Spatial genome organization: contrasting views from chromosome conformation capture and fluorescence in situ hybridization

Williamson, Iain; Berlivet, Soizik; Eskeland, Ragnhild; Boyle, Shelagh; Illingworth, Robert S.; Paquette, Denis; Dostie, Josée; Bickmore, Wendy A.

doi:10.1101/gad.251694.114

Cited by 237 publications

(251 citation statements)

References 74 publications

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“…20), and a chromatin structure in the nuclear space of single cells, on the other hand, is of great interest and has recently come under discussion (see ref. 21). In this study, we used DNA FISH to show that the two TADs splitting the HoxD locus are distinct chromatin units, which rarely overlap despite their close association in space.…”

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confidence: 99%

Nanoscale spatial organization of the HoxD gene cluster in distinct transcriptional states

Fabre

Benke

Joye

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Chromatin condensation plays an important role in the regulation of gene expression. Recently, it was shown that the transcriptional activation of Hoxd genes during vertebrate digit development involves modifications in 3D interactions within and around the HoxD gene cluster. This reorganization follows a global transition from one set of regulatory contacts to another, between two topologically associating domains (TADs) located on either side of the HoxD locus. Here, we use 3D DNA FISH to assess the spatial organization of chromatin at and around the HoxD gene cluster and report that although the two TADs are tightly associated, they appear as spatially distinct units. We measured the relative position of genes within the cluster and found that they segregate over long distances, suggesting that a physical elongation of the HoxD cluster can occur. We analyzed this possibility by super-resolution imaging (STORM) and found that tissues with distinct transcriptional activity exhibit differing degrees of elongation. We also observed that the morphological change of the HoxD cluster in developing digits is associated with its position at the boundary between the two TADs. Such variations in the fine-scale architecture of the gene cluster suggest causal links among its spatial configuration, transcriptional activation, and the flanking chromatin context. DNA FISH | super-resolution microscopy | limb development | topologically associating domains | HoxD

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mentioning

confidence: 99%

Nanoscale spatial organization of the HoxD gene cluster in distinct transcriptional states

Fabre

Benke

Joye

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…The distribution of TADs is also supported by 3D-FISH (Nora et al 2012). However, some genome organization revealed by chromosome conformation capture techniques could not be confirmed by 3D-FISH (Williamson et al 2014). The inconsistency would be derived from the technical difference between FISH and Hi-C.…”

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confidence: 77%

FISH Is in the Limelight Again As More Than a Cytogenetical Technique for Metaphase Chromosomes

Matsunaga

2016

CYTOLOGIA

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Summary Fluorescent in situ hybridization (FISH) has been a powerful technique to reveal chromosomal loci and distribution of DNA sequences in metaphase chromosomes. Three-dimensional FISH (3D-FISH) has also revealed the chromosome territory of interphase chromosomes in the nucleus. Recently, 3D-FISH has played a supportive or complementary role to confirm the genome organization and topologically associated domains, which are provided by chromosome conformation capture techniques.

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“…Live-cell PWS is a natural supplement to superresolution fluorescence techniques, providing quantifiable information about unstained cellular organization to examine the role of the nanoarchitecture on molecular interactions in live cells. In the future, we envision that live-cell PWS can be applied to a broad range of critical studies of structure-function in live cells, leveraging the multimodal potential in conjunction with existing SRM to study: (i) the interaction between chromatin structure and mRNA transport; (ii) the accessibility of euchromatin and heterochromatin to transcription factors (45)(46)(47); (iii) the relationship between chromatin looping, as measured by techniques such as Hi-C, to the physical chromatin structure (6,16,48); (iv) why and how higher-order chromatin structure changes in cancer (14); (v) the role of nuclear architecture as an epigenetic regulator of gene expression (6,16,48); (vi) the effect of metabolism on chromatin structure (40,49); and (vii) the role of chromatin dynamics in stem cell development (50,51).…”

Section: Resultsmentioning

confidence: 99%

Label-free imaging of the native, living cellular nanoarchitecture using partial-wave spectroscopic microscopy

Almassalha

Bauer

Chandler

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The organization of chromatin is a regulator of molecular processes including transcription, replication, and DNA repair. The structures within chromatin that regulate these processes span from the nucleosomal (10-nm) to the chromosomal (>200-nm) levels, with little known about the dynamics of chromatin structure between these scales due to a lack of quantitative imaging technique in live cells. Previous work using partial-wave spectroscopic (PWS) microscopy, a quantitative imaging technique with sensitivity to macromolecular organization between 20 and 200 nm, has shown that transformation of chromatin at these length scales is a fundamental event during carcinogenesis. As the dynamics of chromatin likely play a critical regulatory role in cellular function, it is critical to develop live-cell imaging techniques that can probe the real-time temporal behavior of the chromatin nanoarchitecture. Therefore, we developed a livecell PWS technique that allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in real time. In this work, we use live-cell PWS to study the change in chromatin structure due to DNA damage and expand on the link between metabolic function and the structure of higherorder chromatin. In particular, we studied the temporal changes to chromatin during UV light exposure, show that live-cell DNA-binding dyes induce damage to chromatin within seconds, and demonstrate a direct link between higher-order chromatin structure and mitochondrial membrane potential. Because biological function is tightly paired with structure, live-cell PWS is a powerful tool to study the nanoscale structure-function relationship in live cells.E very cellular and extracellular structure has a complex nanoscale organization ranging from individual macromolecules that are a few nanometers in size (e.g., protein and DNA) to macromolecular assemblies that are tens to hundreds of nanometers in size (e.g., cell membranes, higher-order chromatin structure, cytoskeleton, and extracellular matrix fibers). A major scientific challenge is to understand these macromolecular structures, specifically their function and interactions in structurally and dynamically complex living cellular systems. To meet these goals, the ideal live-cell imaging technology would satisfy six key requirements: being (i) nanoscale sensitive (<200 nm), (ii) label free, (iii) nonperturbing, (iv) quantitative, (v) high throughput, and (vi) molecularly informative.Current approaches are unable to meet all of these criteria alone. The breakthroughs in superresolution fluorescence microscopy (SRM) have enabled new imaging technologies capable of providing unprecedented molecular identification at the highest resolutions currently available in live cells, but require the use of exogenous fluorophores to visualize macromolecular structures (1-3). For some applications, these labels are indispensable to achieve molecular specificity. However, there are significant drawbacks to the exclusive use of molec...

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Spatial genome organization: contrasting views from chromosome conformation capture and fluorescence in situ hybridization

Cited by 237 publications

References 74 publications

Nanoscale spatial organization of the HoxD gene cluster in distinct transcriptional states

Nanoscale spatial organization of the HoxD gene cluster in distinct transcriptional states

FISH Is in the Limelight Again As More Than a Cytogenetical Technique for Metaphase Chromosomes

Label-free imaging of the native, living cellular nanoarchitecture using partial-wave spectroscopic microscopy

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