FIGURE 3:

Complementation of an mgm101-null mutant by C-tail mutant alleles. (A) Meiotic analysis showing that mgm101^K253A, mgm101^W257A, mgm101^Y268A, and mgm101^R259A failed to complement the null mgm101Δ::kan allele for respiratory growth on ethanol plus glycerol medium at 30°C. A tetrad independently segregating the null mgm101Δ::kan and the mutant alanine alleles was dissected on complete glucose medium. The meiotic segregants were then replica plated onto G418, –Ura, and ethanol plus glycerol medium. G418^R marks the presence of the null mgm101Δ::kan allele, and Ura⁺ marks the mutant alanine alleles. Failure to grow on YPGE indicates the lack of complementation by mutant alanine alleles and the loss of mtDNA. (B) Temperature- and H₂O₂-induced petite production in the C-tail mutants. Meiotic segregants expressing only the mgm101^K251A, mgm101^R252A, mgm101^K260A, and mgm101^Y266A showed significantly increased production of petite colonies when challenged with high temperature or increasing concentrations of H₂O₂. The haploid cells were first grown in ethanol plus glycerol medium at 30°C. Cells were then inoculated in liquid YPD medium for 24 h at 37 or at 30°C in the presence of 1 or 2 mM H₂O₂. After dilution in water, cells were plated on YPD and incubated at 30°C for 5 d. Petites were identified as white colonies, and the petite frequency was scored as the percentage of white vs. total number of colonies. The data presented are the averaged petite frequency of at least three independent experiments. Error bars indicate average deviations.