Analyzing Planar Cell Polarity During Zebrafish Gastrulation

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 839)

Abstract

Planar cell polarity was first described in invertebrates over 20 years ago and is defined as the polarity of cells (and cell structures) within the plane of a tissue, such as an epithelium. Studies in the last 10 years have identified critical roles for vertebrate homologs of these planar cell polarity proteins during gastrulation cell movements. In zebrafish, the terms convergence and extension are used to describe the collection of morphogenetic movements and cell behaviors that contribute to narrowing and elongation of the embryonic body plan. Disruption of planar cell polarity gene function causes profound defects in convergence and extension creating an embryo that has a shortened anterior–posterior axis and is broadened mediolaterally. The zebrafish gastrula-stage embryo is transparent and amenable to live imaging using both Nomarski/differential interference contrast and fluorescence microscopy. This chapter describes methods to analyze convergence and extension movements at the cellular level and thereby connect embryonic phenotypes with underlying planar cell polarity defects in migrating cells.

Key words

Convergence and extension Ectoderm Gastrulation Nomarski/DIC Mesoderm Planar cell polarity Van gogh-like 2 Wnt Zebrafish 

Notes

Acknowledgments

I thank my colleagues Diane Sepich and Lila Solnica-Krezel for their investment in my training in all methods concerning zebrafish gastrulation. Work in the Jessen lab is supported by grants from the American Cancer Society (RSG-09-281-01 DDC) and National Science Foundation (IOS 0950849).

References

  1. 1.
    Wong, L. L., and Adler, P. N. (1993) Tissue polarity genes of Drosophila regulate the subcellular location for prehair initiation in pupal wing cells, J Cell Biol 123, 209–221.PubMedCrossRefGoogle Scholar
  2. 2.
    Adler, P. N. (2002) Planar signaling and morphogenesis in Drosophila, Dev Cell 2, 525–535.PubMedCrossRefGoogle Scholar
  3. 3.
    Hammerschmidt, M., Pelegri, F., Mullins, M. C., Kane, D. A., Brand, M., van Eeden, F. J., Furutani-Seiki, M., Granato, M., Haffter, P., Heisenberg, C. P., Jiang, Y. J., Kelsh, R. N., Odenthal, J., Warga, R. M., and Nusslein-Volhard, C. (1996) Mutations affecting morphogenesis during gastrulation and tail formation in the zebrafish, Danio rerio, Development 123, 143–151.Google Scholar
  4. 4.
    Solnica-Krezel, L., Stemple, D. L., Mountcastle-Shah, E., Rangini, Z., Neuhauss, S. C., Malicki, J., Schier, A. F., Stainier, D. Y., Zwartkruis, F., Abdelilah, S., and Driever, W. (1996) Mutations affecting cell fates and cellular rearrangements during gastrulation in zebrafish, Development 123, 67–80.PubMedGoogle Scholar
  5. 5.
    Jessen, J. R., Topczewski, J., Bingham, S., Sepich, D. S., Marlow, F., Chandrasekhar, A., and Solnica-Krezel, L. (2002) Zebrafish trilobite identifies new roles for Strabismus in gastrulation and neuronal movements, Nat Cell Biol 4, 610–615.PubMedGoogle Scholar
  6. 6.
    Solnica-Krezel, L., and Cooper, M. S. (2002) Cellular and genetic mechanisms of convergence and extension, Results Probl Cell Differ 40, 136–165.PubMedGoogle Scholar
  7. 7.
    Mlodzik, M. (2002) Planar cell polarization: do the same mechanisms regulate Drosophila tissue polarity and vertebrate gastrulation?, Trends Genet 18, 564–571.PubMedCrossRefGoogle Scholar
  8. 8.
    Concha, M. L., and Adams, R. J. (1998) Oriented cell divisions and cellular morphogenesis in the zebrafish gastrula and neurula: a time-lapse analysis, Development 125, 983–994.PubMedGoogle Scholar
  9. 9.
    Coyle, R. C., Latimer, A., and Jessen, J. R. (2008) Membrane-type 1 matrix metalloproteinase regulates cell migration during zebrafish gastrulation: evidence for an interaction with non-canonical Wnt signaling, Exp Cell Res 314, 2150–2162.PubMedCrossRefGoogle Scholar
  10. 10.
    Lin, F., Sepich, D. S., Chen, S., Topczewski, J., Yin, C., Solnica-Krezel, L., and Hamm, H. (2005) Essential roles of G{alpha}12/13 signaling in distinct cell behaviors driving zebrafish convergence and extension gastrulation movements, J Cell Biol 169, 777–787.PubMedCrossRefGoogle Scholar
  11. 11.
    Myers, D. C., Sepich, D. S., and Solnica-Krezel, L. (2002) Bmp activity gradient regulates convergent extension during zebrafish gastrulation, Dev Biol 243, 81–98.PubMedCrossRefGoogle Scholar
  12. 12.
    Topczewski, J., Sepich, D. S., Myers, D. C., Walker, C., Amores, A., Lele, Z., Hammerschmidt, M., Postlethwait, J., and Solnica-Krezel, L. (2001) The zebrafish glypican knypek controls cell polarity during gastrulation movements of convergent extension, Dev Cell 1, 251–264.PubMedCrossRefGoogle Scholar
  13. 13.
    Sepich, D. S., and Solnica-Krezel, L. (2005) Analysis of cell movements in zebrafish embryos: morphometrics and measuring movement of labeled cell populations in vivo, Methods Mol Biol 294, 211–233.PubMedGoogle Scholar
  14. 14.
    Kimmel, C. B., Ballard, W. W., Kimmel, S. R., Ullmann, B., and Schilling, T. F. (1995) Stages of embryonic development of the zebrafish, Dev Dyn 203, 253–310.PubMedCrossRefGoogle Scholar
  15. 15.
    Warga, R. M., and Nusslein-Volhard, C. (1999) Origin and development of the zebrafish endoderm, Development 126, 827–838.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Division of Genetic Medicine, Department of MedicineVanderbilt University Medical CenterNashvilleUSA

Personalised recommendations