Skip to main content
SpringerLink
Log in

A genetic screen supports a broad role for the Drosophila insulator proteins BEAF-32A and BEAF-32B in maintaining patterns of gene expression

  • Original Paper
  • Published:
Molecular Genetics and Genomics Aims and scope Submit manuscript

Abstract

Chromatin domain insulators are thought to insulate adjacent genes, including their regulatory elements, from each other by organizing chromatin into functionally independent domains. Thus insulators should play a global role in gene regulation by keeping regulatory domains separated. However, this has never been demonstrated. We previously designed and characterized a transgene that is under GAL4 UAS control and encodes a dominant-negative form of the Boundary Element-Associated Factors BEAF-32A and BEAF-32B. The BID transgene encodes the BEAF self-interaction domain but lacks a DNA binding domain. Expression of BID in eye imaginal discs leads to a rough eye phenotype. Here we screen for dominant mutations that modify this eye phenotype. This assay provides evidence for cross-talk between different classes of insulators, and for a broad role of the BEAF proteins in maintaining patterns of gene expression during eye development. Most identified genes encode other insulator binding proteins, transcription factors involved in head development, or general transcription factors. Because it is unlikely that insulator function is limited to eye development, the present results support the hypothesis that insulators play a widespread role in maintaining global transcription programs.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Armstrong JA, Sperling AS, Deuring R, Manning L, Moseley SL, Papoulas O, Piatek CI, Doe CQ, Tamkun JW (2005) Genetic screens for enhancers of brahma reveal functional interactions between the BRM chromatin-remodeling complex and the delta-notch signal transduction pathway in Drosophila. Genetics 170:1761–1774

    Article  CAS  Google Scholar 

  • Aulner N, Monod C, Mandicourt G, Jullien D, Cuvier O, Sall A, Janssen S, Laemmli UK, Kas E (2002) The AT-hook protein D1 is essential for Drosophila melanogaster development and is implicated in position-effect variegation. Mol Cell Biol 22:1218–1232

    Article  CAS  Google Scholar 

  • Blanton J, Gaszner M, Schedl P (2003) Protein:protein interactions and the pairing of boundary elements in vivo. Genes Dev 17:664–675

    Article  CAS  Google Scholar 

  • Brand AH, Perrimon N (1993) Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. Development 118:401–415

    CAS  Google Scholar 

  • Cai H, Levine M (1995) Modulation of enhancer–promoter interactions by insulators in the Drosophila embryo. Nature 376:533–536

    Article  CAS  Google Scholar 

  • Campuzano S (2001) Emc, a negative HLH regulator with multiple functions in Drosophila development. Oncogene 20:8299–8307

    Article  CAS  Google Scholar 

  • Capelson M, Corces VG (2004) Boundary elements and nuclear organization. Biol Cell 96:617–629

    Article  CAS  Google Scholar 

  • Capelson M, Corces VG (2005) The ubiquitin ligase dTopors directs the nuclear organization of a chromatin insulator. Mol Cell 20:105–116

    Article  CAS  Google Scholar 

  • Cuvier O, Hart CM, Laemmli UK (1998) Identification of a class of chromatin boundary elements. Mol Cell Biol 18:7478–7486

    CAS  Google Scholar 

  • Cuvier O, Hart CM, Kas E, Laemmli UK (2002) Identification of a multicopy chromatin boundary element at the borders of silenced chromosomal domains. Chromosoma 110:519–531

    CAS  Google Scholar 

  • Denell R (1994) Discovery and genetic definition of the Drosophila Antennapedia complex. Genetics 138:549–552

    CAS  Google Scholar 

  • Diederich RJ, Merrill VK, Pultz MA, Kaufman TC (1989) Isolation, structure, and expression of labial, a homeotic gene of the Antennapedia complex involved in Drosophila head development. Genes Dev 3:399–414

    CAS  Google Scholar 

  • Dong PD, Chu J, Panganiban G (2001) Proximodistal domain specification and interactions in developing Drosophila appendages. Development 128:2365–2372

    CAS  Google Scholar 

  • Gaszner M, Vazquez J, Schedl P (1999) The Zw5 protein, a component of the scs chromatin domain boundary, is able to block enhancer–promoter interaction. Genes Dev 13:2098–2107

    CAS  Google Scholar 

  • Gause M, Eissenberg JC, Macrae AF, Dorsett M, Misulovin Z, Dorsett D (2006) Nipped-A, the Tra1/TRRAP subunit of the Drosophila SAGA and Tip60 complexes, has multiple roles in Notch signaling during wing development. Mol Cell Biol 26:2347–2359

    Article  CAS  Google Scholar 

  • Gerasimova TI, Gdula DA, Gerasimov DV, Simonova O, Corces VG (1995) A Drosophila protein that imparts directionality on a chromatin insulator is an enhancer of position-effect variegation. Cell 82:587–597

    Article  CAS  Google Scholar 

  • Geyer PK, Clark I (2002) Protecting against promiscuity: the regulatory role of insulators. Cell Mol Life Sci 59:2112–2127

    Article  CAS  Google Scholar 

  • Geyer PK, Corces VG (1992) DNA position-specific repression of transcription by a Drosophila zinc finger protein. Genes Dev 6:1865–1873

