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Stem Cell Migration pp 169–184Cite as

Imaging Pluripotent Cell Migration in Drosophila

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 750))

Abstract

Drosophila melanogaster offers a powerful system for the analysis of cell migration. In the embryo, ­pluripotent cells of the mesodermal and endodermal primordia undergo epithelial–mesenchymal transitions and cell migration, while primordial germ cells migrate through an endodermal barrier to form the gonads. Visualisation of these migrations has traditionally been achieved by staining fixed embryos at different developmental stages or through live imaging of cells using tissue-specific expression of marker fluorescent proteins. More recently, photoactivatable fluorescence proteins have allowed the labelling of small groups of cells or single cells so that their migratory patterns and fate can be followed. By fusing the photoactivatable fluorescent protein to proteins that mark different subcellular components, it is now possible to visualise different aspects of the cells as they migrate. Here, we review previous studies of the migration of pluripotent embryonic cells and describe, in detail, methods for visualising these cells.

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References

  1. Matthews, K.A., Kaufman, T.C., and Gelbart,W.M. (2005) Research resources for Drosophila: the expanding universe Nat Rev Genet 6,179–93.

    Google Scholar 

  2. Venken, K.J., and Bellen, H.J. (2005) Emerging technologies for gene manipulation in Drosophila melanogaster Nat Rev Genet 6, 167–78.

    Google Scholar 

  3. Bate, M., and Martinez Arias, A. (1993) The Development of Drosophila melanogaster Cold Spring Harbor Laboratory Press.

    Google Scholar 

  4. Chalfie, M., Tu, Y., Euskirchen, G., Ward, W.W., and Prasher, D.C. (1994) Green fluorescent protein as a marker for gene expression Science 263, 802–5.

    Google Scholar 

  5. Brand, A.H. (1995) GFP in Drosophila TIG, 324–5.

    Google Scholar 

  6. Mavrakis, M., Rikhy, R., Lilly, M., Lippincott-Schwartz, J. (2008) Fluorescence imaging techniques for studying Drosophila embryo development. Curr Protoc Cell Biol.4.18.1- 43.

    Google Scholar 

  7. McDonald, J.A., and Montell, D.J. (2005) Analysis of cell migration using Drosophila as a model system Methods Mol Biol, 175–202.

    Google Scholar 

  8. Wood, W., and Jacinto, A. (2005) Imaging cell movement during dorsal closure in Drosophila embryos Methods Mol Biol, 203–10.

    Google Scholar 

  9. Ninov, N., and Martín-Blanco, E. (2007) Live imaging of epidermal morphogenesis during the development of the adult abdominal epidermis of Drosophila Nature protocols 2, 3074–80.

    Google Scholar 

  10. Prasad, M., and Montell, D.J. (2007) Cellular and molecular mechanisms of border cell migration analyzed using time-lapse live-cell imaging Dev Cell 12, 997–1005.

    Google Scholar 

  11. Oda, H., and Tsukita, S. (2001) Real-time imaging of cell-cell adherens junctions reveals that Drosophila mesoderm invagination begins with two phases of apical constriction of cells J Cell Sci 114(Pt 3), 493–501.

    Google Scholar 

  12. Brand, A.H., and Perrimon, N. (1993) Targeted gene expression as a means of ­altering cell fates and generating dominant phenotypes Development 118, 401–15.

    Google Scholar 

  13. Duffy, J.B. (2002) GAL4 system in Drosophila: a fly geneticist’s Swiss army knife Genesis 34, 1–15.

    Google Scholar 

  14. Lin, H., and Schagat, T. (1997) Neuroblasts: a model for the asymmetric division of stem cells Trends Genet 13, 33–9.

    Google Scholar 

  15. Spradling, A., Drummond-Barbosa, D., and Kai, T. (2001) Stem cells find their niche Nature 114, 98–104.

    Google Scholar 

  16. Mandal, L., Martinez-Agosto, J.A., Evans, C.J., Hartenstein, V., and Banerjee, U. (2007) A Hedgehog- and Antennapedia-dependent niche maintains Drosophila haematopoietic precursors Nature 446, 320–4.

    Google Scholar 

  17. Micchelli, C., and Perrimon, N. (2006) Evidence that stem cells reside in the adult Drosophila midgut epithelium Nature 439, 475–9.

    Google Scholar 

  18. Ohlstein, B., and Spradling, A. (2006) The adult Drosophila posterior midgut is maintained by pluripotent stem cells Nature 439,470–4.

