Ligand dependent gene regulation by transient ERα clustered enhancers

Autoři: Bharath Saravanan aff001;  Deepanshu Soota aff001;  Zubairul Islam aff001;  Sudeshna Majumdar aff003;  Rajat Mann aff001;  Sweety Meel aff001;  Umer Farooq aff001;  Kaivalya Walavalkar aff001;  Srimonta Gayen aff003;  Anurag Kumar Singh aff001;  Sridhar Hannenhalli aff005;  Dimple Notani aff001
Působiště autorů: Genetics and Development, National Centre for Biological Sciences, Tata Institute of Fundamental Research, Bangalore, India aff001;  School of Chemical & Biotechnology, SASTRA Deemed University, Thanjavur, Tamil Nadu, India aff002;  Molecular Reproduction, Development and Genetics, Indian Institute of Science, Bangalore, India aff003;  Centre for Functional Genomics and Bio-informatics, The University of Trans-Disciplinary Health Sciences and Technology, Bangalore, India aff004;  Center for Bioinformatics and Computational Biology, University of Maryland, College Park, MD, United States of America aff005
Vyšlo v časopise: Ligand dependent gene regulation by transient ERα clustered enhancers. PLoS Genet 16(1): e32767. doi:10.1371/journal.pgen.1008516
Kategorie: Research Article
doi: 10.1371/journal.pgen.1008516


Unliganded Estrogen receptor alpha (ERα) has been implicated in ligand-dependent gene regulation. Upon ligand exposure, ERα binds to several EREs relatively proximal to the pre-marked, or persistent, ERα-bound sites and affects transient but robust gene expression. However, the underlying mechanisms are not fully understood. Here we demonstrate that upon ligand stimulation, persistent sites interact extensively, via chromatin looping, with the proximal transiently ERα-bound sites, forming Ligand Dependent ERα Enhancer Cluster in 3D (LDEC). The E2-target genes are regulated by these clustered enhancers but not by the H3K27Ac super-enhancers. Further, CRISPR-based deletion of TFF1 persistent site disrupts the formation of its LDEC resulting in the loss of E2-dependent expression of TFF1 and its neighboring genes within the same TAD. The LDEC overlap with nuclear ERα condensates that coalesce in a ligand and persistent site dependent manner. Furthermore, formation of clustered enhancers, as well as condensates, coincide with the active phase of signaling and their later disappearance results in the loss of gene expression even though persistent sites remain bound by ERα. Our results establish, at TFF1 and NRIP1 locus, a direct link between ERα condensates, ERα enhancer clusters, and transient, but robust, gene expression in a ligand-dependent fashion.

Klíčová slova:

Cell binding – Gene expression – Gene regulation – Genomic libraries – Genomic signal processing – Guide RNA – Chromatin – ChIA PET


1. Li W, Notani D, Ma Q, Tanasa B, Nunez E, Chen AY, Merkurjev D, Zhang J, Ohgi K, Song X, Oh S, Kim HS, Glass CK, Rosenfeld MG. Functional roles of enhancer RNAs for oestrogen-dependent transcriptional activation. Nature. 2013. 498(7455):516–20. doi: 10.1038/nature12210 23728302

2. Liu Z, Merkurjev D, Yang F, Li W, Oh S, Friedman MJ, Song X, Zhang F, Ma Q, Ohgi KA, Krones A, Rosenfeld MG. Enhancer activation requires trans-recruitment of a mega transcription factor complex. Cell. 2014. 159(2):358–73. doi: 10.1016/j.cell.2014.08.027 25303530

3. Hah N, Danko CG, Core L, Waterfall JJ, Siepel A, Lis JT, Kraus WL. A Rapid, Extensive, and Transient Transcriptional Response to Estrogen Signaling in Breast Cancer Cells. Cell. 2011. 145(4): 622–634. doi: 10.1016/j.cell.2011.03.042 21549415

4. Dzida T, Iqbal M, Charapitsa I, Reid G, Stunnenberg H, Matarese F, Grote K, Honkela A, Rattray M. Predicting stimulation-dependent enhancer-promoter interactions from ChIP-Seq time course data. PeerJ. 2017. 5:e3742. doi: 10.7717/peerj.3742 28970965

5. Bojcsuk D, Nagy G, Balint BL. Inducible super enhancers are organized based on canonical signal-specific transcription factor binding elements. Nucleic Acids Res. 2017. 45(7): 3693–3706. doi: 10.1093/nar/gkw1283 27994038

