Figure 1. Yeast mating efficiency and shmoo formation depends on the availability of spermidine but is autophagy-independent. (A) Mating efficiency of wild-type and Δspe2 cells with or without external administration of 100 µM spermidine as determined via clonogenic mating assay. Mating efficiency of wild-type control was set at 1. Data represent mean ± S.E.M. (n = 3; ***p < 0.001). (B) Representative sample pictures of wild-type and Δspe2 cells (MATa) after treatment with 1 µM α -pheromone for 3 h with or without external administration of 100 µM spermidine. (C) Microscopic quantification of cells treated as described in (B) and displaying a shmoo. Only viable cells were considered, and at least 3,000 cells per strain and experiment were manually counted. The portion of cells displaying a shmoo was normalized to that of the untreated wild type control, which was set at 1. Data represent means ± S.E.M (n = 4; **p < 0.01). (D) Relative intracellular concentration of spermidine and its precursor putrescine per OD₆₀₀ of wild-type cells (mating type a) either treated with 1 µM α-pheromone or left untreated (control), as determined using LC/MS/MS and normalized to the intracellular concentrations of the untreated control. Data represent means ±S.E.M. (n = 5; ^***p < 0.001). (E) Mating efficiency of wild-type, Δspe2 and Δspe4 strains. Data were determined via clonogenic mating assay and represent means ± S.E.M. (n = 3; ***p < 0.001). (F) Shmoo-formation in wild-type, Δspe2 and Δspe4 cells (mating type a) after treatment with 1 µM α-pheromone for 3 h (compare to Fig. 1B). Data represent means ± S.E.M. (n = 4; **p < 0.01). (G) Mating efficiency of wild-type and Δatg7 cells. Data were determined via clonogenic mating assay and represent means ± S.E.M. (n = 3; ***p < 0.001). Mating efficiency of wild-type control was set at 1. (H) Mating efficiency of wild-type, Δspe2 and Δspe2Δatg7 strains with or without external administration of 100 µM spermidine. Data was determined via clonogenic mating assays and represent means ± S.E.M. (n = 3; ***p < 0.001). Mating efficiency of wild-type control was set at 1.