    Google Scholar 

  19. Takashima, S., Mkrtchyan, M., Younossi-Hartenstein, A., Merriam, J.R., and Hartenstein, V. (2008) The behaviour of Drosophila adult hindgut stem cells is controlled by Wnt and Hh signalling Nature 454, 651–5.

    Google Scholar 

  20. Singh, S.R., Liu, W., and Hou, S.X. (2007) The adult Drosophila malpighian tubules are maintained by multipotent stem cells Cell Stem Cell 1, 191–203.

    Google Scholar 

  21. Kaltschmidt, J.A., Davidson, C.M., Brown, N.H., and Brand, A.H. (2000) Rotation and asymmetry of the mitotic spindle direct asymmetric cell division in the developing central nervous system Nat Cell Biol 2, 7–12.

    Google Scholar 

  22. Cheng, J., Türkel, N., Hemati, N., Fuller, M., Hunt, A., and Yamashita, Y. (2008) Centrosome misorientation reduces stem cell division during ageing Nature 456, 599–604.

    Google Scholar 

  23. Santos, A., and Lehmann, R. (2004) Germ Cell Specification and Migration in Drosophila and beyond Current Biology, R578–R89.

    Google Scholar 

  24. Kunwar, P.S., Starz-Gaiano, M., Bainton, R.J., Heberlein, U., and Lehmann, R. (2003) Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells PLoS Biol 1, E80.

    Google Scholar 

  25. Kunwar, P., Sano, H., Renault, A., Barbosa, V., Fuse, N., and Lehmann, R. (2008) Tre1 GPCR initiates germ cell transepithelial migration by regulating Drosophila melanogaster E-cadherin The Journal of Cell Biology 183, 157–68.

    Google Scholar 

  26. Sano, H., Renault, A.D., and Lehmann, R. (2005) Control of lateral migration and germ cell elimination by the Drosophila melanogaster lipid phosphate phosphatases Wunen and Wunen 2 The Journal of Cell Biology 171, 675–83.

    Google Scholar 

  27. Wilson, R., and Leptin, M. (2000) Fibroblast growth factor receptor-dependent morphogenesis of the Drosophila mesoderm Philos Trans R Soc Lond B Biol Sci 355, 891–5.

    Google Scholar 

  28. Stathopoulos, A., Tam, B., Ronshaugen, M., Frasch, M., and Levine, M. (2004) Pyramus and thisbe: FGF genes that pattern the mesoderm of Drosophila embryos Genes Dev 18, 687–99.

    Google Scholar 

  29. Gryzik, T., and Muller, H.A. (2004) FGF8-like1 and FGF8-like2 encode putative ligands of the FGF receptor Htl and are required for mesoderm migration in the Drosophila gastrula Curr Biol 14, 659–67.

    Google Scholar 

  30. Gisselbrecht, S., Skeath, J.B., Doe, C.Q., and Michelson, A.M. (1996) Heartless encodes a fibroblast growth factor receptor (DFR1/DFGF-R2) involved in the directional migration of early mesodermal cells in the Drosophila embryo Genes Dev 10, 3003–17.

    Google Scholar 

  31. Beiman, M., Shilo, B.Z., and Volk, T. (1996) Heartless, a Drosophila FGF receptor homolog, is essential for cell migration and establishment of several mesodermal lineages Genes Dev 10, 2993–3002.

    Google Scholar 

  32. van Impel, A., Schumacher, S., Draga, M., Herz, H., Goshans, J., and Muller, H. (2009) Regulation of the Rac GTPase pathway by the multi- functional Rho GEF Pebble is essential for mesoderm migration in the Drosophila gastrula Development 136, 813–22.

    Google Scholar 

  33. Smallhorn, M., Murray, M.J., and Saint, R. (2004) The epithelial-mesenchymal transition of the Drosophila mesoderm requires the Rho GTP exchange factor Pebble Development 131, 2641–51.

    Google Scholar 

  34. Schumacher, S., Gryzik, T., Tannebaum, S., and Muller, H.A. (2004) The RhoGEF Pebble is required for cell shape changes during cell migration triggered by the Drosophila FGF receptor Heartless Development 131, 2631–40.

    Google Scholar 

  35. Murray, M.J., and Saint, R. (2007) Photoactivatable GFP resolves Drosophila mesoderm migration behaviour Development 134, 3975–83.

    Google Scholar 

  36. Mcmahon, A., Supatto, W., Fraser, S., and Stathopoulos, A. (2008) Dynamic Analyses of Drosophila Gastrulation Provide Insights into Collective Cell Migration Science 322, 1546–50.