6. Holding AN, Cullen AE, Markowetz F. Genome-wide Estrogen Receptor-α activation is sustained, not cyclical. Elife. 2018. 7. pii: e40854. doi: 10.7554/eLife.40854 30457555

7. Welboren WJ1, van Driel MA, Janssen-Megens EM, van Heeringen SJ, Sweep FC, Span PN, Stunnenberg HG. ChIP-Seq of ERalpha and RNA polymerase II defines genes differentially responding to ligands. EMBO J. 2009. 28(10):1418–28. doi: 10.1038/emboj.2009.88 19339991

8. Caizzi L1, Ferrero G, Cutrupi S, Cordero F, Ballaré C, Miano V, Reineri S, Ricci L, Friard O, Testori A, Corà D, Caselle M, Di Croce L, De Bortoli M. Genome-wide activity of unliganded estrogen receptor-α in breast cancer cells. Proc Natl Acad Sci U S A. 2014. 111(13):4892–7. doi: 10.1073/pnas.1315445111 24639548

9. Rodriguez J, Ren G, Day CR, Zhao K, Chow CC, Larson DR. Intrinsic Dynamics of a Human Gene Reveal the Basis of Expression Heterogeneity. Cell. 2018. pii: S0092-8674(18)31518-6.

10. Jin F, Li Y, Dixon JR, Selvaraj S, Ye Z, Lee AY, Yen CA, Schmitt AD, Espinoza CA, Ren B. A high-resolution map of the three dimensional chromatin interactome in human cells. Nature. 2013 Nov 14;503(7475):290–4. doi: 10.1038/nature12644 24141950

11. Sabari BR, Dall'Agnese A, Boija A, Klein IA, Coffey EL, Shrinivas K, Abraham BJ, Hannett NM, Zamudio AV, Manteiga JC, Li CH, Guo YE, Day DS, Schuijers J, Vasile E, Malik S, Hnisz D, Lee TI, Cisse II, Roeder RG, Sharp PA, Chakraborty AK, Young RA. Coactivator condensation at super-enhancers links phase separation and gene control. Science. 2018. 361(6400).

12. Boija A, Klein IA, Sabari BR, Dall'Agnese A, Coffey EL, Zamudio AV, Li CH, Shrinivas K, Manteiga JC, Hannett NM, Abraham BJ, Afeyan LK, Guo YE, Rimel JK, Fant CB, Schuijers J, Lee TI, Taatjes DJ, Young RA. Transcription Factors Activate Genes through the Phase-Separation Capacity of Their Activation Domains. Cell. 2018. 175(7):1842–1855.e16. doi: 10.1016/j.cell.2018.10.042 30449618

13. Hnisz D, Abraham BJ, Lee TI, Lau A, Saint-André V, Sigova AA, Hoke H, Young RA. Transcriptional super-enhancers connected to cell identity and disease. Cell. 2013. 155(4):10.

14. Swinstead EE, Miranda TB, Paakinaho V, Baek S, Goldstein I, Hawkins M, Karpova TS, Ball D, Mazza D, Lavis LD, Grimm JB, Morisaki T, Grøntved L, Presman DM, Hager GL. Steroid Receptors Reprogram FoxA1 Occupancy through Dynamic Chromatin Transitions. Cell. 2016.165(3):593–605. doi: 10.1016/j.cell.2016.02.067 27062924

15. Hurtado A, Holmes KA, Ross-Innes CS, Schmidt D, Carroll, JS. FOXA1 is a key determinant of estrogen receptor function and endocrine response. Nat. Genet. 2011. 43:27–33. doi: 10.1038/ng.730 21151129

16. Carroll JS, Liu XS, Brodsky AS, Li W, Meyer CA, Szary AJ, Eeckhoute J, Shao W, Hestermann EV, Geistlinger TR, Fox EA, Silver PA, Brown M. Chromosome-wide mapping of estrogen receptor binding reveals long-range regulation requiring the forkhead protein FoxA1. Cell. 2005.122:33–43. doi: 10.1016/j.cell.2005.05.008 16009131

