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Gene Regulation of BMP Ligands in Drosophila

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Bone Morphogenetic Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1891))

Abstract

Drosophila is a valuable system to study bone morphogenetic proteins (BMPs) due to the high functional conservation of the pathway and the molecular genetic tools available. Drosophila has three BMP ligands, decapentaplegic (BMP2/4), screw, and glass bottom boat (BMP5/6/7/8). Of these genes, the transcriptional regulation of decapentaplegic has been studied, and some of the enhancers directing its spatially specific gene expression have been described. These analyses have used many of the standard tools of molecular biology, but a valuable method of analysis often used in Drosophila is the creation of patches of mutant tissue at any stage and in any location by induced somatic recombination. The ability to create transgenic flies and manipulate the Drosophila genome with recombinases is well established. This method can be used to evaluate the requirements for specific transcription factors to act on enhancer elements in vivo, in stage- and tissue-specific manners. The yeast FLP/FRT recombination system facilitates experiments to interrogate loss- or gain-of-function for transcription factor activity on known enhancers. This chapter will outline the necessary steps to create the tools needed and conduct somatic cell recombination experiments to interrogate the function of transcription factors on BMP enhancers.

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References

  1. Arora K, Levine MS, O’Connor MB (1994) The screw gene encodes a ubiquitously expressed member of the TGF-beta family required for specification of dorsal cell fates in the Drosophila embryo. Genes Dev 8(21):2588–2601

    Article  CAS  Google Scholar 

  2. Bivik C, Bahrampour S, Ulvklo C, Nilsson P, Angel A, Fransson F, Lundin E, Renhorn J, Thor S (2015) Novel genes involved in controlling specification of Drosophila FMRFamide neuropeptide cells. Genetics 200(4):1229–1244. https://doi.org/10.1534/genetics.115.178483

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. McCabe BD, Marques G, Haghighi AP, Fetter RD, Crotty ML, Haerry TE, Goodman CS, O’Connor MB (2003) The BMP homolog Gbb provides a retrograde signal that regulates synaptic growth at the Drosophila neuromuscular junction. Neuron 39(2):241–254

    Article  CAS  Google Scholar 

  4. Khalsa O, Yoon JW, Torres-Schumann S, Wharton KA (1998) TGF-beta/BMP superfamily members, Gbb-60A and Dpp, cooperate to provide pattern information and establish cell identity in the Drosophila wing. Development 125(14):2723–2734

    CAS  PubMed  Google Scholar 

  5. Wharton KA, Cook JM, Torres-Schumann S, de Castro K, Borod E, Phillips DA (1999) Genetic analysis of the bone morphogenetic protein-related gene, gbb, identifies multiple requirements during Drosophila development. Genetics 152(2):629–640

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Ballard SL, Jarolimova J, Wharton KA (2010) Gbb/BMP signaling is required to maintain energy homeostasis in Drosophila. Dev Biol 337(2):375–385. https://doi.org/10.1016/j.ydbio.2009.11.011

    Article  CAS  PubMed  Google Scholar 

  7. Kawase E, Wong MD, Ding BC, Xie T (2004) Gbb/Bmp signaling is essential for maintaining germline stem cells and for repressing bam transcription in the Drosophila testis. Development 131(6):1365–1375. https://doi.org/10.1242/dev.01025

    Article  CAS  PubMed  Google Scholar 

  8. Tian A, Jiang J (2014) Intestinal epithelium-derived BMP controls stem cell self-renewal in Drosophila adult midgut. Elife 3:e01857. https://doi.org/10.7554/eLife.01857

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Twombly V, Blackman RK, Jin H, Graff JM, Padgett RW, Gelbart WM (1996) The TGF-beta signaling pathway is essential for Drosophila oogenesis. Development 122(5):1555–1565

    CAS  PubMed  Google Scholar 

  10. Xie T, Spradling AC (1998) Decapentaplegic is essential for the maintenance and division of germline stem cells in the Drosophila ovary. Cell 94(2):251–260

    Article  CAS  Google Scholar 

  11. Irish VF, Gelbart WM (1987) The decapentaplegic gene is required for dorsal-ventral patterning of the Drosophila embryo. Genes Dev 1(8):868–879

    Article  CAS  Google Scholar 

  12. Staehling-Hampton K, Hoffmann FM, Baylies MK, Rushton E, Bate M (1994) dpp induces mesodermal gene expression in Drosophila. Nature 372(6508):783–786

    Article  CAS  Google Scholar 

  13. Frasch M (1995) Induction of visceral and cardiac mesoderm by ectodermal Dpp in the early Drosophila embryo. Nature 374(6521):464–467

    Article  CAS  Google Scholar 

  14. Panganiban GE, Reuter R, Scott MP, Hoffmann FM (1990) A Drosophila growth factor homolog, decapentaplegic, regulates homeotic gene expression within and across germ layers during midgut morphogenesis. Development 110(4):1041–1050