    CAS  Google Scholar 

  • Gilbert MK, Tan YY, Hart CM (2006) The Drosophila boundary element-associated factors BEAF-32A and BEAF-32B affect chromatin structure. Genetics 173:1365–1375

    Article  CAS  Google Scholar 

  • Giot L, Bader JS, Brouwer C, Chaudhuri A, Kuang B, Li Y, Hao YL, Ooi CE, Godwin B, Vitols E, Vijayadamodar G, Pochart P, Machineni H, Welsh M, Kong Y, Zerhusen B, Malcolm R, Varrone Z, Collis A, Minto M, Burgess S, McDaniel L, Stimpson E, Spriggs F, Williams J, Neurath K, Ioime N, Agee M, Voss E, Furtak K, Renzulli R, Aanensen N, Carrolla S, Bickelhaupt E, Lazovatsky Y, DaSilva A, Zhong J, Stanyon CA, Finley RL Jr, White KP, Braverman M, Jarvie T, Gold S, Leach M, Knight J, Shimkets RA, McKenna MP, Chant J, Rothberg JM (2003) A protein interaction map of Drosophila melanogaster. Science 302:1727–1736

    Article  CAS  Google Scholar 

  • Greenberg AJ, Yanowitz JL, Schedl P (2004) The Drosophila GAGA factor is required for dosage compensation in males and for the formation of the male-specific-lethal complex chromatin entry site at 12DE. Genetics 166:279–289

    Article  CAS  Google Scholar 

  • Han Z, Li X, Wu J, Olson EN (2004) A myocardin-related transcription factor regulates activity of serum response factor in Drosophila. Proc Natl Acad Sci USA 101:12567–12572

    Article  CAS  Google Scholar 

  • Hart CM, Zhao K, Laemmli UK (1997) The scs’ boundary element: characterization of boundary element-associated factors. Mol Cell Biol 17:999–1009

    CAS  Google Scholar 

  • Hart CM, Cuvier O, Laemmli UK (1999) Evidence for an antagonistic relationship between the boundary element-associated factor BEAF and the transcription factor DREF. Chromosoma 108:375–383

    Article  CAS  Google Scholar 

  • Janody F, Martirosyan Z, Benlali A, Treisman JE (2003) Two subunits of the Drosophila mediator complex act together to control cell affinity. Development 130:3691–3701

    Article  CAS  Google Scholar 

  • Kellum R, Schedl P (1991) A position-effect assay for boundaries of higher order chromosomal domains. Cell 64:941–950

    Article  CAS  Google Scholar 

  • Kennerdell JR, Yamaguchi S, Carthew RW (2002) RNAi is activated during Drosophila oocyte maturation in a manner dependent on aubergine and spindle-E. Genes Dev 16:1884–1889

    Article  CAS  Google Scholar 

  • Kermekchiev M, Pettersson M, Matthias P, Schaffner W (1991) Every enhancer works with every promoter for all the combinations tested: could new regulatory pathways evolve by enhancer shuffling? Gene Expr 1:71–81

    CAS  Google Scholar 

  • Kuhn EJ, Geyer PK (2003) Genomic insulators: connecting properties to mechanism. Curr Opin Cell Biol 15:259–265

    Article  CAS  Google Scholar 

  • Labrador M, Corces VG (2002) Setting the boundaries of chromatin domains and nuclear organization. Cell 111:151–154

    Article  CAS  Google Scholar 

  • Lee TI, Causton HC, Holstege FC, Shen WC, Hannett N, Jennings EG, Winston F, Green MR, Young RA (2000) Redundant roles for the TFIID and SAGA complexes in global transcription. Nature 405:701–704

    Article  CAS  Google Scholar 

  • Lei EP, Corces VG (2006) RNA interference machinery influences the nuclear organization of a chromatin insulator. Nat Genet 38:936–941

    Article  CAS  Google Scholar 

  • Lewis EB (1998) The bithorax complex: the first fifty years. Int J Dev Biol 42:403–415

    CAS  Google Scholar 

  • Litt MD, Simpson M, Gaszner M, Allis CD, Felsenfeld G (2001) Correlation between histone lysine methylation and developmental changes at the chicken beta-globin locus. Science 293:2453–2455

    Article  CAS  Google Scholar 

  • Maeda RK, Karch F (2006) The ABC of the BX-C: the bithorax complex explained. Development 133:1413–1422

    Article  CAS  Google Scholar 

  • Mukhopadhyay R, Yu W, Whitehead J, Xu J, Lezcano M, Pack S, Kanduri C, Kanduri M, Ginjala V, Vostrov A, Quitschke W, Chernukhin I, Klenova E, Lobanenkov V, Ohlsson R (2004) The binding sites for the chromatin insulator protein CTCF map to DNA methylation-free domains genome-wide. Genome Res 14:1594–1602

    Article  CAS  Google Scholar 

  • Mulholland NM, King IF, Kingston RE (2003) Regulation of Polycomb group complexes by the sequence-specific DNA binding proteins Zeste and GAGA. Genes Dev 17:2741–2746