17. Fu X, Jeselsohn R, Pereira R, Hollingsworth EF, Creighton CJ, Li F, Shea M, Nardone A, De Angelis C, Heiser LM, Anur P, Wang N, Grasso CS, Spellman PT, Griffith OL, Tsimelzon A, Gutierrez C, Huang S, Edwards DP, Trivedi MV, Rimawi MF, Lopez-Terrada D, Hilsenbeck SG, Gray JW, Brown M, Osborne CK, Schiff R. FOXA1 overexpression mediates endocrine resistance by altering the ER transcriptome and IL-8expression in ER-positive breast cancer. Proc Natl Acad Sci U S A. 2016. 25;113(43):E6600–E6609. doi: 10.1073/pnas.1612835113 27791031

18. Hsieh CL, Fei T, Chen Y, Li T, Gao Y, Wang X, Sun T, Sweeney CJ, Lee GS, Chen S, Balk SP, Liu XS, Brown M, Kantoff PW. Enhancer RNAs participate in androgen receptordriven looping that selectively enhances gene activation. Proc Natl Acad Sci U S A. 2014. 111(20):7319–24. doi: 10.1073/pnas.1324151111 24778216

19. Li W, Notani D, Rosenfeld MG. Enhancers as non-coding RNA transcription units: recent insights and future perspectives. Nat Rev Genet. 2016. 17(4):207–23. doi: 10.1038/nrg.2016.4 26948815

20. Fullwood MJ, Liu MH, Pan YF, Liu J, Han X, Mohamed Y, Orlov YL, Velkov S, Ho A, Mei PH. An Oestrogen Receptor α-bound Human Chromatin Interactome. Nature. 2009. 462(7269): 58–64. doi: 10.1038/nature08497 19890323

21. Sanyal A, Lajoie B, Jain G, Dekker J. The long-range interaction landscape of gene promoters. Nature. 2012. 489(7414): 109–113. doi: 10.1038/nature11279 22955621

22. Quintin J, Le Péron C, Palierne G, Bizot M, Cunha S, Sérandour AA, Avner S, Henry C, Percevault F, Belaud-Rotureau MA, Huet S, Watrin E, Eeckhoute J, Legagneux V, Salbert G, Métivier R. Dynamic estrogen receptor interactomes control estrogen-responsive trefoil Factor (TFF) locus cell-specific activities. Mol Cell Biol. 2014. 34(13):2418–36. doi: 10.1128/MCB.00918-13 24752895

23. Rafique S, Thomas JS, Sproul D, Bickmore WA. Estrogen-induced chromatin decondensation and nuclear re-organization linked to regional epigenetic regulation in breast cancer. Genome Biol. 2015. 16:145. doi: 10.1186/s13059-015-0719-9 26235388

24. Larson AG, Elnatan D, Keenen MM, Trnka MJ, Johnston JB, Burlingame AL, Agard DA, Redding S, Narlikar GJ. Liquid droplet formation by HP1α suggests a role for phase separation in heterochromatin. Nature. 2017. 547(7662): 236–240. doi: 10.1038/nature22822 28636604

25. Baran-Gale J, Purvis JE, Sethupathy P. An integrative transcriptomics approach identifies miR-503 as a candidate master regulator of the estrogen response in MCF-7 breast cancer cells. RNA. 2016. 22(10):1592–603. doi: 10.1261/rna.056895.116 27539783

26. Valley CC, Solodin NM, Powers GL, Ellison SJ, Alarid ET. Temporal variation in estrogen receptor-alpha protein turnover in the presence of estrogen. J Mol Endocrinol. 2008. 40(1):23–34. doi: 10.1677/JME-07-0067 18096994

27. Reid G, Hübner MR, Métivier R, Brand H, Denger S, Manu D, Beaudouin J, Ellenberg J, Gannon F. Cyclic, proteasome-mediated turnover of unliganded and liganded ERalpha on responsive promoters is an integral feature of estrogen signaling. Mol Cell. 2003. 11(3):695–707. doi: 10.1016/s1097-2765(03)00090-x 12667452

28. Nawaz Z, Lonard DM, Dennis AP, Smith CL, O'Malley BW. Proteasome-dependent degradation of the human estrogen receptor. Proc Natl Acad Sci U S A.1999. 96:1858–62. doi: 10.1073/pnas.96.5.1858 10051559

29. Lonard DM, Nawaz Z, Smith CL, O'Malley BW. The 26S proteasome is required for estrogen receptor-α and coactivator turnover and for efficient estrogen receptor-α transactivation. Mol Cell. 2000. 5:939–48. doi: 10.1016/s1097-2765(00)80259-2 10911988

