Additive Phenotypes Underlie Epistasis of Fitness Effects

Gene interactions, or epistasis, play a large role in determining evolutionary outcomes. The ruggedness of fitness landscapes, and thus the predictability of evolution and the accessibility of high-fitness genotypes, is determined largely by the pervasiveness of epistasis and the degree of correlation between similar genotypes. We created all possible pairings of three sets of five beneficial first-step mutations fixed during adaptive walks under three different regimes: selection on growth rate alone, on growth rate and thermal stability, and on growth rate and pH stability. All 30 double-mutants displayed negative, antagonistic epistasis with regard to growth rate and fitness, but positive epistasis and additivity were common for the stability phenotypes. This suggested that biophysically simple phenotypes, such as capsid stability, may on average behave more additively than complex phenotypes like viral growth rate. Growth rate epistasis was also smaller in magnitude when the individual effects of single mutations were smaller. Significant sign epistasis, such that the effect of a mutation that is beneficial in the wild-type background is deleterious in combination with a second mutation, emerged more frequently in intragenic mutational pairings than in intergenic pairs, and was evident in nearly half of the double-mutants, indicating that the fitness landscape is moderately uncorrelated and of intermediate ruggedness. Together, our results indicated that mutations may interact additively with regard to phenotype when considered at a basic, biophysical level, but that epistasis arises as a result of pleiotropic interactions between the individual components of complex phenotypes and diminishing returns arising from intermediate phenotypic optima.