Abstract
The relationship between the 4D folding of the genome and its function is an outstanding question in biology. A range of methods that probe the folding of the genome in space and time with unprecedented resolution have been developed. These methods, including chromosome conformation capture and high-resolution light and electron microscopy, are shedding new light on genome architecture and function. Here, we review the emerging picture of genome organization revealed by super-resolution and live-cell imaging. We compare and contrast population-based chromosome conformation capture approaches and imaging-based approaches and highlight future challenges.
This is a preview of subscription content, access via your institution
Access options
Access Nature and 54 other Nature Portfolio journals
Get Nature+, our best-value online-access subscription
$29.99 / 30 days
cancel any time
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Olins, A. L. & Olins, D. E. Spheroid chromatin units (v bodies). Science 183, 330–332 (1974).
Woodcock, C. L. & Ghosh, R. P. Chromatin higher-order structure and dynamics. Cold Spring Harb. Perspect. Biol. 2, a000596 (2010).
Bannister, A. J. & Kouzarides, T. Regulation of chromatin by histone modifications. Cell Res. 21, 381–395 (2011).
Filion, G. J. et al. Systematic protein location mapping reveals five principal chromatin types in Drosophila cells. Cell 143, 212–224 (2010).
Kundaje, A. et al. Integrative analysis of 111 reference human epigenomes. Nature 518, 317–330 (2015).
Breiling, A., Turner, B. M., Bianchi, M. E. & Orlando, V. General transcription factors bind promoters repressed by Polycomb group proteins. Nature 412, 651–655 (2001).
Cremer, T. & Cremer, M. Chromosome territories. Cold Spring Harb. Perspect. Biol. 2, a003889 (2010).
Padeken, J. & Heun, P. Nucleolus and nuclear periphery: Velcro for heterochromatin. Curr. Opin. Cell Biol. 28, 54–60 (2014).
Boyle, S. et al. The spatial organization of human chromosomes within the nuclei of normal and emerin-mutant cells. Hum. Mol. Genet. 10, 211–219 (2001).
Chambeyron, S. & Bickmore, W. A. Chromatin decondensation and nuclear reorganization of the HoxB locus upon induction of transcription. Genes Dev. 18, 1119–1130 (2004).
Grob, S. & Cavalli, G. Technical Review: A hitchhiker’s guide to chromosome conformation capture. Methods Mol. Biol. 1675, 233–246 (2018).
Furey, T. S. ChIP-seq and beyond: new and improved methodologies to detect and characterize protein-DNA interactions. Nat. Rev. Genet. 13, 840–852 (2012).
Rao, S. S. et al. A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell 159, 1665–1680 (2014).
Lieberman-Aiden, E. et al. Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science 326, 289–293 (2009).
Kalhor, R., Tjong, H., Jayathilaka, N., Alber, F. & Chen, L. Genome architectures revealed by tethered chromosome conformation capture and population-based modeling. Nat. Biotechnol. 30, 90–98 (2011).
Rowley, M. J. & Corces, V. G. Organizational principles of 3D genome architecture. Nat. Rev. Genet. 19, 789–800 (2018).
de Wit, E. et al. CTCF binding polarity determines chromatin looping. Mol. Cell 60, 676–684 (2015).
Guo, Y. et al. CRISPR Inversion of CTCF sites alters genome topology and enhancer/promoter function. Cell 162, 900–910 (2015).
Rao, S. S. P. et al. Cohesin loss eliminates all loop domains. Cell 171, 305–320.e324 (2017).
Schwarzer, W. et al. Two independent modes of chromatin organization revealed by cohesin removal. Nature 551, 51–56 (2017).
Gassler, J. et al. A mechanism of cohesin-dependent loop extrusion organizes zygotic genome architecture. EMBO J. 36, 3600–3618 (2017).
Nora, E. P. et al. Targeted degradation of CTCF decouples local insulation of chromosome domains from genomic compartmentalization. Cell 169, 930–944.e922 (2017).
Wutz, G. et al. Topologically associating domains and chromatin loops depend on cohesin and are regulated by CTCF, WAPL, and PDS5 proteins. EMBO J. 36, 3573–3599 (2017).
Sanborn, A. L. et al. Chromatin extrusion explains key features of loop and domain formation in wild-type and engineered genomes. Proc. Natl Acad. Sci. USA 112, E6456–E6465 (2015).