30. Callige M, Kieffer I, Richard-Foy H. CSN5/Jab1 is involved in ligand-dependent degradation of estrogen receptor {α} by the proteasome. Mol Cell Biol. 2005. 25:4349–58. doi: 10.1128/MCB.25.11.4349-4358.2005 15899841

31. Grøntved L, John S, Baek S, Liu Y, Buckley JR, Vinson C, Aguilera G et al. C/EBP maintains chromatin accessibility in liver and facilitates glucocorticoid receptor recruitment to steroid response elements. The EMBO journal. 2013. 32(11):1568–83. doi: 10.1038/emboj.2013.106 23665916

32. Grøntved L, Waterfall JJ, Kim DW, Baek S, Sung MH, Zhao L, Park JW, Nielsen R, Walker RL, Zhu YJ, Meltzer PS, Hager GL, Cheng SY. Transcriptional activation by the thyroid hormone receptor through ligand-dependent receptorrecruitment and chromatin remodelling. Nat Commun. 2015. 28;6:7048.

33. Johnson TA, Chereji RV, Stavreva DA, Morris SA, Hager GL, Clark DJ. Conventional and pioneer modes of glucocorticoid receptor interaction with enhancer chromatin in vivo. Nucleic Acids Res. 2018 Jan 9;46(1):203–214. doi: 10.1093/nar/gkx1044 29126175

34. Voss TC1, Schiltz RL, Sung MH, Yen PM, Stamatoyannopoulos JA, Biddie SC, Johnson TA, Miranda TB, John S, Hager GL. Dynamic exchange at regulatory elements during chromatin remodeling underlies assisted loading mechanism. Cell. 2011 Aug 19;146(4):544–54. doi: 10.1016/j.cell.2011.07.006 21835447

35. He HH, Meyer CA, Chen MW, Jordan VC, Brown M, Liu XS. Differential DNase I hypersensitivity reveals factor-dependent chromatin dynamics. Genome Res. 2012. (22):1015–1025.

36. Carleton JB, Berrett KC, Gertz J. Multiplex Enhancer Interference Reveals Collaborative Control of Gene Regulation by Estrogen Receptor Alpha Bound Enhancers. Cell Syst. 2017. 5(4): 333–344.e5. doi: 10.1016/j.cels.2017.08.011 28964699

37. Shin HY, Willi M, HyunYoo K, Zeng X, Wang C, Metser G, Hennighausen L. Hierarchy within the mammary STAT5-driven Wap super-enhancer. Nat Genet. 2016. 48(8): 904–911. doi: 10.1038/ng.3606 27376239

38. Williamson I, Berlivet S, Eskeland R, Boyle S, Illingworth RS, Paquette D, Dostie J, Bickmore WA. Spatial genome organization: contrasting views from chromosome conformation capture and fluorescence in situ hybridization. Genes Dev. 2014. 28(24):2778–91. doi: 10.1101/gad.251694.114 25512564

39. Wang LH, Yang XY, Zhang X, An P, Kim HJ, Huang J, Clarke R, Osborne CK, Inman JK, Appella E, Farrar WL. Disruption of estrogen receptor DNA binding domain and related intramolecular communication restores tamoxifen sensitivity in resistant breast cancer. Cancer Cell. 2006. 10(6):487–99. doi: 10.1016/j.ccr.2006.09.015 17157789

40. Shin Y, Chang YC, Lee DSW, Berry J, Sanders DW, Ronceray P, Wingreen NS, Haataja M, Brangwynne CP.Liquid Nuclear Condensates Mechanically Sense and Restructure the Genome. Cell. 2018. 175(6):1481–1491. doi: 10.1016/j.cell.2018.10.057 30500535

41. Kang YK1, Guermah M, Yuan CX, Roeder RG. The TRAP/Mediator coactivator complex interacts directly with estrogen receptors alpha and beta through the TRAP220 subunit and directly enhances estrogen receptor function in vitro. Proc Natl Acad Sci U S A. 2002. 99(5):2642–7. doi: 10.1073/pnas.261715899 11867769

42. Malik S, Roeder RG. Nuclear Receptors. Elsevier Methods in Enzymology. 2003. 364257–284.

43. Tanida T, Matsuda KI, Yamada S, Hashimoto T, Kawata M. Estrogen-related Receptor β Reduces the Subnuclear Mobility of Estrogen Receptor α and Suppresses Estrogen-dependent Cellular Function. J Biol Chem. 2015. 290(19):12332–45. doi: 10.1074/jbc.M114.619098 25805499

44. Liu J, Perumal NB, Oldfield CJ, Su EW, Uversky VN, Dunker AK. Intrinsic disorder in transcription factors. Biochemistry. 2006 45(22): 6873–88. doi: 10.1021/bi0602718 16734424

45. Harmon TS, Holehouse AS, Rosen MK, Pappu RV. Intrinsically disordered linkers determine the interplay between phase separation and gelation in multivalent proteins. elife. 2017. 6, e30294. doi: 10.7554/eLife.30294 29091028

46. Lai F, Orom UA, Cesaroni M, Beringer M, Taatjes DJ, Blobel GA, Shiekhattar R. Activating RNAs associate with Mediator to enhance chromatin architecture and transcription. Nature. 2013. 494(7438):497–50. doi: 10.1038/nature11884 23417068

47. Cheng D, Vemulapalli V, Lu Y, Shen J, Aoyagi S, Fry CJ, Yang Y, Foulds CE, Stossi F, Treviño LS, Mancini MA, O'Malley BW, Walker CL, Boyer TG, Bedford MT. CARM1 methylates MED12 to regulate its RNA-binding ability. Life Sci Alliance. 2018. 1(5).

48. Lin Y, Protter DS, Rosen MK, Parker R. Formation and Maturation of Phase-Separated Liquid Droplets by RNA-Binding Proteins. Mol Cell. 2015. 60(2):208–19. doi: 10.1016/j.molcel.2015.08.018 26412307

49. Banani SF, Rice AM, Peeples WB, Lin Y, Jain S, Parker R, Rosen MK. Compositional Control of Phase-Separated Cellular Bodies. Cell. 2016. 166(3):651–663. doi: 10.1016/j.cell.2016.06.010 27374333

50. Fan M, Nakshatri H, Nephew KP. Inhibiting proteasomal proteolysis sustains estrogen receptor-alpha activation. Mol Endocrinol. 2004. (11):2603–15. doi: 10.1210/me.2004-0164 15284335

51. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods. 2012 Mar 4;9(4):357–9. doi: 10.1038/nmeth.1923 22388286

52. Heinz S, Benner C, Spann N, Bertolino E, Lin YC, Laslo P, Cheng JX, Murre C, Singh H, Glass CK. Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell. 2010 May 28;38(4):576–89. doi: 10.1016/j.molcel.2010.05.004 20513432

53. van Berkum NL, Dekker J. Determining spatial chromatin organization of large genomic regions using 5C technology. Methods Mol Biol. 2009. 567:189–213. doi: 10.1007/978-1-60327-414-2_13 19588094

54. Ferraiuolo MA, Sanyal A, Naumova N, Dekker J, and Dostie J. Mapping chromatin interactions with 5C technology. 5C; a quantitative approach to capturing chromatin conformation over large genomic distances. Methods. 2012. 58(3): 10.1016.

55. Notani D, Gottimukkala KP, Jayani RS, Limaye AS, Damle MV, Mehta S, Purbey PK, Joseph J, Galande S.Global regulator SATB1 recruits beta-catenin and regulates T(H)2 differentiation in Wnt-dependent manner. PLoS Biol. 2010. 8(1):e1000296. doi: 10.1371/journal.pbio.1000296 20126258

56. Gayen S, Maclary E, Buttigieg E, Hinten M, Kalantry S. A Primary Role for the Tsix lncRNA in Maintaining Random X-Chromosome Inactivation. Cell Rep. 2015. 11(8):1251–65. doi: 10.1016/j.celrep.2015.04.039 25981039

57. Schindelin J1, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, Preibisch S, Rueden C, Saalfeld S, Schmid B, Tinevez JY, White DJ, Hartenstein V, Eliceiri K, Tomancak P, Cardona A. Fiji: an open-source platform for biological-image analysis. Nat Methods. 2012. 289(7):676–82.

58. Sprague BL, Pego RL, Stavreva DA, McNally JG. Analysis of binding reactions by fluorescence recovery after photobleaching. Biophys J. 2004. 86:3473–3495. doi: 10.1529/biophysj.103.026765 15189848

59. Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Genome engineering using the CRISPR-Cas9 system. Nat Protoc. 2013. 8(11):2281–308. doi: 10.1038/nprot.2013.143 24157548

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