A) Artificial mating pairs using the S. macrospora pheromone and receptor. First line: the mating pair using S. cerevisiae pheromones described in Figure 2A. Second line: the mating pair that expresses heterologous α-factor-like pheromone and the corresponding receptor from S. macrospora described in Figure 2A. Third line: a MATa cell expressing the S. macrospora α-factor pheromone (αFmac, green inverted α) mating with MATα cell expressing the α-factor receptor from S. macrospora (αRmac, green rounded receptor). This pair can communicate via α-factor like peptides only, but maintains the asymmetry at the mating locus (MATa and MATα cells). Fourth line: a MATα strain expressing S. cerevisiae’s α-factor receptor, usually expressed in MATa cells (αFcer, blue rounded receptor), and the pheromone from S. macrospora (αFmac, green inverted α). This cell can now communicate with the MATα αFcer αRmac cell described above, using only α-factor like peptides, in a mating with both cells expressing MATα. B) Visualizing mating with heterologous pheromone-receptor pairs. The cells described in A) were streaked, replica plated on top of each other, and allowed to mate over night in complete media. They were then replica plated onto media where only diploids could grow. The absence of off-diagonal mating shows that only strains expressing complementary pheromones and receptor pairs can mate. C) Quantitative mating data. The indicated crosses were allowed to mate on filters for 7 hours. Filters were then washed and cells plated on selective media to select for diploids. Mating efficiency is calculated as described in the Materials and Methods. Mating pairs where both strains express the MATα locus mate about 450 times worse than mating pairs that express different mating loci. Errors represent standard deviations of at least 3 independent trials. See Materials and Methods and main text for more details. Legend: Red: a-factor (aFcer) and a-factor (aRcer) receptor from S. cerevisiae. Blue: α-factor (αFcer) and α-factor receptor (αRcer) from S. cerevisiae. Green: α-factor (αFmac) and α-factor receptor (αRmac) from S. macrospora.