Fudenberg, G. et al. Formation of chromosomal domains by loop extrusion. Cell Rep. 15, 2038–2049 (2016).
Davidson, I. F. et al. DNA loop extrusion by human cohesin. Science 366, 1338–1345 (2019).
Kim, Y., Shi, Z., Zhang, H., Finkelstein, I. J. & Yu, H. Human cohesin compacts DNA by loop extrusion. Science 366, 1345–1349 (2019).
Hsieh, T. H. et al. Mapping nucleosome resolution chromosome folding in yeast by Micro-C. Cell 162, 108–119 (2015).
Song, F. et al. Cryo-EM study of the chromatin fiber reveals a double helix twisted by tetranucleosomal units. Science 344, 376–380 (2014).
Scheffer, M. P., Eltsov, M. & Frangakis, A. S. Evidence for short-range helical order in the 30-nm chromatin fibers of erythrocyte nuclei. Proc. Natl Acad. Sci. USA 108, 16992–16997 (2011).
McDowall, A. W., Smith, J. M. & Dubochet, J. Cryo-electron microscopy of vitrified chromosomes in situ. EMBO J. 5, 1395–1402 (1986).
Maeshima, K. & Eltsov, M. Packaging the genome: the structure of mitotic chromosomes. J. Biochem. 143, 145–153 (2008).
König, P., Braunfeld, M. B., Sedat, J. W. & Agard, D. A. The three-dimensional structure of in vitro reconstituted Xenopus laevis chromosomes by EM tomography. Chromosoma 116, 349–372 (2007).
Fussner, E. et al. Open and closed domains in the mouse genome are configured as 10-nm chromatin fibres. EMBO Rep. 13, 992–996 (2012).
Ricci, M. A., Manzo, C., García-Parajo, M. F., Lakadamyali, M. & Cosma, M. P. Chromatin fibers are formed by heterogeneous groups of nucleosomes in vivo. Cell 160, 1145–1158 (2015).
Ou, H. D. et al. ChromEMT: visualizing 3D chromatin structure and compaction in interphase and mitotic cells. Science 357, eaag0025 (2017).
Lakadamyali, M. & Cosma, M. P. Advanced microscopy methods for visualizing chromatin structure. FEBS Lett. 589(20 Pt A), 3023–3030 (2015).
Oddone, A., Vilanova, I. V., Tam, J. & Lakadamyali, M. Super-resolution imaging with stochastic single-molecule localization: concepts, technical developments, and biological applications. Microsc. Res. Tech. 77, 502–509 (2014).
Otterstrom, J. et al. Super-resolution microscopy reveals how histone tail acetylation affects DNA compaction within nucleosomes in vivo. Nucleic Acids Res. 47, 8470–8484 (2019).
Fang, K. et al. Super-resolution imaging of individual human subchromosomal regions in situ reveals nanoscopic building blocks of higher-order structure. ACS Nano 12, 4909–4918 (2018).
Nozaki, T. et al. Dynamic organization of chromatin domains revealed by super-resolution live-cell imaging. Mol. Cell 67, 282–293.e287 (2017).
Xu, J. et al. Super-resolution imaging of higher-order chromatin structures at different epigenomic states in single mammalian cells. Cell Rep. 24, 873–882 (2018).
Ferrai, C., de Castro, I. J., Lavitas, L., Chotalia, M. & Pombo, A. Gene positioning. Cold Spring Harb. Perspect. Biol. 2, a000588 (2010).
Finn, E. H. et al. Extensive heterogeneity and intrinsic variation in spatial genome organization. Cell 176, 1502–1515.e1510 (2019).
Beliveau, B. J. et al. Versatile design and synthesis platform for visualizing genomes with Oligopaint FISH probes. Proc. Natl Acad. Sci. USA 109, 21301–21306 (2012).
Beliveau, B. J. et al. In situ super-resolution imaging of genomic DNA with OligoSTORM and OligoDNA-PAINT. Methods Mol. Biol. 1663, 231–252 (2017).
Boettiger, A. N. et al. Super-resolution imaging reveals distinct chromatin folding for different epigenetic states. Nature 529, 418–422 (2016).
Bintu, B. et al. Super-resolution chromatin tracing reveals domains and cooperative interactions in single cells. Science 362, eaau1783 (2018).
Mateo, L. J. et al. Visualizing DNA folding and RNA in embryos at single-cell resolution. Nature 568, 49–54 (2019).
Nir, G. et al. Walking along chromosomes with super-resolution imaging, contact maps, and integrative modeling. PLoS Genet. 14, e1007872 (2018).
Wang, S. et al. Spatial organization of chromatin domains and compartments in single chromosomes. Science 353, 598–602 (2016).
Szabo, Q. et al. TADs are 3D structural units of higher-order chromosome organization in Drosophila. Sci. Adv. 4, eaar8082 (2018).
Imakaev, M. V., Fudenberg, G. & Mirny, L. A. Modeling chromosomes: beyond pretty pictures. FEBS Lett. 589(20 Pt A), 3031–3036 (2015).
Marti-Renom, M. A. & Mirny, L. A. Bridging the resolution gap in structural modeling of 3D genome organization. PLOS Comput. Biol. 7, e1002125 (2011).
Ozer, G., Luque, A. & Schlick, T. The chromatin fiber: multiscale problems and approaches. Curr. Opin. Struct. Biol. 31, 124–139 (2015).
Nicodemi, M. & Pombo, A. Models of chromosome structure. Curr. Opin. Cell Biol. 28, 90–95 (2014).
Mirny, L. A. The fractal globule as a model of chromatin architecture in the cell. Chromosome Res. 19, 37–51 (2011).
Tsukamoto, T. et al. Visualization of gene activity in living cells. Nat. Cell Biol. 2, 871–878 (2000).
Germier, T., Audibert, S., Kocanova, S., Lane, D. & Bystricky, K. Real-time imaging of specific genomic loci in eukaryotic cells using the ANCHOR DNA labelling system. Methods 142, 16–23 (2018).
Chen, B. et al. Dynamic imaging of genomic loci in living human cells by an optimized CRISPR/Cas system. Cell 155, 1479–1491 (2013).
Miyanari, Y., Ziegler-Birling, C. & Torres-Padilla, M. E. Live visualization of chromatin dynamics with fluorescent TALEs. Nat. Struct. Mol. Biol. 20, 1321–1324 (2013).
Chen, B., Zou, W., Xu, H., Liang, Y. & Huang, B. Efficient labeling and imaging of protein-coding genes in living cells using CRISPR-Tag. Nat. Commun. 9, 5065 (2018).
Ma, H. et al. Multicolor CRISPR labeling of chromosomal loci in human cells. Proc. Natl Acad. Sci. USA 112, 3002–3007 (2015).
Neguembor, M. V. et al. (Po)STAC (Polycistronic SunTAg modified CRISPR) enables live-cell and fixed-cell super-resolution imaging of multiple genes. Nucleic Acids Res. 46, e30 (2018).
Qin, P. et al. Live cell imaging of low- and non-repetitive chromosome loci using CRISPR-Cas9. Nat. Commun. 8, 14725 (2017).
Tanenbaum, M. E., Gilbert, L. A., Qi, L. S., Weissman, J. S. & Vale, R. D. A protein-tagging system for signal amplification in gene expression and fluorescence imaging. Cell 159, 635–646 (2014).
Gu, B. et al. Transcription-coupled changes in nuclear mobility of mammalian cis-regulatory elements. Science 359, 1050–1055 (2018).
Alexander, J. M. et al. Live-cell imaging reveals enhancer-dependent Sox2 transcription in the absence of enhancer proximity. Elife 8, e41769 (2019).
Mazza, D., Ganguly, S. & McNally, J. G. Monitoring dynamic binding of chromatin proteins in vivo by single-molecule tracking. Methods Mol. Biol. 1042, 117–137 (2013).
Liu, Z. & Tjian, R. Visualizing transcription factor dynamics in living cells. J. Cell Biol. 217, 1181–1191 (2018).
Cisse, I. I. et al. Real-time dynamics of RNA polymerase II clustering in live human cells. Science 341, 664–667 (2013).
Cho, W. K. et al. RNA Polymerase II cluster dynamics predict mRNA output in living cells. Elife 5, e13617 (2016).
Cho, W. K. et al. Mediator and RNA polymerase II clusters associate in transcription-dependent condensates. Science 361, 412–415 (2018).
Sabari, B. R. et al. Coactivator condensation at super-enhancers links phase separation and gene control. Science 361, eaar3958 (2018).
