The K6001 yeast with background W303; wild-type BY4741yeast strain; ∆sod1, ∆sod2,uth1, ∆skn7, ∆cat, ∆gpx, ∆atg2, andatg32 mutants with K6001 background; YOM36 and YOM38 containing pR316-GFP-ATG8 plasmid were used. The K6001 yeast strain was obtained from Professor Michael Breitenbach (University of Salzburg, Austria). The K6001 mutant strains of ∆sod1, ∆sod2, ∆uth1, ∆skn7, ∆cat, ∆gpx, ∆atg2, ∆atg32, BY4741, YOM36, and YOM38 containing pR316-GFP-ATG8 plasmid were provided by Professor Akira Matsuura (Chiba University, Japan). The genotypes of yeast strains and mutants are described in our previous research [19], please see them in Supplementary Table S1.

Replicative lifespan assays were performed following a previous method [20]. In brief, the K6001 yeast strain was collected from a −30 °C freezer after washing with PBS thrice, inoculated in galactose liquid medium (YPG, 1% yeast extract, 2% hipolypeptone, and 3% galactose), and cultured with shaking at 28 °C and 180 rpm for 24 h. After the yeast cells reached the logarithmic growth phase, 4000 yeast cells were inoculated in glucose solid medium (YPD, 2% glucose, 2% hipolypeptone, 1% yeast extract, and 2% agar), which was added with samples in advance and incubated at 28 °C for 48 h. Forty mother cells were randomly selected under the microscope, and surrounding daughter cells were counted. The method for determining conducting the replicative lifespans of ∆sod1, ∆sod2, ∆uth1, ∆skn7, ∆cat, ∆gpx, ∆atg2, and ∆atg32 mutant strains with the background of K6001 was the same as that for K6001 yeast.

The chronological lifespan assay was conducted as described in our previous study [20]. In brief, the YOM36 yeast strain was collected from a −30 °C freezer and washed with PBS, and about 200 yeast cells were inoculated in glucose solid medium and cultured at 28 °C for 48 h in the stationary phase. A single colony was picked out to a synthetic defined (SD) medium (0.17% yeast nitrogen base without amino acids and ammonium sulfate, 0.5% ammonium sulfate, and 2% glucose) and cultured with shaking at 28 °C and 180 rpm for 24 h. At day 0, yeast cells were inoculated into a new 100 mL SD medium with 0.01 OD600 value; treated with 0, 1, and 3 µM GENI; and cultured with shaking at 28 °C and 180 rpm. At day 3, about 200 yeast cells were spread onto glucose solid medium and cultured at 28 °C for 48 h, and the colony forming unit (CFU) of each plate was counted. The CFU of the third day was denoted as 100% survival. These steps were repeated every two days, and the survival rate of each group (CFU per plate/CFU of the same plate on the third day × 100%) was calculated until the survival rate dropped below 5%.

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