Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Multiplexed analysis of chromosome conformation at vastly improved sensitivity

Abstract

Methods for analyzing chromosome conformation in mammalian cells are either low resolution or low throughput and are technically challenging. In next-generation (NG) Capture-C, we have redesigned the Capture-C method to achieve unprecedented levels of sensitivity and reproducibility. NG Capture-C can be used to analyze many genetic loci and samples simultaneously. High-resolution data can be produced with as few as 100,000 cells, and single-nucleotide polymorphisms can be used to generate allele-specific tracks. The method is straightforward to perform and should greatly facilitate the investigation of many questions related to gene regulation as well as the functional dissection of traits examined in genome-wide association studies.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Overview of the method.
Figure 2: Single and double oligonucleotide capture.
Figure 3: Identification of regulatory elements using comparative analysis.
Figure 4: SNP-specific interaction profiles.

Accession codes

Primary accessions

Gene Expression Omnibus

References

  1. 1

    Wang, Z., Gerstein, M. & Snyder, M. RNA-seq: a revolutionary tool for transcriptomics. Nat. Rev. Genet. 10, 57–63 (2009).

    CAS  Article  Google Scholar 

  2. 2

    Mikkelsen, T.S. et al. Genome-wide maps of chromatin state in pluripotent and lineage-committed cells. Nature 448, 553–560 (2007).

    CAS  Article  Google Scholar 

  3. 3

    Robertson, G. et al. Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencing. Nat. Methods 4, 651–657 (2007).

    CAS  Article  Google Scholar 

  4. 4

    Hesselberth, J.R. et al. Global mapping of protein-DNA interactions in vivo by digital genomic footprinting. Nat. Methods 6, 283–289 (2009).

    CAS  Article  Google Scholar 

  5. 5

    Buenrostro, J.D., Giresi, P.G., Zaba, L.C., Chang, H.Y. & Greenleaf, W.J. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat. Methods 10, 1213–1218 (2013).

    CAS  Article  Google Scholar 

  6. 6

    Dekker, J., Rippe, K., Dekker, M. & Kleckner, N. Capturing chromosome conformation. Science 295, 1306–1311 (2002).

    CAS  Article  Google Scholar 

  7. 7

    Tolhuis, B., Palstra, R.J., Splinter, E., Grosveld, F. & de Laat, W. Looping and interaction between hypersensitive sites in the active beta-globin locus. Mol. Cell 10, 1453–1465 (2002).

    CAS  Article  Google Scholar 

  8. 8

    Noordermeer, D. et al. The dynamic architecture of Hox gene clusters. Science 334, 222–225 (2011).

    CAS  Article  Google Scholar 

  9. 9

    Sanyal, A., Lajoie, B.R., Jain, G. & Dekker, J. The long-range interaction landscape of gene promoters. Nature 489, 109–113 (2012).

    CAS  Article  Google Scholar 

  10. 10

    van de Werken, H.J. et al. Robust 4C-seq data analysis to screen for regulatory DNA interactions. Nat. Methods 9, 969–972 (2012).

    CAS  Article  Google Scholar 

  11. 11

    de Laat, W. & Duboule, D. Topology of mammalian developmental enhancers and their regulatory landscapes. Nature 502, 499–506 (2013).

    CAS  Article  Google Scholar 

  12. 12

    Hughes, J.R. et al. Analysis of hundreds of cis-regulatory landscapes at high resolution in a single, high-throughput experiment. Nat. Genet. 46, 205–212 (2014).

    CAS  Article  Google Scholar 

  13. 13

    Pasquali, L. et al. Pancreatic islet enhancer clusters enriched in type 2 diabetes risk-associated variants. Nat. Genet. 46, 136–143 (2014).

    CAS  Article  Google Scholar 

  14. 14

    Maurano, M.T. et al. Systematic localization of common disease-associated variation in regulatory DNA. Science 337, 1190–1195 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. 15

    Parker, S.C. et al. Chromatin stretch enhancer states drive cell-specific gene regulation and harbor human disease risk variants. Proc. Natl. Acad. Sci. USA 110, 17921–17926 (2013).

    CAS  Article  Google Scholar 

  16. 16

    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).

    CAS  Article  Google Scholar 

  17. 17

    Jäger, R. et al. Capture Hi-C identifies the chromatin interactome of colorectal cancer risk loci. Nat. Commun. 6, 6178 (2015).

    Article  Google Scholar 

  18. 18

    Schoenfelder, S. et al. The pluripotent regulatory circuitry connecting promoters to their long-range interacting elements. Genome Res. 25, 582–597 (2015).

    CAS  Article  Google Scholar 

  19. 19

    Vernimmen, D., De Gobbi, M., Sloane-Stanley, J.A., Wood, W.G. & Higgs, D.R. Long-range chromosomal interactions regulate the timing of the transition between poised and active gene expression. EMBO J. 26, 2041–2051 (2007).