    Google Scholar 

  37. Reuter, R., Grunewald, B., and Leptin, M. (1993) A role for the mesoderm in endodermal migration and morphogenesis in Drosophila Development 11, 1135–45.

    Google Scholar 

  38. Martin-Bermudo, M.D., Alvarez-Garcia, I., and Brown, N.H. (1999) Migration of the Drosophila primordial midgut cells requires coordination of diverse PS integrin functions Development 126, 5161–9.

    Google Scholar 

  39. Devenport, D., and Brown, N.H. (2004) Morphogenesis in the absence of integrins: mutation of both Drosophila beta subunits prevents midgut migration Development 131, 5405–15.

    Google Scholar 

  40. Lukyanov, K.A., Chudakov, D.M., Lukyanov, S., and Verkhusha, V.V. (2005) Innovation: Photoactivatable fluorescent proteins Nat Rev Mol Cell Biol 6, 885–91.

    Google Scholar 

  41. Patterson, G.H., and Lippincott-Schwartz, J. (2002) A photoactivatable GFP for selective photolabeling of proteins and cells Science 297, 1873–7.

    Google Scholar 

  42. Patterson, G.H., and Lippincott-Schwartz, J. (2002) Selective photolabeling of proteins using photoactivatable GFP Methods, 445–50.

    Google Scholar 

  43. Pantazis, P., and Gonzalez-Gaitan, M. (2007) Localized multiphoton photoactivation of paGFP in Drosophila wing imaginal discs J Biomed Opt 12, 044004.

    Google Scholar 

  44. Post, J.N., Lidke, K.A., Rieger, B., and Arndt-Jovin, D.J. (2005) One- and two-photon photoactivation of a paGFP-fusion protein in live Drosophila embryos FEBS Lett 579, 325–30.

    Google Scholar 

  45. Dutta, D., Bloor, J.W., Ruiz-Gomez ,M., VijayRaghavan, K., and Kiehart, D.P. (2002) Real-time imaging of morphogenetic movements in Drosophila using Gal4-UAS-driven expression of GFP fused to the actin-binding domain of moesin Genesis 34, 146–51.

    Google Scholar 

  46. Greenspan, R.J. (1997) Fly Pushing: The Theory and Practice of Drosophila genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press.

    Google Scholar 

  47. Campos-Ortega, J.A., and Hartenstein, V. (1997) The Embryonic Development of Drosophila melanogaster 2nd ed. Berlin: Springer-Verlag.

    Google Scholar 

  48. Pawley, J.B. (2006) Handbook of Biological Confocal Microscopy 3 ed: Springer.

    Google Scholar 

  49. Collins, T.J. (2007) ImageJ for microscopy Biotechniques 43(1 Suppl), 25–30.

    Google Scholar 

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Acknowledgments

We thank Michael Zavortink and Ursula Wiedemann for ­construction of the PAGFP-α-Tub84B and PAGFP-MoeABD ­constructs, respectively and Maria Leptin for the gift of the ­anti-Twist antibody. The PAGFP vectors were kindly provided by George Patterson. This work was supported by an NHMRC ­project grant to M.J.M. and R.S., the ARC Special Research Centre for the Molecular Genetics of Development, and the Institute of Advanced Studies at The Australian National University.

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Authors and Affiliations

  1. Molecular Genetics and Evolution, Research School of Biological Sciences, Australian National University, Acton, ACT, Australia

    Robert Saint

Authors
  1. Michael J. Murray
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  2. Robert Saint
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Correspondence to Robert Saint .

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Editors and Affiliations

  1. Medical Center, Division of Experimental Hematology, Cincinnati Children's Hospital, Burnet Ave. 3333, Cincinnati, 45229, Ohio, USA

    Marie-Dominique Filippi

  2. Medical Center, Division of Experimental Hematology, Cincinnati Children's Hospital, Burnet Avenue 3333, Cincinnati, 45229, Ohio, USA

    Hartmut Geiger

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Murray, M.J., Saint, R. (2011). Imaging Pluripotent Cell Migration in Drosophila . In: Filippi, MD., Geiger, H. (eds) Stem Cell Migration. Methods in Molecular Biology, vol 750. Humana Press. https://doi.org/10.1007/978-1-61779-145-1_12

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  • DOI: https://doi.org/10.1007/978-1-61779-145-1_12

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-144-4

  • Online ISBN: 978-1-61779-145-1

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