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.

The regulation of gene expression is mediated by interactions between chromatin and protein complexes. The importance of where and when these interactions take place in the nucleus is currently a subject of intense investigation. Increasing evidence indicates that gene activation or silencing is often associated with repositioning of the locus relative to nuclear compartments and other genomic loci. At the same time, however, structural constraints impose limits on chromatin mobility. Understanding how the dynamic nature of the positioning of genetic material in the nuclear space and the higher-order architecture of the nucleus are integrated is therefore essential to our overall understanding of gene regulation.

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We demonstrate that the nuclear topological arrangement of chromosome territories (CTs) has been conserved during primate evolution over a period of about 30 million years. Recent evidence shows that the positioning of chromatin in human lymphocyte nuclei is correlated with gene density. For example, human chromosome 19 territories, which contain mainly gene-dense and early replicating chromatin, are located toward the nuclear center, whereas chromosome 18 territories, which consist mainly of genepoor and later replicating chromatin, is located close to the nuclear border. In this study, we subjected seven different primate species to comparative analysis of the radial distribution pattern of human chromosome 18-and 19-homologous chromatin by three-dimensional fluorescence in situ hybridization. Our data demonstrate that gene-density-correlated radial chromatin arrangements were conserved during higher-primate genome evolution, irrespective of the major karyotypic rearrangements that occurred in different phylogenetic lineages. The evolutionarily conserved positioning of homologous chromosomes or chromosome segments in related species supports evidence for a functionally relevant higher-order chromatin arrangement that is correlated with gene-density.

hether chromosomes maintain their nuclear positions during interphase and from one cell cycle to the next has been controversially discussed. To address this question, we performed long-term live-cell studies using a HeLa cell line with GFP-tagged chromatin. Positional changes of the intensity gravity centers of fluorescently labeled chromosome territories (CTs) on the order of several m were observed in early G1, suggesting a role of CT mobility in establishing interphase nuclear architecture. Thereafter, the positions were highly constrained within a range of ‫ف‬ 1 m until the end of G2. To analyze possible changes of chromosome arrangements from one cell cycle to the next, nuclei were photobleached in G2 W maintaining a contiguous zone of unbleached chromatin at one nuclear pole. This zone was stably preserved until the onset of prophase, whereas the contiguity of unbleached chromosome segments was lost to a variable extent, when the metaphase plate was formed. Accordingly, chromatin patterns observed in daughter nuclei differed significantly from the mother cell nucleus. We conclude that CT arrangements were stably maintained from mid G1 to late G2/early prophase, whereas major changes of CT neighborhoods occurred from one cell cycle to the next. The variability of CT neighborhoods during clonal growth was further confirmed by chromosome painting experiments.

In spite of strong evidence that the nucleus is a highly organized organelle, a consensus on basic principles of the global nuclear architecture has not so far been achieved. The chromosome territory-interchromatin compartment (CT-IC) model postulates an IC which expands between chromatin domains both in the interior and the periphery of CT. Other models, however, dispute the existence of the IC and claim that numerous chromatin loops expand between and within CTs. The present study was undertaken to resolve these conflicting views. (1) We demonstrate that most chromatin exists in the form of higher-order chromatin domains with a compaction level at least 10 times above the level of extended 30 nm chromatin fibers. A similar compaction level was obtained in a detailed analysis of a particularly gene-dense chromosome region on HSA 11, which often expanded from its CT as a finger-like chromatin protrusion. (2) We further applied an approach which allows the experimental manipulation of both chromatin condensation and the width of IC channels in a fully reversible manner. These experiments, together with electron microscopic observations, demonstrate the existence of the IC as a dynamic, structurally distinct nuclear compartment, which is functionally linked with the chromatin compartment.

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.

BackgroundA Xist RNA decorated Barr body is the structural hallmark of the compacted inactive X territory in female mammals. Using super-resolution three-dimensional structured illumination microscopy (3D-SIM) and quantitative image analysis, we compared its ultrastructure with active chromosome territories (CTs) in human and mouse somatic cells, and explored the spatio-temporal process of Barr body formation at onset of inactivation in early differentiating mouse embryonic stem cells (ESCs).ResultsWe demonstrate that all CTs are composed of structurally linked chromatin domain clusters (CDCs). In active CTs the periphery of CDCs harbors low-density chromatin enriched with transcriptionally competent markers, called the perichromatin region (PR). The PR borders on a contiguous channel system, the interchromatin compartment (IC), which starts at nuclear pores and pervades CTs. We propose that the PR and macromolecular complexes in IC channels together form the transcriptionally permissive active nuclear compartment (ANC). The Barr body differs from active CTs by a partially collapsed ANC with CDCs coming significantly closer together, although a rudimentary IC channel system connected to nuclear pores is maintained. Distinct Xist RNA foci, closely adjacent to the nuclear matrix scaffold attachment factor-A (SAF-A) localize throughout Xi along the rudimentary ANC. In early differentiating ESCs initial Xist RNA spreading precedes Barr body formation, which occurs concurrent with the subsequent exclusion of RNA polymerase II (RNAP II). Induction of a transgenic autosomal Xist RNA in a male ESC triggers the formation of an ‘autosomal Barr body’ with less compacted chromatin and incomplete RNAP II exclusion.Conclusions3D-SIM provides experimental evidence for profound differences between the functional architecture of transcriptionally active CTs and the Barr body. Basic structural features of CT organization such as CDCs and IC channels are however still recognized, arguing against a uniform compaction of the Barr body at the nucleosome level. The localization of distinct Xist RNA foci at boundaries of the rudimentary ANC may be considered as snap-shots of a dynamic interaction with silenced genes. Enrichment of SAF-A within Xi territories and its close spatial association with Xist RNA suggests their cooperative function for structural organization of Xi.