    CAS  Article  Google Scholar 

  20. 20

    Hughes, J.R. et al. High-resolution analysis of cis-acting regulatory networks at the alpha-globin locus. Phil. Trans. R. Soc. Lond. B 368, 20120361 (2013).

    Article  Google Scholar 

  21. 21

    Baù, D. et al. The three-dimensional folding of the α-globin gene domain reveals formation of chromatin globules. Nat. Struct. Mol. Biol. 18, 107–114 (2011).

    Article  Google Scholar 

  22. 22

    Simonis, M. et al. Nuclear organization of active and inactive chromatin domains uncovered by chromosome conformation capture-on-chip (4C). Nat. Genet. 38, 1348–1354 (2006).

    CAS  Article  Google Scholar 

  23. 23

    Kang, J.H. et al. Genomic organization, tissue distribution and deletion mutation of human pyridoxine 5′-phosphate oxidase. Eur. J. Biochem. 271, 2452–2461 (2004).

    CAS  Article  Google Scholar 

  24. 24

    Love, M.I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).

    Article  Google Scholar 

  25. 25

    Klein, F.A. et al. FourCSeq: analysis of 4C sequencing data. Bioinformatics 31, 3085–3091 (2015).

    CAS  Article  Google Scholar 

  26. 26

    Thongjuea, S., Stadhouders, R., Grosveld, F.G., Soler, E. & Lenhard, B. r3Cseq: an R/Bioconductor package for the discovery of long-range genomic interactions from chromosome conformation capture and next-generation sequencing data. Nucleic Acids Res. 41, e132 (2013).

    CAS  Article  Google Scholar 

  27. 27

    Osborne, C.S. et al. Active genes dynamically colocalize to shared sites of ongoing transcription. Nat. Genet. 36, 1065–1071 (2004).

    CAS  Article  Google Scholar 

  28. 28

    Noordermeer, D. et al. Variegated gene expression caused by cell-specific long-range DNA interactions. Nat. Cell Biol. 13, 944–951 (2011).

    CAS  Article  Google Scholar 

  29. 29

    Bernet, A. et al. Targeted inactivation of the major positive regulatory element (HS-40) of the human alpha-globin gene locus. Blood 86, 1202–1211 (1995).

    CAS  PubMed  Google Scholar 

  30. 30

    Anguita, E. et al. Deletion of the mouse alpha-globin regulatory element (HS-26) has an unexpectedly mild phenotype. Blood 100, 3450–3456 (2002).

    CAS  Article  Google Scholar 

  31. 31

    de Wit, E. & de Laat, W. A decade of 3C technologies: insights into nuclear organization. Genes Dev. 26, 11–24 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. 32

    Takahashi, K. & Yamanaka, S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126, 663–676 (2006).

    CAS  Article  Google Scholar 

  33. 33

    Takahashi, K. & Yamanaka, S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126, 663–676 (2006).

    CAS  Article  Google Scholar 

  34. 34

    Magoč, T. & Salzberg, S.L. FLASH: fast length adjustment of short reads to improve genome assemblies. Bioinformatics 27, 2957–2963 (2011).

    Article  Google Scholar 

  35. 35

    Kent, W.J. et al. The human genome browser at UCSC. Genome Res. 12, 996–1006 (2002).

    CAS  Article  Google Scholar 

  36. 36

    Raney, B.J. et al. Track data hubs enable visualization of user-defined genome-wide annotations on the UCSC Genome Browser. Bioinformatics 30, 1003–1005 (2014).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

J.O.J.D. thanks the Wellcome Trust for funding his work (Wellcome Trust Clinical Research Training Fellowship reference 098931/Z/12/Z). The work was also supported by a Wellcome Trust Strategic Award (reference 106130/Z/14/Z) and the Medical Research Council (MRC Core Funding and Centenary Award reference 4050189188). We thank E. Repapi for statistical advice and L. Hanssen, M. Oudelaar, D. Jeziorska, B. Graham, M. Kassouf, M. Suciu, H. Long, S. Pasricha, V. Buckle, T. Milne, T. Fulga, T. Sauka-Spengler, D. Downes and A. Drakesmith for their critique of the manuscript. We thank S. Thongjuea for discussions on analysis.

Author information

Affiliations

Authors

Contributions

J.O.J.D. performed the experiments, analyzed the data and wrote the manuscript. J.R.H. designed the experiments, assisted with the bioinformatic analysis and wrote the manuscript. N.A.R. assisted with the experiments. J.M.T. analyzed the data. S.J.M. and S.T. assisted with the bioinformatics analysis and prepared the software for public release. D.R.H. wrote the manuscript.

Corresponding author

Correspondence to Jim R Hughes.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–22 and Supplementary Note (PDF 20583 kb)

Supplementary Data

NG Capture-C supplementary data file (XLSX 55 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Davies, J., Telenius, J., McGowan, S. et al. Multiplexed analysis of chromosome conformation at vastly improved sensitivity. Nat Methods 13, 74–80 (2016). https://doi.org/10.1038/nmeth.3664

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing