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An insulator blocks access to enhancers by an illegitimate promoter, preventing repression by transcriptional interference


Autoři: Miki Fujioka aff001;  Anastasiya Nezdyur aff001;  James B. Jaynes aff001
Působiště autorů: Thomas Jefferson University, Philadelphia, Pennsylvania, United States of America aff001
Vyšlo v časopise: An insulator blocks access to enhancers by an illegitimate promoter, preventing repression by transcriptional interference. PLoS Genet 17(4): e1009536. doi:10.1371/journal.pgen.1009536
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.pgen.1009536

Souhrn

Several distinct activities and functions have been described for chromatin insulators, which separate genes along chromosomes into functional units. Here, we describe a novel mechanism of functional separation whereby an insulator prevents gene repression. When the homie insulator is deleted from the end of a Drosophila even skipped (eve) locus, a flanking P-element promoter is activated in a partial eve pattern, causing expression driven by enhancers in the 3’ region to be repressed. The mechanism involves transcriptional read-through from the flanking promoter. This conclusion is based on the following. Read-through driven by a heterologous enhancer is sufficient to repress, even when homie is in place. Furthermore, when the flanking promoter is turned around, repression is minimal. Transcriptional read-through that does not produce anti-sense RNA can still repress expression, ruling out RNAi as the mechanism in this case. Thus, transcriptional interference, caused by enhancer capture and read-through when the insulator is removed, represses eve promoter-driven expression. We also show that enhancer-promoter specificity and processivity of transcription can have decisive effects on the consequences of insulator removal. First, a core heat shock 70 promoter that is not activated well by eve enhancers did not cause read-through sufficient to repress the eve promoter. Second, these transcripts are less processive than those initiated at the P-promoter, measured by how far they extend through the eve locus, and so are less disruptive. These results highlight the importance of considering transcriptional read-through when assessing the effects of insulators on gene expression.

Klíčová slova:

DNA transcription – Gene expression – Genetic interference – Genetic loci – In situ hybridization – Insulators – Non-coding RNA – Embryonic pattern formation


Zdroje

1. Schwartz YB, Cavalli G. Three-Dimensional Genome Organization and Function in Drosophila. Genetics. 2017;205(1):5–24. doi: 10.1534/genetics.115.185132 28049701.

2. Geyer PK, Corces VG. DNA position-specific repression of transcription by a Drosophila zinc finger protein. Genes Dev. 1992;6(10):1865–73. doi: 10.1101/gad.6.10.1865 1327958.

3. Kellum R, Schedl P. A position-effect assay for boundaries of higher order chromosomal domains. Cell. 1991;64(5):941–50. doi: 10.1016/0092-8674(91)90318-s 1848159.

4. Holdridge C, Dorsett D. Repression of hsp70 heat shock gene transcription by the suppressor of hairy-wing protein of Drosophila melanogaster. Mol Cell Biol. 1991;11(4):1894–900. doi: 10.1128/mcb.11.4.1894 1900919

5. Bell AC, West AG, Felsenfeld G. The protein CTCF is required for the enhancer blocking activity of vertebrate insulators. Cell. 1999;98(3):387–96. doi: 10.1016/s0092-8674(00)81967-4 10458613.

6. Hark AT, Schoenherr CJ, Katz DJ, Ingram RS, Levorse JM, Tilghman SM. CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus.[see comment]. Nature. 2000;405(6785):486–9. doi: 10.1038/35013106 10839547.

7. Fujioka M, Wu X, Jaynes JB. A chromatin insulator mediates transgene homing and very long-range enhancer-promoter communication. Development. 2009;136(18):3077–87. doi: 10.1242/dev.036467 19675129.

8. Sigrist CJ, Pirrotta V. Chromatin insulator elements block the silencing of a target gene by the Drosophila polycomb response element (PRE) but allow trans interactions between PREs on different chromosomes. Genetics. 1997;147(1):209–21. 9286681.

9. Mallin DR, Myung JS, Patton JS, Geyer PK. Polycomb group repression is blocked by the Drosophila suppressor of Hairy-wing [su(Hw)] insulator. Genetics. 1998;148(1):331–9. 9475743.

10. Comet I, Schuettengruber B, Sexton T, Cavalli G. A chromatin insulator driving three-dimensional Polycomb response element (PRE) contacts and Polycomb association with the chromatin fiber. Proc Natl Acad Sci USA. 2011;108(6):2294–9. doi: 10.1073/pnas.1002059108 21262819.

11. Kahn TG, Schwartz YB, Dellino GI, Pirrotta V. Polycomb complexes and the propagation of the methylation mark at the Drosophila Ubx gene. J Biol Chem. 2006;281(39):29064–75. doi: 10.1074/jbc.M605430200 16887811.

12. Fujioka M, Sun G, Jaynes JB. The Drosophila eve insulator Homie promotes eve expression and protects the adjacent gene from repression by Polycomb spreading. PLoS Genet. 2013;9(10):e1003883. doi: 10.1371/journal.pgen.1003883 24204298.

13. Muller M, Hagstrom K, Gyurkovics H, Pirrotta V, Schedl P. The mcp element from the Drosophila melanogaster bithorax complex mediates long-distance regulatory interactions. Genetics. 1999;153(3):1333–56. 10545463.

14. Blanton J, Gaszner M, Schedl P. Protein:protein interactions and the pairing of boundary elements in vivo. Genes Dev. 2003;17(5):664–75. doi: 10.1101/gad.1052003 12629048.

15. Parnell TJ, Viering MM, Skjesol A, Helou C, Kuhn EJ, Geyer PK. An endogenous suppressor of hairy-wing insulator separates regulatory domains in Drosophila. Proc Natl Acad Sci USA. 2003;100(23):13436–41. doi: 10.1073/pnas.2333111100 14597701.

16. Spilianakis CG, Flavell RA. Long-range intrachromosomal interactions in the T helper type 2 cytokine locus. Nat Immunol. 2004;5(10):1017–27. doi: 10.1038/ni1115 15378057.

17. Lomvardas S, Barnea G, Pisapia DJ, Mendelsohn M, Kirkland J, Axel R. Interchromosomal interactions and olfactory receptor choice. Cell. 2006;126(2):403–13. doi: 10.1016/j.cell.2006.06.035 16873069

18. Hou C, Zhao H, Tanimoto K, Dean A. CTCF-dependent enhancer-blocking by alternative chromatin loop formation. Proc Natl Acad Sci USA. 2008;105(51):20398–403. doi: 10.1073/pnas.0808506106 19074263.

19. Fujioka M, Mistry H, Schedl P, Jaynes JB. Determinants of Chromosome Architecture: Insulator Pairing in cis and in trans. PLoS Genet. 2016;12(2):e1005889. doi: 10.1371/journal.pgen.1005889 26910731.

20. Viets K, Sauria ME, Chernoff C, Viales RR, Echterling M, Anderson C, et al. Characterization of button loci that promote homologous chromosome pairing and cell-type-specific interchromosomal gene regulation. Dev Cell. 2019;51(3):341–56. e7. doi: 10.1016/j.devcel.2019.09.007 31607649

21. Gyurkovics H, Gausz J, Kummer J, Karch F. A new homeotic mutation in the Drosophila bithorax complex removes a boundary separating two domains of regulation. The EMBO journal. 1990;9(8):2579–85. 1973385

22. Galloni M, Gyurkovics H, Schedl P, Karch F. The bluetail transposon: evidence for independent cis-regulatory domains and domain boundaries in the bithorax complex. The EMBO Journal. 1993;12(3):1087–97. 8384551

23. Karch F, Galloni M, Sipos L, Gausz J, Gyurkovics H, Schedl P. Mcp and Fab-7: molecular analysis of putative boundaries of cis-regulatory domains in the bithorax complex of Drosophila melanogaster. Nucleic Acids Res. 1994;22(15):3138–46. doi: 10.1093/nar/22.15.3138 7915032.

24. Hagstrom K, Muller M, Schedl P. Fab-7 functions as a chromatin domain boundary to ensure proper segment specification by the Drosophila bithorax complex. Genes Dev. 1996;10(24):3202–15. doi: 10.1101/gad.10.24.3202 8985188.

25. Bender W, Hudson A. P element homing to the Drosophila bithorax complex. Development. 2000;127(18):3981–92. 10952896.

26. Mihaly J, Hogga I, Gausz J, Gyurkovics H, Karch F. In situ dissection of the Fab-7 region of the bithorax complex into a chromatin domain boundary and a Polycomb-response element. Development. 1997;124(9):1809–20. 9165128.

27. Hagstrom K, Muller M, Schedl P. A Polycomb and GAGA dependent silencer adjoins the Fab-7 boundary in the Drosophila bithorax complex. Genetics. 1997;146(4):1365–80. 9258680.

28. Negre N, Brown CD, Shah PK, Kheradpour P, Morrison CA, Henikoff JG, et al. A Comprehensive Map of Insulator Elements for the Drosophila Genome. PLoS Genet. 2010;6(6):e1000814. doi: 10.1371/journal.pgen.1000814 20084099

29. Matthews NE, White R. Chromatin Architecture in the Fly: Living without CTCF/Cohesin Loop Extrusion? 2019.

30. Rowley MJ, Corces VG. Organizational principles of 3D genome architecture. Nature Reviews Genetics. 2018;19(12):789–800. doi: 10.1038/s41576-018-0060-8 30367165

31. Mateo LJ, Murphy SE, Hafner A, Cinquini IS, Walker CA, Boettiger AN. Visualizing DNA folding and RNA in embryos at single-cell resolution. Nature. 2019;568(7750):49. doi: 10.1038/s41586-019-1035-4 30886393

32. Bender W, Lucas M. The Border Between the Ultrabithorax and abdominal-A Regulatory Domains in the Drosophila Bithorax Complex. Genetics. 2013;193(4):1135–47. doi: 10.1534/genetics.112.146340 23288934 [Available on 04/01/14].

33. Savitsky M, Kim M, Kravchuk O, Schwartz YB. Distinct Roles of Chromatin Insulator Proteins in Control of the Drosophila Bithorax Complex. Genetics. 2016;202(2):601–17. Epub 2015/12/31. doi: 10.1534/genetics.115.179309 26715665.

34. Narendra V, Rocha PP, An D, Raviram R, Skok JA, Mazzoni EO, et al. CTCF establishes discrete functional chromatin domains at the Hox clusters during differentiation. Science. 2015;347(6225):1017–21. doi: 10.1126/science.1262088 25722416

35. Narendra V, Bulajić M, Dekker J, Mazzoni EO, Reinberg D. CTCF-mediated topological boundaries during development foster appropriate gene regulation. Genes Dev. 2016;30(24):2657–62. doi: 10.1101/gad.288324.116 28087711

36. Rao SS, Huntley MH, Durand NC, Stamenova EK, Bochkov ID, Robinson JT, et al. A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell. 2014;159(7):1665–80. doi: 10.1016/j.cell.2014.11.021 25497547.

37. Dixon JR, Selvaraj S, Yue F, Kim A, Li Y, Shen Y, et al. Topological domains in mammalian genomes identified by analysis of chromatin interactions. Nature. 2012;485(7398):376–80. doi: 10.1038/nature11082 22495300.

38. Schwartz YB, Linder-Basso D, Kharchenko PV, Tolstorukov MY, Kim M, Li H-B, et al. Nature and function of insulator protein binding sites in the Drosophila genome.[Erratum appears in Genome Res. 2013 Feb;23(2):409]. Genome Res. 2012;22(11):2188–98. doi: 10.1101/gr.138156.112 22767387.

39. Lieberman-Aiden E, van Berkum NL, Williams L, Imakaev M, Ragoczy T, Telling A, et al. Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science. 2009;326(5950):289–93. doi: 10.1126/science.1181369 19815776;

40. Rowley MJ, Nichols MH, Lyu X, Ando-Kuri M, Rivera ISM, Hermetz K, et al. Evolutionarily conserved principles predict 3D chromatin organization. Mol Cell. 2017;67(5):837–52. e7. doi: 10.1016/j.molcel.2017.07.022 28826674

41. Kaikkonen MU, Adelman K. Emerging roles of non-coding RNA transcription. Trends Biochem Sci. 2018;43(9):654–67. doi: 10.1016/j.tibs.2018.06.002 30145998

42. Mellor J, Woloszczuk R, Howe FS. The Interleaved Genome. Trends Genet. 2016;32(1):57–71. doi: 10.1016/j.tig.2015.10.006 26613890.

43. Kornienko AE, Guenzl PM, Barlow DP, Pauler FM. Gene regulation by the act of long non-coding RNA transcription. BMC Biol. 2013;11(1):59. doi: 10.1186/1741-7007-11-59 23721193

44. Erokhin M, Davydova A, Parshikov A, Studitsky VM, Georgiev P, Chetverina D. Transcription through enhancers suppresses their activity in Drosophila. Epigenetics & chromatin. 2013;6(1):31. doi: 10.1186/1756-8935-6-31 24279291.

45. Bender W, Fitzgerald DP. Transcription activates repressed domains in the Drosophila bithorax complex. Development. 2002;129(21):4923–30. 12397101

46. Hogga I, Karch F. Transcription through the iab-7 cis-regulatory domain of the bithorax complex interferes with maintenance of Polycomb-mediated silencing. Development. 2002;129(21):4915–22. 12397100

47. Gummalla M, Maeda RK, Castro Alvarez JJ, Gyurkovics H, Singari S, Edwards KA, et al. abd-A regulation by the iab-8 noncoding RNA. PLoS Genet. 2012;8(5):e1002720. doi: 10.1371/journal.pgen.1002720 22654672.

48. Fujioka M, Yusibova GL, Patel NH, Brown SJ, Jaynes JB. The repressor activity of Even-skipped is highly conserved, and is sufficient to activate engrailed and to regulate both the spacing and stability of parasegment boundaries. Development. 2002;129(19):4411–21. 12223400

49. Fujioka M, Jaynes JB, Goto T. Early even-skipped stripes act as morphogenetic gradients at the single cell level to establish engrailed expression. Development. 1995;121(12):4371–82. 8575337.

50. Ruden DM, Sollars V, Wang X, Mori D, Alterman M, Lu X. Membrane fusion proteins are required for oskar mRNA localization in the Drosophila egg chamber. Dev Biol. 2000;218(2):314–25. doi: 10.1006/dbio.1999.9583 10656772.

51. Leon A, McKearin D. Identification of TER94, an AAA ATPase protein, as a Bam-dependent component of the Drosophila fusome. Molecular Biology of the Cell. 1999;10(11):3825–34. doi: 10.1091/mbc.10.11.3825 10564274.

52. Pinter M, Jekely G, Szepesi RJ, Farkas A, Theopold U, Meyer HE, et al. TER94, a Drosophila homolog of the membrane fusion protein CDC48/p97, is accumulated in nonproliferating cells: in the reproductive organs and in the brain of the imago. Insect Biochemistry & Molecular Biology. 1998;28(2):91–8. doi: 10.1016/s0965-1748(97)00095-7 9639875.

53. Bateman JR, Lee AM, Wu CT. Site-specific transformation of Drosophila via phiC31 integrase-mediated cassette exchange. Genetics. 2006;173(2):769–77. doi: 10.1534/genetics.106.056945 16547094.

54. Sackerson C, Fujioka M, Goto T. The even-skipped locus is contained in a 16-kb chromatin domain. Dev Biol. 1999;211(1):39–52. doi: 10.1006/dbio.1999.9301 10373303

55. Fujioka M, Emi-Sarker Y, Yusibova GL, Goto T, Jaynes JB. Analysis of an even-skipped rescue transgene reveals both composite and discrete neuronal and early blastoderm enhancers, and multi-stripe positioning by gap gene repressor gradients. Development. 1999;126(11):2527–38. 10226011

56. Small S, Blair A, Levine M. Regulation of two pair-rule stripes by a single enhancer in the Drosophila embryo. Dev Biol. 1996;175(May 1):314–24. doi: 10.1006/dbio.1996.0117 8626035

57. Small S, Blair A, Levine M. Regulation of even-skipped stripe 2 in the Drosophila embryo. The EMBO journal. 1992;11(11):4047–57. 1327756

58. Harding K, Hoey T, Warrior R, Levine M. Autoregulatory and gap gene response elements of the even-skipped promoter of Drosophila. The EMBO Journal. 1989;8:1205–12. 2743979

59. Goto T, Macdonald P, Maniatis T. Early and late periodic patterns of even skipped expression are controlled by distinct regulatory elements that respond to different spatial cues. Cell. 1989;57:413–22. doi: 10.1016/0092-8674(89)90916-1 2720776

60. Hodgetts RB, O’Keefe SL. The mutant phenotype associated with P-element alleles of the vestigial locus in Drosophila melanogaster may be caused by a readthrough transcript initiated at the P-element promoter. Genetics. 2001;157(4):1665–72. 11290721

61. Kaufman PD, Rio DC. Drosophila P-element transposase is a transcriptional repressor in vitro. Proc Natl Acad Sci USA. 1991;88(7):2613–7. doi: 10.1073/pnas.88.7.2613 1849267.

62. Rio DC, Rubin GM. Identification and purification of a Drosophila protein that binds to the terminal 31-base-pair inverted repeats of the P transposable element. Proceedings of the National Academy of Sciences. 1988;85(23):8929–33. doi: 10.1073/pnas.85.23.8929 2848246

63. Rubin GM, Spradling AC. Genetic transformation of Drosophila with transposable element vectors. Science. 1982;218(4570):348–53. doi: 10.1126/science.6289436 6289436

64. Theurkauf WE, Baum H, Bo J, Wensink PC. Tissue-specific and constitutive alpha-tubulin genes of Drosophila melanogaster code for structurally distinct proteins. Proceedings of the National Academy of Sciences. 1986;83(22):8477–81. doi: 10.1073/pnas.83.22.8477 3095837

65. Connelly S, Manley JL. A functional mRNA polyadenylation signal is required for transcription termination by RNA polymerase II. Genes Dev. 1988;2(4):440–52. doi: 10.1101/gad.2.4.440 2836265

66. Hiromi Y, Gehring WJ. Regulation and function of the Drosophila segmentation gene fushi tarazu. Cell. 1987;50(6):963–74. doi: 10.1016/0092-8674(87)90523-x 2887293

67. Venken KJT, Schulze KL, Haelterman NA, Pan H, He Y, Evans-Holm M, et al. MiMIC: a highly versatile transposon insertion resource for engineering Drosophila melanogaster genes. Nat Methods. 2011;8(9):737–43. doi: 10.1038/nmeth.1662 21985007.

68. Ip YT, Park RE, Kosman D, Bier E, Levine M. The dorsal gradient morphogen regulates stripes of rhomboid expression in the presumptive neuroectoderm of the Drosophila embryo. Genes Dev. 1992;6(9):1728–39. doi: 10.1101/gad.6.9.1728 1325394

69. Zabidi MA, Arnold CD, Schernhuber K, Pagani M, Rath M, Frank O, et al. Enhancer–core-promoter specificity separates developmental and housekeeping gene regulation. Nature. 2015;518(7540):556. doi: 10.1038/nature13994 25517091

70. Maeda RK, Karch F. The ABC of the BX-C: the bithorax complex explained. Development. 2006;133(8):1413–22. doi: 10.1242/dev.02323 16556913.

71. Karess RE, Rubin GM. Analysis of P transposable element functions in Drosophila. Cell. 1984;38(1):135–46. doi: 10.1016/0092-8674(84)90534-8 6088058

72. Laski FA, Rio DC, Rubin GM. Tissue specificity of Drosophila P element transposition is regulated at the level of mRNA splicing. Cell. 1986;44(1):7–19. doi: 10.1016/0092-8674(86)90480-0 3000622

73. Petruk S, Sedkov Y, Riley KM, Hodgson J, Schweisguth F, Hirose S, et al. Transcription of bxd noncoding RNAs promoted by trithorax represses Ubx in cis by transcriptional interference. Cell. 2006;127(6):1209–21. doi: 10.1016/j.cell.2006.10.039 17174895.

74. Thurmond J, Goodman JL, Strelets VB, Attrill H, Gramates LS, Marygold SJ, et al. FlyBase 2.0: the next generation. Nucleic Acids Res. 2019;47(D1):D759–D65. doi: 10.1093/nar/gky1003 30364959

75. Chen H, Levo M, Barinov L, Fujioka M, Jaynes JB, Gregor T. Dynamic interplay between enhancer–promoter topology and gene activity. Nat Genet. 2018;50(9):1296. doi: 10.1038/s41588-018-0175-z 30038397

76. Schwartz YB, Kahn TG, Nix DA, Li XY, Bourgon R, Biggin M, et al. Genome-wide analysis of Polycomb targets in Drosophila melanogaster. Nat Genet. 2006;38(6):700–5. doi: 10.1038/ng1817 16732288.

77. Tolhuis B, de Wit E, Muijrers I, Teunissen H, Talhout W, van Steensel B, et al. Genome-wide profiling of PRC1 and PRC2 Polycomb chromatin binding in Drosophila melanogaster. [erratum appears in Nat Genet. 2006 Jul;38(7):850]. Nat Genet. 2006;38(6):694–9. doi: 10.1038/ng1792 16628213.

78. Eaton JD, West S. Termination of Transcription by RNA Polymerase II: BOOM! Trends Genet. 2020. doi: 10.1016/j.tig.2020.05.008 32527618

79. Miki TS, Carl SH, Großhans H. Two distinct transcription termination modes dictated by promoters. Genes Dev. 2017;31(18):1870–9. doi: 10.1101/gad.301093.117 29021241


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