Ultrastructural Analysis of Cell Fusion in Yeast
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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 WordsUltrastructure 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.
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