CoChat
Sign Up
Cited by 289Genomics and Chromatin DynamicsChromatin Remodeling and CancerCancer-related molecular mechanisms research

Promoter-proximal CTCF binding promotes distal enhancer-dependent gene activation

Open access
Kubo, Naoki, Ishii, Haruhiko, Xiong, Xiong, Bianco, Simona, Meitinger, Franz, Hu, Rong, Hocker, James D., Conte, Mattia +8 more·2021·Nature Structural & Molecular Biology
DOI 10.1038/s41594-020-00539-5PMC PMC7913465

Published version of a preprint

Open this paper in CoChat — free account

Abstract

The CCCTC-binding factor (CTCF) works together with the cohesin complex to drive the formation of chromatin loops and topologically associating domains, but its role in gene regulation has not been fully defined. Here, we investigated the effects of acute CTCF loss on chromatin architecture and transcriptional programs in mouse embryonic stem cells undergoing differentiation to neural precursor cells. We identified CTCF-dependent enhancer-promoter contacts genome-wide and found that they disproportionately affect genes that are bound by CTCF at the promoter and are dependent on long-distance enhancers. Disruption of promoter-proximal CTCF binding reduced both long-range enhancer-promoter contacts and transcription, which were restored by artificial tethering of CTCF to the promoter. Promoter-proximal CTCF binding is correlated with the transcription of over 2,000 genes across a diverse set of adult tissues. Taken together, the results of our study show that CTCF binding to promoters may promote long-distance enhancer-dependent transcription at specific genes in diverse cell types.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1038/s41594-020-00539-5

Cites

25 of 25 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Using techniques with antibodies to enhancer- and promoter-associated histone modifications or RNA polymerase has revealed large numbers of 3D enhancer-promoter contacts.

    “Using these techniques with antibodies to enhancer- and promoter-associated histone modifications or those to RNA polymerase has also revealed large numbers of 3D enhancer–promoter contacts [244, 295–301].”

    From Transcriptional enhancers and their communication with gene promoters (2021) · cites this paper as [301]

    The Results section (page unavailable) says the authors used H3K4me3 antibodies with PLAC-seq to detect promoter-centered chromatin contacts and found thousands of contacts involving distal enhancers or promoters. This supports the histone-modification portion, but the paper does not report using RNA-polymerase antibodies or enhancer-associated histone antibodies to detect those contacts. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

  • Rapid depletion of cohesin or CTCF quickly abolishes TADs but has only relatively mild effects on gene expression.

    “rapid depletion of cohesin or CTCF quickly abolishes TADs but, as mentioned above, has only relatively mild effects on gene expression [301, 345–347, 448].”

    From Transcriptional enhancers and their communication with gene promoters (2021) · cites this paper as [301]

    The Results report that acute CTCF depletion significantly weakened TAD boundaries and left expression of most genes unaffected, supporting much of the claim. But the same Results section says some TAD boundaries remained relatively well preserved—not that TADs were abolished—and reports significant expression decreases in hundreds of genes. The paper discusses cohesin's limited expression effects but does not report a comparable rapid cohesin-depletion experiment. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

  • Cohesin-dependent enhancers tend to be longer-range than their cohesin-independent counterparts.

    “Cohesin-dependent enhancers also tend to be longer-range than their cohesin-independent counterparts [301, 349, 514].”

    From Transcriptional enhancers and their communication with gene promoters (2021) · cites this paper as [301]

    The Results section (page not provided) reports that CTCF-dependent genes generally have long-range enhancer–promoter contacts (≥100 kb), whereas CTCF-independent genes appear to rely on short-range contacts (≤50 kb), especially in neural precursor cells. This supports the distance comparison, but the paper tests dependence on CTCF loss, not cohesin loss, so it does not establish the claim specifically for cohesin-dependent versus cohesin-independent enhancers. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • Artificial tethering of CTCF to the Vcan promoter using a dCas9-CTCF construct effectively rescues the loop and, partially, the expression of Vcan.

    “Artificial tethering of CTCF to the Vcan promoter using a dCas9-CTCF construct effectively rescues the loop and, partially, the expression of Vcan [301].”

    From Transcriptional enhancers and their communication with gene promoters (2021) · cites this paper as [301]

  • Loops that require CTCF might be preferentially long-range (>100 kb) and situated in loci where enhancers are generally sparse.

    “recent studies suggest that loops that require CTCF might be preferentially long-range (> 100 kb) and situated in loci where enhancers are generally sparse [301, 349].”

