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Stable morphology, but dynamic internal reorganisation, of interphase human chromosomes in living cells.

Müller I, Boyle S, Singer RH, Bickmore WA, Chubb JR - PLoS ONE (2010)

Bottom Line: This contrasted with the behaviour of specific loci on labelled chromosomes, which showed more progressive reorganisation, and revealed that "looping out" of chromatin from chromosome territories is a dynamic state.Chromosome structure showed tremendous resistance to inhibitors of transcription, histone deacetylation and chromatin remodelling.Together, these data indicate steric constraints determine structure, rather than innate chromosome architecture or function-driven anchoring, with interphase chromatin organisation governed primarily by opposition between needs for decondensation and the space available for this to happen.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee, UK.

ABSTRACT
Despite the distinctive structure of mitotic chromosomes, it has not been possible to visualise individual chromosomes in living interphase cells, where chromosomes spend over 90% of their time. Studies of interphase chromosome structure and dynamics use fluorescence in-situ hybridisation (FISH) on fixed cells, which potentially damages structure and loses dynamic information. We have developed a new methodology, involving photoactivation of labelled histone H3 at mitosis, to visualise individual and specific human chromosomes in living interphase cells. Our data revealed bulk chromosome volume and morphology are established rapidly after mitosis, changing only incrementally after the first hour of G1. This contrasted with the behaviour of specific loci on labelled chromosomes, which showed more progressive reorganisation, and revealed that "looping out" of chromatin from chromosome territories is a dynamic state. We measured considerable heterogeneity in chromosome decondensation, even between sister chromatids, which may reflect local structural impediments to decondensation and could potentially amplify transcriptional noise. Chromosome structure showed tremendous resistance to inhibitors of transcription, histone deacetylation and chromatin remodelling. Together, these data indicate steric constraints determine structure, rather than innate chromosome architecture or function-driven anchoring, with interphase chromatin organisation governed primarily by opposition between needs for decondensation and the space available for this to happen.

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Imaging a specific locus and its chromosome territory in living human cells.Representative images of three different nuclei revealing the relative distribution of chromosome 11 and the 11q13 locus 135 minutes post-mitosis. Maximal projections of 3D stacks of chromosome 11 (green, left) and lacO repeats (red, middle) or merged (right) at 11q13 in 3 different positions: outside main territory (A), edge of territory (B) and inside territory (C). Bar 10 µm.
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pone-0011560-g006: Imaging a specific locus and its chromosome territory in living human cells.Representative images of three different nuclei revealing the relative distribution of chromosome 11 and the 11q13 locus 135 minutes post-mitosis. Maximal projections of 3D stacks of chromosome 11 (green, left) and lacO repeats (red, middle) or merged (right) at 11q13 in 3 different positions: outside main territory (A), edge of territory (B) and inside territory (C). Bar 10 µm.

Mentions: Using the 11q13-lacO HT-1080 cell line, we were able to observe 11q13 and the chromosome 11 territory together in living cells and assess whether looping out can be detected without fixation. FISH experiments on asynchronous fixed 11q13-lacO HT-1080 cells, revealed 39% of lacO hybridisation signals detected outside the territory. In living cells, we observed examples for three different states of lacO repeat position: “out” (Figure 6A), “edge” (Figure 6B) or “in” (Figure 6C) the chromosome territory. The data from 18 lacO spots at 90 min after cell division showed 2 spots outside the territory, with 5 at the edge and 11 inside (Figure 7A).


Stable morphology, but dynamic internal reorganisation, of interphase human chromosomes in living cells.

Müller I, Boyle S, Singer RH, Bickmore WA, Chubb JR - PLoS ONE (2010)

Imaging a specific locus and its chromosome territory in living human cells.Representative images of three different nuclei revealing the relative distribution of chromosome 11 and the 11q13 locus 135 minutes post-mitosis. Maximal projections of 3D stacks of chromosome 11 (green, left) and lacO repeats (red, middle) or merged (right) at 11q13 in 3 different positions: outside main territory (A), edge of territory (B) and inside territory (C). Bar 10 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2903487&req=5

pone-0011560-g006: Imaging a specific locus and its chromosome territory in living human cells.Representative images of three different nuclei revealing the relative distribution of chromosome 11 and the 11q13 locus 135 minutes post-mitosis. Maximal projections of 3D stacks of chromosome 11 (green, left) and lacO repeats (red, middle) or merged (right) at 11q13 in 3 different positions: outside main territory (A), edge of territory (B) and inside territory (C). Bar 10 µm.
Mentions: Using the 11q13-lacO HT-1080 cell line, we were able to observe 11q13 and the chromosome 11 territory together in living cells and assess whether looping out can be detected without fixation. FISH experiments on asynchronous fixed 11q13-lacO HT-1080 cells, revealed 39% of lacO hybridisation signals detected outside the territory. In living cells, we observed examples for three different states of lacO repeat position: “out” (Figure 6A), “edge” (Figure 6B) or “in” (Figure 6C) the chromosome territory. The data from 18 lacO spots at 90 min after cell division showed 2 spots outside the territory, with 5 at the edge and 11 inside (Figure 7A).

Bottom Line: This contrasted with the behaviour of specific loci on labelled chromosomes, which showed more progressive reorganisation, and revealed that "looping out" of chromatin from chromosome territories is a dynamic state.Chromosome structure showed tremendous resistance to inhibitors of transcription, histone deacetylation and chromatin remodelling.Together, these data indicate steric constraints determine structure, rather than innate chromosome architecture or function-driven anchoring, with interphase chromatin organisation governed primarily by opposition between needs for decondensation and the space available for this to happen.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee, UK.

ABSTRACT
Despite the distinctive structure of mitotic chromosomes, it has not been possible to visualise individual chromosomes in living interphase cells, where chromosomes spend over 90% of their time. Studies of interphase chromosome structure and dynamics use fluorescence in-situ hybridisation (FISH) on fixed cells, which potentially damages structure and loses dynamic information. We have developed a new methodology, involving photoactivation of labelled histone H3 at mitosis, to visualise individual and specific human chromosomes in living interphase cells. Our data revealed bulk chromosome volume and morphology are established rapidly after mitosis, changing only incrementally after the first hour of G1. This contrasted with the behaviour of specific loci on labelled chromosomes, which showed more progressive reorganisation, and revealed that "looping out" of chromatin from chromosome territories is a dynamic state. We measured considerable heterogeneity in chromosome decondensation, even between sister chromatids, which may reflect local structural impediments to decondensation and could potentially amplify transcriptional noise. Chromosome structure showed tremendous resistance to inhibitors of transcription, histone deacetylation and chromatin remodelling. Together, these data indicate steric constraints determine structure, rather than innate chromosome architecture or function-driven anchoring, with interphase chromatin organisation governed primarily by opposition between needs for decondensation and the space available for this to happen.

Show MeSH
Related in: MedlinePlus