Fig 1.
A model for how MMR incompatible populations arise in nature [22,27].
In this cartoon, a common ancestor bearing the Mlh1 Gly 761 and Pms1 Arg 818/822 alleles can sustain mutations, neutral or beneficial, that give rise to the derived S288c (purple, Asp 761, Arg 818/822) and SK1 (green, Gly 761, Lys 818/822) group strains. Mating between the derived strains can yield an allele combination (Mlh1 Asp 761, Pms1 Lys 818/822) that had not been selected for function, leading to a negative epistatic interaction and a mutator phenotype. Sequencing analysis of a 32-kb region in the derived groups provided evidence for recombination between the two, supporting the idea that these two groups can meet in nature, exchange genetic information, and form a hybrid mutator [22].
Fig 2.
Incompatible strains display a fitness advantage in high salt media.
Independent cultures of compatible (kMLH1-kPMS1, EAY3242) and incompatible (cMLH1-kPMS1, EAY3236) strains were grown for up to Transfer 16 (~ 2 x 107 cells per transfer) in YPD (unevolved) or YPD + 1.2 M NaCl (evolved). Cultures at the indicated transfers were diluted to an OD600 of 0.1 (~ 2 x 107 cells per transfer) in YPD + 1.2 M NaCl and monitored for growth for 12 hrs. A representative experiment involving three replicates for each genotype is shown. Mean OD600, +/- standard deviation, is presented for each time point. See Materials and Methods for details.
Table 1.
Fitness of incompatible relative to compatible strains following competition.
Fig 3.
A single locus is likely responsible for high salt adaptation in compatible and incompatible strains.
A, High salt resistant clones (red) isolated from evolved compatible and incompatible cultures were crossed to an unevolved strain (blue). Diploids were selected and streaked onto YPD + 1.2 M NaCl plates to determine if NaClr in the evolved strain was dominant or recessive. Subsequently, diploids were sporulated and tetrad dissected, and spore clones were analyzed for resistance to NaCl. B. Left panel. An evolved NaCl resistant clone showing a dominant phenotype. Growth of the indicated haploid and diploid strains on YPD + 1.2 M NaCl plates is shown. Right panel, example of 2:2 NaClr:NaClssegregation. Growth on YPD + 1.2 M NaCl plates is shown for the spore clones of a single tetrad obtained by mating an evolved NaClr clone to an unevolved strain. C. The indicated incompatible and compatible evolved NaClr clones were each mated to an unevolved haploid strain and analyzed for segregation of NaCl resistance in tetrad analysis. For each mating the vast majority displayed 2:2 segregation of resistance to sensitivity. In total nine tetrads deviated from this pattern (“other” category) with eight showing 3:1 or 1:3 segregation and one showing 4:0 segregation.
Table 2.
Whole genome sequencing indicates PMR1 linkage to NaCl resistance.
Fig 4.
Location of pmr1 mutation alleles found in evolved strains.
21 pmr1 mutations (see Table 3 for the exact locations) identified in this study were mapped onto the Pmr1 structure predicted by Uniprot (Materials and Methods). * indicates the presence of a frameshift mutation.
Table 3.
pmr1 mutations identified in this study.
Fig 5.
pmr1 mutations identified in evolved lines are causative for resistance to 1.2 M NaCl and 0.4 M LiCl.
In both A and B, wild type and pmr1Δ unevolved strains were plated in 10-fold serial dilutions onto YPD, YPD + 1.2 M NaCl, and YPD + 0.4 M LiCl plates. In panel A, representative NaCl-evolved strains (I, incompatible, C, compatible, with the Transfer indicated) bearing pmr1 mutations are shown. In panel B, unevolved strains transformed to contain the indicated pmr1 alleles (Replacement) are shown, with the corresponding evolved strain plated side by side.