    From Transcriptional enhancers and their communication with gene promoters (2021) · cites this paper as [301]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 56

  1. [2]Long HK, Prescott SL & Wysocka J. Ever-Changing Landscapes: Transcriptional Enhancers in Development and Evolution. Cell 167, 1170– 1187 (2016). 27863239 PMC5123704
  2. [5]Consortium EP An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57– 74 (2012). 22955616 PMC3439153
  3. [6]Yu M. & Ren B. The Three-Dimensional Organization of Mammalian Genomes. Annu Rev Cell Dev Biol 33, 265– 289 (2017). 28783961 PMC5837811
  4. [7]Benabdallah NS Decreased Enhancer-Promoter Proximity Accompanying Enhancer Activation. Mol Cell 76, 473–484 e7 (2019). PMC6838673 31494034
  5. [8]Alexander JM Live-cell imaging reveals enhancer-dependent Sox2 transcription in the absence of enhancer proximity. Elife 8(2019). PMC6534382 31124784
  6. [10]Phillips-Cremins JE Architectural protein subclasses shape 3D organization of genomes during lineage commitment. Cell 153, 1281– 95 (2013). 23706625 PMC3712340
  7. [12]Rao SS A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell 159, 1665– 80 (2014). 25497547 PMC5635824
  8. [16]Monahan K. Role of CCCTC binding factor (CTCF) and cohesin in the generation of single-cell diversity of protocadherin-alpha gene expression. Proc Natl Acad Sci U S A 109, 9125– 30 (2012). 22550178 PMC3384188
  9. [17]Zhang X. Fundamental roles of chromatin loop extrusion in antibody class switching. Nature 575, 385– 389 (2019). 31666703 PMC6856444
  10. [18]Lee J, Krivega I, Dale RK & Dean A. The LDB1 Complex Co-opts CTCF for Erythroid Lineage-Specific Long-Range Enhancer Interactions. Cell Rep 19, 2490– 2502 (2017). 28636938 PMC5564295
  11. [20]Arzate-Mejia RG, Recillas-Targa F. & Corces VG Developing in 3D: the role of CTCF in cell differentiation. Development 145(2018). PMC5897592 29567640
  12. [21]Stik G. CTCF is dispensable for immune cell transdifferentiation but facilitates an acute inflammatory response. Nat Genet (2020). 32514124
  13. [22]Nishimura K, Fukagawa T, Takisawa H, Kakimoto T. & Kanemaki M. An auxin-based degron system for the rapid depletion of proteins in nonplant cells. Nat Methods 6, 917– 22 (2009). 19915560
  14. [23]Holland AJ, Fachinetti D, Han JS & Cleveland DW Inducible, reversible system for the rapid and complete degradation of proteins in mammalian cells. Proc Natl Acad Sci U S A 109, E3350–7 (2012). PMC3523849 23150568
  15. [26]Mumbach MR HiChIP: efficient and sensitive analysis of protein-directed genome architecture. Nat Methods 13, 919– 922 (2016). 27643841 PMC5501173
  16. [27]Dowen JM Control of cell identity genes occurs in insulated neighborhoods in mammalian chromosomes. Cell 159, 374– 387 (2014). 25303531 PMC4197132
  17. [30]Li Y. CRISPR reveals a distal super-enhancer required for Sox2 expression in mouse embryonic stem cells. PLoS One 9, e114485 (2014). PMC4259346 25486255
  18. [31]Li G. Extensive promoter-centered chromatin interactions provide a topological basis for transcription regulation. Cell 148, 84– 98 (2012). 22265404 PMC3339270
  19. [32]Diao Y. A tiling-deletion-based genetic screen for cis-regulatory element identification in mammalian cells. Nat Methods 14, 629– 635 (2017). 28417999 PMC5490986
  20. [33]Thiecke MJ Cohesin-Dependent and -Independent Mechanisms Mediate Chromosomal Contacts between Promoters and Enhancers. Cell Rep 32, 107929 (2020). PMC7383238 32698000
  21. [34]Landolt RM, Vaughan L, Winterhalter KH & Zimmermann DR Versican is selectively expressed in embryonic tissues that act as barriers to neural crest cell migration and axon outgrowth. Development 121, 2303– 12 (1995). 7671797
  22. [36]Chiariello AM, Annunziatella C, Bianco S, Esposito A. & Nicodemi M. Polymer physics of chromosome large-scale 3D organisation. Sci Rep 6, 29775 (2016). PMC4942835 27405443
  23. [37]Bianco S. Polymer physics predicts the effects of structural variants on chromatin architecture. Nat Genet 50, 662– 667 (2018). 29662163
  24. [38]Herz HM Enhancer-associated H3K4 monomethylation by Trithorax-related, the Drosophila homolog of mammalian Mll3/Mll4. Genes Dev 26, 2604– 20 (2012). 23166019 PMC3521626
  25. [39]Hu D. The MLL3/MLL4 branches of the COMPASS family function as major histone H3K4 monomethylases at enhancers. Mol Cell Biol 33, 4745– 54 (2013). 24081332 PMC3838007
  26. [40]Yan J. Histone H3 lysine 4 monomethylation modulates long-range chromatin interactions at enhancers. Cell Res 28, 387 (2018). 29497152 PMC5835777
  27. [41]He Y. Spatiotemporal DNA methylome dynamics of the developing mouse fetus. Nature 583, 752– 759 (2020). 32728242 PMC7398276
