Pheromone Induces Programmed Cell Death in S. cerevisiae

Programmed cell death is a ubiquitous process in multicellular organisms. The pathways by which cells in multicellular organisms trigger cell death have been extensively characterized. A typical scenario for programmed cell death development includes activation of the MAP kinase cascade, accumulation of reactive oxygen species, release of cytochrome c from mitochondria into cytoplasm as a consequence of opening the permeability transition pore (PTP) in the mitochondrial membrane, and activation of caspases (reviewed in [1]). It remains unclear whether single cell organisms have evolved such pathways. And it is unclear what benefit would accrue to a single cell organism undergoing programmed cell death. One reason for a unicellular organism to commit altruistic suicide may be to benefit the cell community. Such a mechanism would improve the genetic fund of the community by eliminating the weak individuals. Yeast show aspects of communal behaviour when they mate. There are two mating types in yeast: α α and a. Cells of α α mating type produce alpha-factor, triggering cells of a type to mate and vice versa. High doses or prolonged exposure to sexual pheromone is toxic for yeast (see [2] for review). The altruistic death of yeast cells unable to mate after a long time in contact with cells of the opposite mating type might be beneficial for the cell community. We decided to test whether the toxicity of alpha-factor could be explained by induction of programmed cell death. One of the key mechanisms by which cells trigger programmed cell death is by production of reactive oxygen species (see [1] for review). 2′ ′,7′ ′-dichlorodihydro-fluorescein diacetate (H 2 DCF-DA) can be used as a probe for reactive oxygen species production; H 2 DCF-DA is oxidized by reactive oxygen species to DCF which fluoresces green [3]. We tested whether alpha-factor addition to a cells could trigger DCF accumulation. Experiments showed that after 1.5 hr exposure of a cells to alpha-factor approximately 30% of a-type cells fluoresce, whereas no fluorescing cells are found either in the absence of pheromone (Figure 1A), or when alpha-factor is added to α α cells (Table 1). We conclude that addition of alpha-factor to cells of the opposite mating type can induce the formation of reactive oxygen species, a marker of programmed cell death. We titrated the concentration of pheromone and monitored the appearance of shmoos, morphological markers of mating. We found that reactive oxygen species induction requires approximately 10-fold …