Cell fusion during yeast mating requires high levels of a-factor mating pheromone

During conjugation, two yeast cells fuse to form a single zygote. Cell fusion requires extensive remodeling of the cell wall, both to form a seal between the two cells and to remove the intervening material. The two plasma membranes then fuse to produce a continuous cytoplasm. We report the characterization of two cell fusion defective (Fus-) mutants, fus5 and fus8, isolated previously in our laboratory. Fluorescence and electron microscopy demonstrated that the fus5 and fus8 mutant zygotes were defective for cell wall remodeling/removal but not plasma membrane fusion. Strikingly, fus5 and fus8 were a specific; both mutations caused the mutant phenotype when present in the MATa parent but not in the MAT alpha parent. Consistent with an a-specific defect, the fus5 and fus8 mutants produced less a-factor than the isogenic wild-type strain. FUS5 and FUS8 were determined to be allelic to AXL1 and RAM1, respectively, two genes known to be required for biogenesis of a-factor. Several experiments demonstrated that the partial defect in a-factor production resulted in the Fus- phenotype. First, overexpression of a-factor in the fus mutants suppressed the Fus- defect. Second, matings to an MAT alpha partner supersensitive to mating pheromone (sst2 delta) suppressed the Fus- defect in trans. Finally, the gene encoding a-factor, MFA1, was placed under the control of a repressible promoter; reduced levels of wild-type a-factor caused an identical cell fusion defect during mating. We conclude that high levels of pheromone are required as one component of the signal for prezygotes to initiate cell fusion.