  28. [42]Martinez O. & Reyes-Valdes MH Defining diversity, specialization, and gene specificity in transcriptomes through information theory. Proc Natl Acad Sci U S A 105, 9709– 14 (2008). 18606989 PMC2443819
  29. [43]Lee DP Robust CTCF-Based Chromatin Architecture Underpins Epigenetic Changes in the Heart Failure Stress-Gene Response. Circulation 139, 1937– 1956 (2019). 30717603
  30. [44]Wutz G. ESCO1 and CTCF enable formation of long chromatin loops by protecting cohesin(STAG1) from WAPL. Elife 9(2020). PMC7054000 32065581
  31. [45]Hsieh TS Resolving the 3D Landscape of Transcription-Linked Mammalian Chromatin Folding. Mol Cell 78, 539–553 e8 (2020). PMC7703524 32213323
  32. [46]Weintraub AS YY1 Is a Structural Regulator of Enhancer-Promoter Loops. Cell 171, 1573–1588 e28 (2017). PMC5785279 29224777
  33. [48]Caputo L. The Isl1/Ldb1 Complex Orchestrates Genome-wide Chromatin Organization to Instruct Differentiation of Multipotent Cardiac Progenitors. Cell Stem Cell 17, 287– 99 (2015). 26321200 PMC5870759
  34. [49]Monahan K, Horta A. & Lomvardas S. LHX2- and LDB1-mediated trans interactions regulate olfactory receptor choice. Nature 565, 448– 453 (2019). 30626972 PMC6436840
  35. [50]Schoenfelder S. The pluripotent regulatory circuitry connecting promoters to their long-range interacting elements. Genome Res 25, 582– 97 (2015). 25752748 PMC4381529
  36. [51]Hnisz D, Shrinivas K, Young RA, Chakraborty AK & Sharp PA A Phase Separation Model for Transcriptional Control. Cell 169, 13– 23 (2017). 28340338 PMC5432200
  37. [53]Renda M. Critical DNA binding interactions of the insulator protein CTCF: a small number of zinc fingers mediate strong binding, and a single finger-DNA interaction controls binding at imprinted loci. J Biol Chem 282, 33336–45 (2007). 17827499
  38. [54]Leitch HG Naive pluripotency is associated with global DNA hypomethylation. Nat Struct Mol Biol 20, 311– 6 (2013). 23416945 PMC3591483
  39. [55]Ficz G. FGF signaling inhibition in ESCs drives rapid genome-wide demethylation to the epigenetic ground state of pluripotency. Cell Stem Cell 13, 351– 9 (2013). 23850245 PMC3765959
  40. [56]Maurano MT Role of DNA Methylation in Modulating Transcription Factor Occupancy. Cell Rep 12, 1184– 95 (2015). 26257180
  41. [57]Nanan KK TET-Catalyzed 5-Carboxylcytosine Promotes CTCF Binding to Suboptimal Sequences Genome-wide. iScience 19, 326– 339 (2019). 31404833 PMC6699469
  42. [58]Katainen R. CTCF/cohesin-binding sites are frequently mutated in cancer. Nat Genet 47, 818– 21 (2015). 26053496
  43. [59]Kaiser VB, Taylor MS & Semple CA Mutational Biases Drive Elevated Rates of Substitution at Regulatory Sites across Cancer Types. PLoS Genet 12, e1006207 (2016). PMC4973979 27490693
  44. [61]Gribnau J, Hochedlinger K, Hata K, Li E. & Jaenisch R. Asynchronous replication timing of imprinted loci is independent of DNA methylation, but consistent with differential subnuclear localization. Genes Dev 17, 759– 73 (2003). 12651894 PMC196021
  45. [62]Strubing C. Differentiation of pluripotent embryonic stem cells into the neuronal lineage in vitro gives rise to mature inhibitory and excitatory neurons. Mech Dev 53, 275– 87 (1995). 8562428
  46. [63]Bain G, Kitchens D, Yao M, Huettner JE & Gottlieb DI Embryonic stem cells express neuronal properties in vitro. Dev Biol 168, 342– 57 (1995). 7729574
  47. [64]Li H. & Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25, 1754– 60 (2009). 19451168 PMC2705234
  48. [65]Ramirez F. deepTools2: a next generation web server for deep-sequencing data analysis. Nucleic Acids Res 44, W160– 5 (2016). 27079975 PMC4987876
  49. [66]Zhang Y. Model-based analysis of ChIP-Seq (MACS). Genome Biol 9, R137 (2008). 18798982 PMC2592715
  50. [69]Anders S, Pyl PT & Huber W. HTSeq--a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166– 9 (2015). 25260700 PMC4287950
  51. [70]Robinson MD, McCarthy DJ & Smyth GK edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26, 139– 40 (2010). 19910308 PMC2796818
  52. [71]Hu M. HiCNorm: removing biases in Hi-C data via Poisson regression. Bioinformatics 28, 3131– 3 (2012). 23023982 PMC3509491
  53. [72]Durand NC Juicebox Provides a Visualization System for Hi-C Contact Maps with Unlimited Zoom. Cell Syst 3, 99– 101 (2016). 27467250 PMC5596920
  54. [73]Crane E. Condensin-driven remodelling of X chromosome topology during dosage compensation. Nature 523, 240– 4 (2015). 26030525 PMC4498965
  55. [76]Bhattacharyya S, Chandra V, Vijayanand P. & Ay F. Identification of significant chromatin contacts from HiChIP data by FitHiChIP. Nat Commun 10, 4221 (2019). 31530818 PMC6748947
  56. [77]Kremer K. & Grest GS Dynamics of entangled linear polymer melts: A molecular-dynamics simulation. The Journal of Chemical Physics 92, 5057– 5086 (1990).