    CAS  PubMed  Google Scholar 

  15. Hursh DA, Padgett RW, Gelbart WM (1993) Cross regulation of decapentaplegic and Ultrabithorax transcription in the embryonic visceral mesoderm of Drosophila. Development 117(4):1211–1222

    CAS  PubMed  Google Scholar 

  16. Vincent S, Ruberte E, Grieder NC, Chen CK, Haerry T, Schuh R, Affolter M (1997) DPP controls tracheal cell migration along the dorsoventral body axis of the Drosophila embryo. Development 124(14):2741–2750

    CAS  PubMed  Google Scholar 

  17. Spencer FA, Hoffmann FM, Gelbart WM (1982) Decapentaplegic: a gene complex affecting morphogenesis in Drosophila melanogaster. Cell 28(3):451–461

    Article  CAS  Google Scholar 

  18. Stultz BG, Lee HJ, Ramon K, Hursh DA (2006) Decapentaplegic head capsule mutations disrupt novel peripodial expression controlling the morphogenesis of the Drosophila ventral head. Dev Biol 296(2):329–339

    Article  CAS  Google Scholar 

  19. Pennetier D, Oyallon J, Morin-Poulard I, Dejean S, Vincent A, Crozatier M (2012) Size control of the Drosophila hematopoietic niche by bone morphogenetic protein signaling reveals parallels with mammals. Proc Natl Acad Sci U S A 109(9):3389–3394. https://doi.org/10.1073/pnas.1109407109

    Article  PubMed  PubMed Central  Google Scholar 

  20. Wharton KA, Thomsen GH, Gelbart WM (1991) Drosophila 60A gene, another transforming growth factor beta family member, is closely related to human bone morphogenetic proteins. Proc Natl Acad Sci U S A 88(20):9214–9218

    Article  CAS  Google Scholar 

  21. St Johnston RD, Hoffmann FM, Blackman RK, Segal D, Grimaila R, Padgett RW, Irick HA, Gelbart WM (1990) Molecular organization of the decapentaplegic gene in Drosophila melanogaster. Genes Dev 4(7):1114–1127

    Article  CAS  Google Scholar 

  22. Huang JD, Schwyter DH, Shirokawa JM, Courey AJ (1993) The interplay between multiple enhancer and silencer elements defines the pattern of decapentaplegic expression. Genes Dev 7(4):694–704

    Article  CAS  Google Scholar 

  23. Zaffran S, Kuchler A, Lee HH, Frasch M (2001) biniou (FoxF), a central component in a regulatory network controlling visceral mesoderm development and midgut morphogenesis in Drosophila. Genes Dev 15(21):2900–2915

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Schaub C, Frasch M (2013) Org-1 is required for the diversification of circular visceral muscle founder cells and normal midgut morphogenesis. Dev Biol 376(2):245–259. https://doi.org/10.1016/j.ydbio.2013.01.022

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Sun B, Hursh DA, Jackson D, Beachy PA (1995) Ultrabithorax protein is necessary but not sufficient for full activation of decapentaplegic expression in the visceral mesoderm. EMBO J 14(3):520–535

    Article  CAS  Google Scholar 

  26. Stultz BG, Jackson DG, Mortin MA, Yang X, Beachy PA, Hursh DA (2006) Transcriptional activation by extradenticle in the Drosophila visceral mesoderm. Dev Biol 290(2):482–494

    Article  CAS  Google Scholar 

  27. Capovilla M, Brandt M, Botas J (1994) Direct regulation of decapentaplegic by Ultrabithorax and its role in Drosophila midgut morphogenesis. Cell 76(3):461–475

    Article  CAS  Google Scholar 

  28. Manak JR, Mathies LD, Scott MP (1994) Regulation of a decapentaplegic midgut enhancer by homeotic proteins. Development 120(12):3605–3619

    CAS  PubMed  Google Scholar 

  29. Capovilla M, Botas J (1998) Functional dominance among Hox genes: repression dominates activation in the regulation of Dpp. Development 125(24):4949–4957

    CAS  PubMed  Google Scholar 

  30. Yang X, van Beest M, Clevers H, Jones T, Hursh DA, Mortin MA (2000) Decapentaplegic is a direct target of dTcf repression in the Drosophila visceral mesoderm. Development 127(17):3695–3702

    CAS  PubMed  Google Scholar 

  31. Johnson AN, Bergman CM, Kreitman M, Newfeld SJ (2003) Embryonic enhancers in the dpp disk region regulate a second round of Dpp signaling from the dorsal ectoderm to the mesoderm that represses Zfh-1 expression in a subset of pericardial cells. Dev Biol 262(1):137–151

