Abstract
Studies of higher-order chromatin arrangements are an essential part of ongoing attempts to explore changes in epigenome structure and their functional implications during development and cell differentiation. However, the extent and cell-type-specificity of three-dimensional (3D) chromosome arrangements has remained controversial. In order to overcome technical limitations of previous studies, we have developed tools that allow the quantitative 3D positional mapping of all chromosomes simultaneously. We present unequivocal evidence for a probabilistic 3D order of prometaphase chromosomes, as well as of chromosome territories (CTs) in nuclei of quiescent (G0) and cycling (early S-phase) human diploid fibroblasts (46, XY). Radial distance measurements showed a probabilistic, highly nonrandom correlation with chromosome size: small chromosomes—independently of their gene density—were distributed significantly closer to the center of the nucleus or prometaphase rosette, while large chromosomes were located closer to the nuclear or rosette rim. This arrangement was independently confirmed in both human fibroblast and amniotic fluid cell nuclei. Notably, these cell types exhibit flat-ellipsoidal cell nuclei, in contrast to the spherical nuclei of lymphocytes and several other human cell types, for which we and others previously demonstrated gene-density-correlated radial 3D CT arrangements. Modeling of 3D CT arrangements suggests that cell-type-specific differences in radial CT arrangements are not solely due to geometrical constraints that result from nuclear shape differences. We also found gene-density-correlated arrangements of higher-order chromatin shared by all human cell types studied so far. Chromatin domains, which are gene-poor, form a layer beneath the nuclear envelope, while gene-dense chromatin is enriched in the nuclear interior. We discuss the possible functional implications of this finding.
Highlights
Somatic cells within an organism possess genomes that are, with only a few minor exceptions, identical
Early-passage human fibroblast cultures (46, XY) were grown to confluence and maintained at this stage for several significantly higher probability of being localized at the nuclear periphery, while modeled large chromosome territory (CT) adopt a more internal localization. (C) In agreement with normalized 3D center of the nucleus (CN)–CT distances, normalized 3D CR–prometaphase chromosome (PC) distances show a positive correlation with the DNA content or size of chromosomes: small PCs were preferentially located near the CR, large chromosomes at the rosette periphery
spherical ~1-Mbp chromatin domain (SCD) Model Nuclei, and Prometaphase Rosettes (A) Normalized 3D radial CN–CT distances show a positive correlation with chromosome size: CTs of small chromosomes were preferentially located in the center of the nucleus, whereas CTs of large chromosomes were found more often at the nuclear periphery
Summary
Somatic cells within an organism possess genomes that are, with only a few minor exceptions, identical. For the three-dimensional (3D) and 4D (3D plus spatiotemporal) arrangements of the two haploid chromosome complements in a diploid somatic cell nucleus have been lacking so far Such 3D and 4D maps would provide the necessary foundation for studying the effect of higher-order chromatin distribution on nuclear functions, and are needed for different cell types at various stages of the cell cycle and at various stages of terminal differentiation. In addition to their importance for epigenome research, these maps should help to understand karyotype evolution [9,10,11,12] and the formation of chromosomal rearrangements in irradiated or cancer cells [13,14,15,16,17]
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