Modular epistasis and the compensatory evolution of gene deletion mutants

Abstract

Screens for epistatic interactions have long been used to characterize functional relationships corresponding to protein complexes, metabolic pathways, and other functional modules. Although epistasis between adaptive mutations is also common in laboratory evolution experiments, the functional basis for these interactions is less well characterized. Here, we quantify the extent to which gene function (as determined by a genome-wide screen for epistasis among deletion mutants) influences the rate and genetic basis of compensatory adaptation in a set of 37 gene deletion mutants nested within 16 functional modules. We find that functional module has predictive power: mutants with deletions in the same module tend to adapt more similarly, on average, than those with deletions in different modules. At the same time, initial fitness also plays a role: independent of the specific functional modules involved, adaptive mutations tend to be systematically more beneficial in less-fit genetic backgrounds, consistent with a general pattern of diminishing returns epistasis. We measured epistatic interactions between initial gene deletion mutations and the mutations that accumulate during compensatory adaptation and found a general trend towards positive epistasis (i.e. mutations tend to be more beneficial in the background in which they arose). In two functional modules, epistatic interactions between the initial gene deletions and the mutations in their descendant lines caused evolutionary entrenchment, indicating an intimate functional relationship. Our results suggest that genotypes with similar epistatic interactions with gene deletion mutations will also have similar epistatic interactions with adaptive mutations, meaning that genome scale maps of epistasis between gene deletion mutations can be predictive of evolutionary dynamics.

Fig 1. Experimental design showing genotypes of important strains.

Schematic showing 5 of the 37 gene deletion founders, from 2 of the 16 different functional modules studied here (see Table 1 for the full list). After constructing each gene deletion founder, we founded 20 replicate populations from each and allowed them to evolve for 500 generations. Finally, we reverted the initial gene deletion in a subset of clones from these evolved populations.

Fig 2. Fitness evolution.

(A) Relationship between the initial fitness of the 37 Founder gene deletion mutants and the fitness of each of the 20 replicate populations after 500 generations of evolution, colored according to functional module. The x-coordinate is jittered slightly for visibility. (B) Relationship between initial fitness of the 37 Founder gene deletion mutants and the mean fitness gain of the 20 replicate populations descended from that Founder after 500 generations of evolution. Horizontal error bars show standard errors of the mean initial fitness and vertical error bars show standard deviations in the fitness of descendant populations. (C) Fraction of the variance between populations in fitness gain after 500 generations of evolution that is attributable to each indicated component. All variance components are significant (Table 3).

Fig 3. Epistasis between Founder gene deletion and evolved mutations.

(A) Relationship between the fitness effects of evolved mutations in the Founder deletion background compared to their effect in the wild type background. Boxed region shows area expanded in (B). (C) Epistasis, ϵ, between evolved mutations and Founder deletion mutation plotted against the fitness effect of Founder deletion mutation. Refer to Fig 2 for the symbol legend. (D) Fraction of the variance between populations in epistasis between Founder gene deletion and evolved mutations that is attributable to each indicated component. The analysis was performed with all reverted populations (All) and repeated excluding descendants of ade2Δ, hxk2Δ and tps1Δ (Without entrenched). All variance components are significant (Table 4).

Fig 4. Effects of reverting founding gene deletion mutations.

Fitness effects of reverting the initial gene deletion mutations in the evolved background plotted against the fitness effect of reverting the initial gene deletion in the wild-type background. Refer to Fig 2 for the symbol legend.

Fig 5. List of multi-hit mutations.

Genes that were independently mutated in at least two different populations by Founder gene deletion (number of populations analyzed), grouped by mutation module and Founder module.