Cell Fusion pp 197-211 | Cite as

Ultrastructural Analysis of Cell Fusion in Yeast

  • Alison E. Gammie
Part of the Methods in Molecular Biology™ book series (MIMB, volume 475)


The process of creating a single cell from two progenitor cells requires molecular precision to coordinate the events leading to cytoplasmic continuity while preventing lethal cell lysis. Cell fusion characteristically involves the mobilization of fundamental processes, including signaling, polarization, adhesion, and membrane fusion. The yeast Saccharomyces cerevisiae is an ideal model system for examining the events of this critical and well-conserved process. Researchers employ yeast cells because they are rapidly growing, easy to manipulate, amenable to long-term storage, genetically tractable, readily transformed, and nonhazardous. The genetic and morphological characterizations of cell fusion in wild-type and fusion mutants have helped define the mechanism and temporal regulation required for efficient cell fusion. Ultrastructural studies, in particular, have contributed to the characterization of and revealed striking similarities within cell fusion events in higher organisms. This chapter details two yeast cell fusion ultrastructural methods. The first utilizes an ambient temperature chemical fixation, and the second employs a combination of high-pressure freezing and freeze substitution.

Key Words

Ultrastructure cell fusion yeast mating electron microscopy freeze substitution high-pressure freezing 



I thank Kent McDonald, Max Heiman, and Peter Walter for sharing valuable information about the high-pressure freezing and freeze substitution procedures and Margaret Bisher for helping to implement the procedure at Princeton University.


  1. 1.
    Elion, E. A. (2000) Pheromone response, mating and cell biology. Curr. Opin. Microbiol. 3, 573–581.CrossRefPubMedGoogle Scholar
  2. 2.
    White, J. M. and Rose, M. D. (2001) Yeast mating: getting close to membrane merger. Curr. Biol. 11, R16–R20.CrossRefPubMedGoogle Scholar
  3. 3.
    Marsh, L. and Rose, M. D. (1997) The pathway of cell and nuclear fusion during mating in S. cerevisiae, in The Molecular and Cellular Biology of the Yeast Saccharomyces: Cell Cycle and Cell Biology (J. R. Pringle, J. R. Broach, and E. W. Jones. eds.), vol. 3. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 827–888.Google Scholar
  4. 4.
    Gammie, A. E., Brizzio, V. , and Rose, M. D. (1998) Distinct morphological phe-notypes of cell fusion mutants. Mol. Biol. Cell. 9, 1395–1410.PubMedGoogle Scholar
  5. 5.
    Wright, R. (2000) Transmission electron microscopy of yeast. Microsc. Res. Tech. 51, 496–510.CrossRefPubMedGoogle Scholar
  6. 6.
    Gammie, A. E. and Rose, M. D. (2002) Assays of cell and nuclear fusion. Methods Enzymol. 351, 477–498.CrossRefPubMedGoogle Scholar
  7. 7.
    Heiman, M. G., Engel, A., and Walter, P. (2007) The Golgi-resident protease Kex2 acts in conjunction with Prm1 to facilitate cell fusion during yeast mating. J. Cell Biol. 176, 209–222.CrossRefPubMedGoogle Scholar
  8. 8.
    McDonald, K. L. and Auer, M. (2006) High-pressure freezing, cellular tomography, and structural cell biology. Biotechniques 41, 137, 139, 141 passim. CrossRefGoogle Scholar
  9. 9.
    Dawes, C. J. (1979) Biological Techniques for Transmission and Scanning Electron Microscopy. Ladd Research Industries, Burlington, VT.Google Scholar
  10. 10.
    Hayat, M. A. (2000) Principles and Techniques of Electron Microscopy: Biological Applications, 4th ed. Cambridge University Press, Cambridge, England.Google Scholar
  11. 11.
    Robinson, D. G. (1987) Methods of Preparation for Electron Microscopy: An Introduction for the Biomedical Sciences. Springer-Verlag, Berlin.Google Scholar
  12. 12.
    Perlin, A. S. (2006) Glycol-cleavage oxidation. Adv. Carbohydr. Chem. Biochem. 60, 183–250.CrossRefPubMedGoogle Scholar
  13. 13.
    van Tuinen, E. and Riezman, H. (1987) Immunolocalization of glyceraldehyde-3-phosphate dehydrogenase, hexokinase, and carboxypeptidase Y in yeast cells at the ultrastructural level. J. Histochem. Cytochem. 35, 327–333.PubMedGoogle Scholar
  14. 14.
    McDonald, K. (1999) High-pressure freezing for preservation of high resolution fine structure and antigenicity for immunolabeling. Methods Mol. Biol. 117, 77–97.CrossRefPubMedGoogle Scholar
  15. 15.
    Walther, P. and Ziegler, A. (2002) Freeze substitution of high-pressure frozen samples: the visibility of biological membranes is improved when the substitution medium contains water. J. Microsc. 208, 3–10.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science + Business Media, LLC 2008

Authors and Affiliations

  • Alison E. Gammie
    • 1
  1. 1.Department of Molecular BiologyPrinceton UniversityPrinceton

Personalised recommendations