Determinants of divergent adaptation and Dobzhansky-Muller interaction in experimental yeast populations

Abstract

Divergent adaptation can be associated with reproductive isolation in speciation [1]. We recently demonstrated the link between divergent adaptation and the onset of reproductive isolation in experimental populations of the yeast Saccharomyces cerevisiae evolved from a single progenitor in either a high-salt or a low-glucose environment [2]. Here, whole-genome resequencing and comparative genome hybridization of representatives of three populations revealed 17 mutations, six of which explained the adaptive increases in mitotic fitness. In two populations evolved in high salt, two different mutations occurred in the proton efflux pump gene PMA1 and the global transcriptional repressor gene CYC8; the ENA genes encoding sodium efflux pumps were overexpressed once through expansion of this gene cluster and once because of mutation in the regulator CYC8. In the population from low glucose, one mutation occurred in MDS3, which modulates growth at high pH, and one in MKT1, a global regulator of mRNAs encoding mitochondrial proteins, the latter recapitulating a naturally occurring variant. A Dobzhansky-Muller (DM) incompatibility between the evolved alleles of PMA1 and MKT1 strongly depressed fitness in the low-glucose environment. This DM interaction is the first reported between experimentally evolved alleles of known genes and shows how reproductive isolation can arise rapidly when divergent selection is strong.

Figure 1. Contribution of S2 and S6 evolved alleles to fitness in high salt

Shown are fitness measurements (OD₆₀₀, mean and standard error, normalized to the progenitor value) for 48 offspring fully genotyped for all coding alleles identified by sequencing – from each of the crosses S2 × P (A, C, 5 loci) and S6 × P (B, D, 3 loci). Data are aggregated by specific alleles as marked (in each marked category, e.g. “PMA1–2”, the other alleles are segregating). Full data (including intergenic loci) are available in Tables S3 and S4. (A, B). The bars represent the average fitness effect of each variant across all offspring. Light gray bars, ancestral alleles; dark bars, evolved alleles. Fitness of evolved parent is shown at the upper right corner. Significant differences are noted with P-value. (C, D) Average pair-wise effects of the two most advantageous mutations in each strain. Shown are the same data as in A and B, but averaged for two-locus genotypes showing positive interaction. Superscript a = ancestral allele; superscript e = evolved allele. Interaction was tested by ANOVA; all P values appear in Table S8.

Figure 2. Contribution of M8 evolved alleles to fitness in low glucose

(A, B) Average fitness effect of each variant across the segregant offspring at log-phase (20h) and post-diauxic shift (30h) during growth in low-glucose. Shown are fitness measurements (OD₆₀₀, mean and standard error, normalized to the progenitor value) for 48 progeny from an M8 × P cross – fully genotyped for all five coding loci identified by sequencing, at 20h (A) and 30h (B) of growth on glucose. Data are aggregated by specific alleles, as marked (in each marked category, e.g. “MKT1”, the other alleles are segregating). Full data are available in Table S5. Light gray bars, ancestral alleles; dark bars, evolved alleles. Fitness of evolved parent is shown at the upper right corner. Significant differences are noted with P-value. All P values appear in Table S8. (C) Evolved alleles of MDS3 and MKT1 (MDS3e and MKT1e) account for the M8 phenotype. Shown are growth curves (OD₆₀₀) from three tetrads from each of two independent crosses segregating for MDS3 and MKT1, and no other evolved alleles (based on full genotyping). The number of replicates for each time course varied between four and eight, reflecting independent assortment. The evolved allele of MDS3 (green) confers a benefit early, while that of MKT1 (blue) confers a benefit late in the growth cycle, relative to the ancestral genotype (black). Together these two alleles produce a phenotype (red) that matches that of the M8 strain (dashed).

Figure 3. Global expression changes in evolved strains associated with the adaptive genetic changes

(A) Genome wide expression profiles from P, S2, S6, and M8 strains grown in YPD, low glucose (LG), and high salt (HS) environments. Red – induced compared to mean of all strains in that condition; green – repressed compared to mean of all strains in that condition. (B) Genes with high expression specific to S6 across all conditions are enriched for Cyc8-Tup1 targets and for osmotic response genes. Shown is a zoomed in cluster from (A). Yellow bar – genes whose expression is induced in a deletion of the TUP1 gene [19]; purple bar – genes whose expression is induced during the Osmotic Stress Response (OSR) to high salt [20]. Genes are re-ordered by the TUP1 and OSR annotations. (C) Genes with high expression specific to M8 across all conditions are induced in the RM-11 wine strain and enriched for Puf3 targets. Top panel – zoomed in cluster from (A). Bottom panel – expression of the same genes in the laboratory strain BY and in the wild wine strain RM. Blue bar – genes in the Puf3 module [22], whose eQTL in a cross between BY and RM has been linked to the same genetic change in MKT1 found also in the M8 strain. Genes are re-ordered by membership in the Puf3 module.

Figure 4. DM interactions between the evolved alleles of PMA1 and MKT1

(A) DM interaction between the evolved alleles of PMA1 and MKT1 at 24h in low-glucose. Shown are the fitness measurements (OD₆₀₀, mean and standard error, normalized to the progenitor value) of 96 offspring of a cross between S2 and M8 in the low-glucose environment at 24h grouped by their two-locus genotypes for PMA1 and MKT1 (e - evolved allele; a - ancestral allele); note the depressed fitness of the genotype carrying both evolved alleles of these genes. ANOVA: evolved allele of PMA1, P < 10⁻⁴; evolved allele of MKT1, P < 10⁻⁴; and interaction of the evolved alleles of PMA1 and MKT1, P < 0.015. Full data are available in Table S6 and all P values of all tests are listed in Table S8. (B) DM interaction between the evolved alleles of PMA1 and MKT1 along the growth curve. Shown are growth curves from three tetrads from each of two independent crosses segregating for PMA1 and MKT1, and carrying no other evolved alleles (based on full genotyping). The number of replicates for each time course varied between four and eight, reflecting independent assortment. The genotype carrying the evolved alleles of PMA1 and MKT1 (red) shows poor growth at all time points (up to 27h) relative to the other genotypes. The other genotypes are marked as PMA1e (green); MKT1e (blue); ancestral (PMA1a MKT1a, black); and M8 (dashed). (C) Absence of an interaction between PMA1 and MDS3; analysis as in B: PMA1e (green); MDS3e (blue), PMA1e MDS3e (red); ancestral (PMA1a MDS3a, black); and M8 (dashed).