Cited by

Sorted by citations

Authors
Kubo, N., Ishii, H., Xiong, X., Bianco, S., Meitinger, F., Hu, R., Hocker, J. D., Conte, M., Gorkin, D., Yu, M., Li, B., Dixon, J. R., Hu, M., Nicodemi, M., Zhao, H., & Ren, B.
Affiliations
  • University of California San Diego (US)
  • University of Illinois Urbana-Champaign (US)
  • Istituto Nazionale di Fisica Nucleare, Sezione di Napoli (IT)
  • Fudan University (CN)
  • Salk Institute for Biological Studies (US)
  • Cleveland Clinic (US)
  • Max Delbrück Center (DE)
Corresponding author
Ren, Bing
ORCID
Venue
Nature Structural & Molecular Biology
Publisher
Springer Science and Business Media LLC
Published
2021-01-04
Volume
28
Issue
2
Pages
152-161
ISSN
  • 1545-9985 (ISSN-L)
  • 1545-9993
DOI
10.1038/s41594-020-00539-5
PMC
PMC7913465
OpenAlex
W3120280701
Cited by
289
References
25
Citations by year
2026: 25 · 2025: 66 · 2024: 59 · 2023: 55 · 2022: 55 · 2021: 29
FWCI
15.54
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Structural & Molecular Biology (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion)
  • MDC Repository (Max-Delbrueck-Center for Molecular Medicine) (submittedVersion)
  • eScholarship (California Digital Library) (submittedVersion)
Funders
  • National Institute of Mental Health (U19MH114830)
  • National Human Genome Research Institute (UM1HG011585)
  • National Cancer Institute (P30CA014195)
  • National Institute of General Medical Sciences (T32GM008666, T32GM007198)
  • National Institute of Diabetes and Digestive and Kidney Diseases (U54DK107977, U54DK107965)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chromatin Remodeling and Cancer
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer-related molecular mechanisms research
Keywords
CTCF, long-range enhancers, enhancer-promoter interactions, gene activation, chromatin looping, topologically associating domains, cohesin, chromatin structure, neural progenitor cells, mouse embryonic stem cells, transcriptional regulation
MeSH terms
Mouse Embryonic Stem Cells, CCCTC-Binding Factor, Animals, Binding Sites, Cell Line, Chromatin, Enhancer Elements, Genetic, Gene Expression Regulation, Promoter Regions, Genetic, Protein Binding, Transcriptional Activation, Mice, Neural Stem Cells

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review