    Article  CAS  Google Scholar 

  • Noma K, Allis CD, Grewal SI (2001) Transitions in distinct histone H3 methylation patterns at the heterochromatin domain boundaries. Science 293:1150–1155

    Article  CAS  Google Scholar 

  • Norton JD, Deed RW, Craggs G, Sablitzky F (1998) Id helix-loop-helix proteins in cell growth and differentiation. Trends Cell Biol 8:58–65

    Article  CAS  Google Scholar 

  • Ohtsuki S, Levine M (1998) GAGA mediates the enhancer blocking activity of the eve promoter in the Drosophila embryo. Genes Dev 12:3325–3330

    CAS  Google Scholar 

  • Pal-Bhadra M, Leibovitch BA, Gandhi SG, Rao M, Bhadra U, Birchler JA, Elgin SC (2004) Heterochromatic silencing and HP1 localization in Drosophila are dependent on the RNAi machinery. Science 303:669–672

    Article  CAS  Google Scholar 

  • Prioleau MN, Nony P, Simpson M, Felsenfeld G (1999) An insulator element and condensed chromatin region separate the chicken beta-globin locus from an independently regulated erythroid-specific folate receptor gene. EMBO J 18:4035–4048

    Article  CAS  Google Scholar 

  • Read D, Butte MJ, Dernburg AF, Frasch M, Kornberg TB (2000) Functional studies of the BTB domain in the Drosophila GAGA and Mod(mdg4) proteins. Nucleic Acids Res 28:3864–3870

    Article  CAS  Google Scholar 

  • Recillas-Targa F, Pikaart MJ, Burgess-Beusse B, Bell AC, Litt MD, West AG, Gaszner M, Felsenfeld G (2002) Position-effect protection and enhancer blocking by the chicken beta-globin insulator are separable activities. Proc Natl Acad Sci USA 99:6883–6888

    Article  CAS  Google Scholar 

  • Rodriguez Alfageme C, Rudkin GT, Cohen LH (1980) Isolation, properties and cellular distribution of D1, a chromosomal protein of Drosophila. Chromosoma 78:1–31

    Article  CAS  Google Scholar 

  • Schwartz CJ, Sampson HM, Hlousek D, Percival-Smith A, Copeland JW, Simmonds AJ, Krause HM (2001) FTZ-Factor1 and Fushi tarazu interact via conserved nuclear receptor and coactivator motifs. EMBO J 20:510–519

    Article  CAS  Google Scholar 

  • Scott KS, Geyer PK (1995) Effects of the su(Hw) insulator protein on the expression of the divergently transcribed Drosophila yolk protein genes. EMBO J 14:6258–6267

    CAS  Google Scholar 

  • Thomas BJ, Wassarman DA (1999) A fly’s eye view of biology. Trends Genet 15:184–190

    Article  CAS  Google Scholar 

  • Tsukiyama T, Becker PB, Wu C (1994) ATP-dependent nucleosome disruption at a heat-shock promoter mediated by binding of GAGA transcription factor. Nature 367:525–532

    Article  CAS  Google Scholar 

  • West AG, Gaszner M, Felsenfeld G (2002) Insulators: many functions, many mechanisms. Genes Dev 16:271–288

    Article  Google Scholar 

  • Yamaguchi M, Yoshida H, Hirose F, Inoue YH, Hayashi Y, Yamagishi M, Nishi Y, Tamai K, Sakaguchi K, Matsukage A (2001) Ectopic expression of BEAF32A in the Drosophila eye imaginal disc inhibits differentiation of photoreceptor cells and induces apoptosis. Chromosoma 110:313–321

    CAS  Google Scholar 

  • Zhao K, Hart CM, Laemmli UK (1995) Visualization of chromosomal domains with boundary element-associated factor BEAF-32. Cell 81:879–889

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank Cindy Henk and Ying Xiao of the Socolofsky Microscopy Center for assistance with the SEM; Matt Gilbert and Nan Jiang for comments on the manuscript; and Christal DeCuir-Charbonnet for assistance with the deficiency crosses. MRTF flies were kindly provided by Z. Han and E.N. Olson (University of Texas Southwestern Medical Center) and ftz flies were kindly provided by H.M. Krause (University of Toronto). This work was supported by grants from the Louisiana Board of Regents [LEQSF(2002-04)-RD-A-06 and LEQSF(2004-05)-RD-A-11] and a LSU Faculty Research Grant.

Author information

Authors and Affiliations

  1. Department of Biological Sciences, Louisiana State University, 70803, LA, USA

    Swarnava Roy, Yian Yee Tan & Craig M. Hart

Authors
  1. Swarnava Roy
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yian Yee Tan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Craig M. Hart
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Craig M. Hart.

Additional information

Communicated by G. Reuter.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Cite this article

Roy, S., Tan, Y.Y. & Hart, C.M. A genetic screen supports a broad role for the Drosophila insulator proteins BEAF-32A and BEAF-32B in maintaining patterns of gene expression. Mol Genet Genomics 277, 273–286 (2007). https://doi.org/10.1007/s00438-006-0187-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00438-006-0187-8

Keywords

Search

Navigation