Territories

Chromosome territories (CTs) constitute a major feature of nuclear architecture. In a brief statement, the possible contribution of nuclear architecture studies to the field of epigenomics is considered, followed by a historical account of the CT concept and the final compelling experimental evidence of a territorial organization of chromosomes in all eukaryotes studied to date. Present knowledge of nonrandom CT arrangements, of the internal CT architecture, and of structural interactions with other CTs is provided as well as the dynamics of CT arrangements during cell cycle and postmitotic terminal differentiation. The article concludes with a discussion of open questions and new experimental strategies to answer them.Impressive progress has been achieved during the last decade with regard to the functional implications of DNA methylation, histone modifications, and chromatin remodeling events for gene regulation (Fuks 2005; Kouzarides 2007; Maier et al. 2008; Jiang and Pugh 2009). It has, however, also become obvious that decoding the chromatin language does not suffice to fully understand the ways in which the diploid genome contributes to the formation of the different epigenomes present in the various cell types of a multicellular organism.Different epigenomes and their functional implications also depend on differences in higher‐order chromatin organization and nuclear architecture at large. Epigenomic research aims for an integrated understanding of the structural and functional aspects of epigenetics with nuclear architecture during the differentiation of toti- or pluripotent cells to functionally distinct cell types.The territorial organization of chromosomes in interphase (chromosome territories, CTs) constitutes a basic feature of nuclear architecture. This article starts with a brief historical account of the CT concept and the compelling experimental evidence in favor of a territorial organization of chromosomes in all eukaryotes studied to date. A survey of what is presently known about nonrandom arrangements of CTs, about changes of such arrangements in cycling cells as a result of internal or external influences and about the internal architecture of CTs and their structural interactions with each other is provided. The article concludes with a discussion of open questions on CT organization and new experimental strategies to answer them.     

Dynamic 1H NMR-based extracellular metabonomic analysis of oligodendroglia cells infected with herpes simplex virus type 1

Herpes simplex virus type 1 (HSV-1) is a large, neurotropic, double-stranded DNA virus that establishes a lifelong latent infection in neurons and glial cells. Previous studies reveal that several metabolic perturbations are associated with HSV-1 infection. However, the extracellular metabolic alterations associated with HSV-1 infection have not been systematically profiled in human cells. Here, a proton nuclear magnetic resonance-based metabonomic approach was applied to differentiate the extracellular metabonomic profiles of HSV-1 infected human oligodendroglia cells (n = 18) and matched control cells (n = 18) at three time points (12, 24, and 36 h post-infection). Resulting spectra were analyzed by chemometric and statistical methods. Metabonomic profiling revealed perturbations in 21 extracellular metabolites. Partial least squares discriminant analysis demonstrated that the whole metabolic patterns enabled statistical discrimination between HSV-1 infected human oligodendroglia cells and control cells. Eight extracellular metabolites, seven of which were amino acids, were primarily responsible for score plot discrimination between HSV-1 infected human oligodendroglia cells and control cells at 36 h post-infection: alanine, glycine, isoleucine, leucine, glutamate, glutamine, histidine, and lactate. HSV-1 infection alters amino acid metabolism in human oligodendroglia cells cultured in vitro. HSV-1 infection may disturb these host cellular pathways to support viral replication. Through elucidating the extracellular metabolic changes incident to HSV-1 infection, this study also provides future directions for investigation into the pathogenic mechanism of HSV-1.     

Cell Type Specific Alterations in Interchromosomal Networks across the Cell Cycle

The interchromosomal organization of a subset of human chromosomes (#1, 4, 11, 12, 16, 17, and 18) was examined in G1 and S phase of human WI38 lung fibroblast and MCF10A breast epithelial cells. Radial positioning of the chromosome territories (CTs) was independent of gene density, but size dependent. While no changes in radial positioning during the cell cycle were detected, there were stage-specific differences between cell types. Each CT was in close proximity (interaction) with a similar number of other CT except the gene rich CT17 which had significantly more interactions. Furthermore, CT17 was a member of the highest pairwise CT combinations with multiple interactions. Major differences were detected in the pairwise interaction profiles of MCF10A versus WI38 including cell cycle alterations from G1 to S. These alterations in interaction profiles were subdivided into five types: overall increase, overall decrease, switching from 1 to ≥2 interactions, vice versa, or no change. A global data mining program termed the chromatic median determined the most probable overall association network for the entire subset of CT. This probabilistic interchromosomal network was nearly completely different between the two cell lines. It was also strikingly altered across the cell cycle in MCF10A, but only slightly in WI38. We conclude that CT undergo multiple and preferred interactions with other CT in the nucleus and form preferred -albeit probabilistic- interchromosomal networks. This network of interactions is altered across the cell cycle and between cell types. It is intriguing to consider the relationship of these alterations to the corresponding changes in the gene expression program across the cell cycle and in different cell types.     

Changes in chromosome positioning may contribute to the development of diseases related to X-chromosome aneuploidy

The radial positions of the centromeric regions of chromosomes 1 and X were determined in normal male fibroblasts (XY) and in fibroblasts from a patient with a rare case of XXXXY polysomy. The centromeric regions and presumably the whole territories of active X chromosomes were demonstrated to occupy similar, although not identical, positions in XY and XXXXY cells. The centromeres of inactive X chromosomes (Barr bodies) were located closer to the nuclear periphery as compared with the centromeres of active X chromosomes. In addition, it was established that the nuclear radial position of gene-rich chromosome 1 was changed in XXXXY cells as compared to normal XY cells. The data are discussed in the context of the hypothesis postulating that changes in nuclear positioning of chromosomal territories induced by the presence of extra copies of individual chromosomes may contribute to the development of diseases related to different polysomies.     

Inter- and intra-specific gene-density-correlated radial chromosome territory arrangements are conserved in Old World monkeys

Recently it has been shown that the gene-density correlated radial distribution of human 18 and 19 homologous chromosome territories (CTs) is conserved in higher primates in spite of chromosomal rearrangements that occurred during evolution. However, these observations were limited to apes and New World monkey species. In order to provide further evidence for the evolutionary conservation of gene-density-correlated CT arrangements, we extended our previous study to Old World monkeys. They comprise the remaining species group to be analyzed in order to obtain a comprehensive overview of the nuclear topology of human 18 and 19 homologous CTs in higher primates. In the present study we investigated four lymphoblastoid cell lines from three species of Old World monkeys by three-dimensional fluorescence in situ hybridization (3D-FISH): two individuals of Japanese macaque (Macaca fuscata), crab-eating macaque (Macaca fascicularis), and an interspecies hybrid individual between African green monkey (Cercopithecus aethiops) and Patas monkey (Erythrocebus patas). Our data demonstrate that gene-poor human 18 homologous CTs are located preferentially close to the nuclear periphery, whereas gene-dense human 19 homologous CTs are oriented towards the nuclear center in all cell lines analyzed. The gene-density-correlated positioning of human 18 and 19 homologous CTs is evolutionarily conserved throughout all major higher primate lineages, despite chromosomal inversions, fusions, fissions or reciprocal translocations that occurred in the course of evolution in these species. This remarkable preservation of a gene-density-correlated chromatin arrangement gives further support for a functionally relevant higher-order chromatin architecture.     

染色体领域的研究及其进展

间期核中的染色体并不是散乱分布的,而是每条染色体占据了一块特定的核区域,即染色体领域（chromosome territory, CTs）,染色体领域在间期核中的排列与定位是经过严格组织的,并具有一定的动力学特征,染色体领域的这些严格的定位和空间组织与基因的表达调控密切相关。文章综述了这几个方面的研究进展。