Hansen, A. S., Pustova, I., Cattoglio, C., Tjian, R. & Darzacq, X. CTCF and cohesin regulate chromatin loop stability with distinct dynamics. Elife 6, e25776 (2017).
Luppino, J.M. et al. Cohesin promotes stochastic domain intermingling to ensure proper regulation of boundary-proximal genes. Preprint at https://doi.org/10.1101/649335 (2019).
Larson, A. G. et al. Liquid droplet formation by HP1α suggests a role for phase separation in heterochromatin. Nature 547, 236–240 (2017).
Hilbert, L. et al. Transcription organizes euchromatin similar to an active microemulsion. Preprint at https://doi.org/10.1101/234112 (2018).
Gibson, B. et al. Organization of chromatin by intrinsic and regulated phase separation. Cell 179, 470–484.e421 (2019).
Williamson, I. et al. Spatial genome organization: contrasting views from chromosome conformation capture and fluorescence in situ hybridization. Genes Dev. 28, 2778–2791 (2014).
Chen, K. H., Boettiger, A. N., Moffitt, J. R., Wang, S. & Zhuang, X. RNA imaging. Spatially resolved, highly multiplexed RNA profiling in single cells. Science 348, aaa6090 (2015).
Gómez-García, P. A., Garbacik, E. T., Otterstrom, J. J., Garcia-Parajo, M. F. & Lakadamyali, M. Excitation-multiplexed multicolor superresolution imaging with fm-STORM and fm-DNA-PAINT. Proc. Natl Acad. Sci. USA 115, 12991–12996 (2018).
Wade, O. K. et al. 124-color super-resolution imaging by engineering DNA-PAINT blinking kinetics. Nano Lett. 19, 2641–2646 (2019).
Beghin, A. et al. Localization-based super-resolution imaging meets high-content screening. Nat. Methods 14, 1184–1190 (2017).
Nagano, T. et al. Single-cell Hi-C reveals cell-to-cell variability in chromosome structure. Nature 502, 59–64 (2013).
Flyamer, I. M. et al. Single-nucleus Hi-C reveals unique chromatin reorganization at oocyte-to-zygote transition. Nature 544, 110–114 (2017).
Sun, J. H. et al. Disease-associated short tandem repeats co-localize with chromatin domain boundaries. Cell 175, 224–238.e215 (2018).
Gwosch, K.C. et al. MINFLUX nanoscopy delivers 3D multicolor nanometer resolution in cells. Nat. Methods (2020).
Kim, J. H. et al. LADL: light-activated dynamic looping for endogenous gene expression control. Nat. Methods 16, 633–639 (2019).
Acknowledgements
This work was supported by the University of Pennsylvania Epigenetics Pilot Award (to M.L.), the Center for Engineering and Mechanobiology (CEMB), an NSF Science and Technology Center Pilot Award under grant agreement CMMI 15-48571 (to M.L.), a Linda Pechenik Montague Investigator Award (to M.L.), the European Union’s Horizon 2020 Research and Innovation Programme (CellViewer No 686637 to M.L. and M.P.C.), Ministerio de Ciencia e Innovación, grant BFU2017-86760-P (AEI/FEDER, UE), and an AGAUR grant from Secretaria d’Universitats i Recerca del Departament d’Empresa i Coneixement de la Generalitat de Catalunya (2017 SGR 689 to M.P.C.). We acknowledge the support of the Spanish Ministry of Science and Innovation to the EMBL partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme / Generalitat de Catalunya.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interests.
Additional information
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Lakadamyali, M., Cosma, M.P. Visualizing the genome in high resolution challenges our textbook understanding. Nat Methods 17, 371–379 (2020). https://doi.org/10.1038/s41592-020-0758-3
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/s41592-020-0758-3
This article is cited by
-
Enhancer selectivity in space and time: from enhancer–promoter interactions to promoter activation
Nature Reviews Molecular Cell Biology (2024)
-
Chromatin structure and dynamics: one nucleosome at a time
Histochemistry and Cell Biology (2024)
-
Histone FRET reports the spatial heterogeneity in nanoscale chromatin architecture that is imparted by the epigenetic landscape at the level of single foci in an intact cell nucleus
Chromosoma (2024)
-
3D genomics and its applications in precision medicine
Cellular & Molecular Biology Letters (2023)
-
Determining chromatin architecture with Micro Capture-C
Nature Protocols (2023)