    Article  CAS  Google Scholar 

  32. Takaesu NT, Bulanin DS, Johnson AN, Orenic TV, Newfeld SJ (2008) A combinatorial enhancer recognized by Mad, TCF and Brinker first activates then represses dpp expression in the posterior spiracles of Drosophila. Dev Biol 313(2):829–843. https://doi.org/10.1016/j.ydbio.2007.10.021

    Article  CAS  PubMed  Google Scholar 

  33. Hepker J, Blackman RK, Holmgren R (1999) Cubitus interruptus is necessary but not sufficient for direct activation of a wing-specific decapentaplegic enhancer. Development 126(16):3669–3677

    CAS  PubMed  Google Scholar 

  34. Muller B, Basler K (2000) The repressor and activator forms of Cubitus interruptus control Hedgehog target genes through common generic gli-binding sites. Development 127(14):2999–3007

    CAS  PubMed  Google Scholar 

  35. Stultz BG, Park SY, Mortin MA, Kennison JA, Hursh DA (2012) Hox proteins coordinate peripodial decapentaplegic expression to direct adult head morphogenesis in Drosophila. Dev Biol 369(2):362–376

    Article  CAS  Google Scholar 

  36. Lee H, Stultz BG, Hursh DA (2007) The Zic family member, odd-paired, regulates the Drosophila BMP, decapentaplegic, during adult head development. Development 134(7):1301–1310

    Article  CAS  Google Scholar 

  37. Biehs B, Francois V, Bier E (1996) The Drosophila short gastrulation gene prevents Dpp from autoactivating and suppressing neurogenesis in the neuroectoderm. Genes Dev 10(22):2922–2934

    Article  CAS  Google Scholar 

  38. Chanut F, Heberlein U (1997) Role of decapentaplegic in initiation and progression of the morphogenetic furrow in the developing Drosophila retina. Development 124(2):559–567

    CAS  PubMed  Google Scholar 

  39. Pignoni F, Zipursky SL (1997) Induction of Drosophila eye development by decapentaplegic. Development 124(2):271–278

    CAS  PubMed  Google Scholar 

  40. Wiersdorff V, Lecuit T, Cohen SM, Mlodzik M (1996) Mad acts downstream of Dpp receptors, revealing a differential requirement for dpp signaling in initiation and propagation of morphogenesis in the Drosophila eye. Development 122(7):2153–2162

    CAS  PubMed  Google Scholar 

  41. Chou TB, Perrimon N (1992) Use of a yeast site-specific recombinase to produce female germline chimeras in Drosophila. Genetics 131(3):643–653

    CAS  PubMed  PubMed Central  Google Scholar 

  42. Golic KG (1991) Site-specific recombination between homologous chromosomes in Drosophila. Science 252(5008):958–961

    Article  CAS  Google Scholar 

  43. Xu T, Rubin GM (1993) Analysis of genetic mosaics in developing and adult Drosophila tissues. Development 117:1223–1237

    CAS  PubMed  Google Scholar 

  44. Siegal ML, Hartl DL (1996) Transgene Coplacement and high efficiency site-specific recombination with the Cre/loxP system in Drosophila. Genetics 144(2):715–726

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Groth AC, Fish M, Nusse R, Calos MP (2004) Construction of transgenic Drosophila by using the site-specific integrase from phage phiC31. Genetics 166(4):1775–1782

    Article  CAS  Google Scholar 

  46. Blair SS (2003) Genetic mosaic techniques for studying Drosophila development. Development 130(21):5065–5072. https://doi.org/10.1242/dev.00774

    Article  CAS  PubMed  Google Scholar 

  47. de Navascues J, Modolell J (2010) The pronotum LIM-HD gene tailup is both a positive and a negative regulator of the proneural genes achaete and scute of Drosophila. Mech Dev 127(9–12):393–406. https://doi.org/10.1016/j.mod.2010.05.001

    Article  CAS  PubMed  Google Scholar 

  48. Greenspan RJ (2004) Fly pushing: the theory and practice of Drosophila genetics, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

    Google Scholar 

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Acknowledgments

We thank Drs. John Thomas, Brent McCright, and Mark Mortin for helpful comments that improved the manuscript.

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Authors and Affiliations

  1. Division of Cell and Gene Therapy, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD, USA

    Brian G. Stultz & Deborah A. Hursh

Authors
  1. Brian G. Stultz
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  2. Deborah A. Hursh
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Corresponding author

Correspondence to Deborah A. Hursh .

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Editors and Affiliations

  1. New Jersey Medical School, Rutgers Biomedical and Health Sciences, Newark, NJ, USA

    Melissa B. Rogers

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Stultz, B.G., Hursh, D.A. (2019). Gene Regulation of BMP Ligands in Drosophila. In: Rogers, M. (eds) Bone Morphogenetic Proteins. Methods in Molecular Biology, vol 1891. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8904-1_7

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  • DOI: https://doi.org/10.1007/978-1-4939-8904-1_7

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8903-4

  • Online ISBN: 978-1-4939-8904-1

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