Expression Profiles Reveal Parallel Evolution of Epistatic Interactions Involving the CRP Regulon in Escherichia coli

Abstract

The extent and nature of epistatic interactions between mutations are issues of fundamental importance in evolutionary biology. However, they are difficult to study and their influence on adaptation remains poorly understood. Here, we use a systems-level approach to examine epistatic interactions that arose during the evolution of Escherichia coli in a defined environment. We used expression arrays to compare the effect on global patterns of gene expression of deleting a central regulatory gene, crp. Effects were measured in two lineages that had independently evolved for 20,000 generations and in their common ancestor. We found that deleting crp had a much more dramatic effect on the expression profile of the two evolved lines than on the ancestor. Because the sequence of the crp gene was unchanged during evolution, these differences indicate epistatic interactions between crp and mutations at other loci that accumulated during evolution. Moreover, a striking degree of parallelism was observed between the two independently evolved lines; 115 genes that were not crp-dependent in the ancestor became dependent on crp in both evolved lines. An analysis of changes in crp dependence of well-characterized regulons identified a number of regulatory genes as candidates for harboring beneficial mutations that could account for these parallel expression changes. Mutations within three of these genes have previously been found and shown to contribute to fitness. Overall, these findings indicate that epistasis has been important in the adaptive evolution of these lines, and they provide new insight into the types of genetic changes through which epistasis can evolve. More generally, we demonstrate that expression profiles can be profitably used to investigate epistatic interactions. The effect of a genetic mutation can depend on the genotype of the organism in which it occurs. For example, a mutation that is beneficial in one genetic background might be neutral or even deleterious in another. The interactions between genes that cause this dependence—known as epistasis—play an important role in many evolutionary theories. However, they are difficult to study and remain poorly understood. We used a novel approach to examine the evolution of interactions arising between a key regulatory gene, crp, and mutations that occurred during the adaptation of a bacterium, Escherichia coli, to a laboratory environment. To do this, we measured the effect of deleting crp on the expression of all genes in the organism, providing a sensitive measure to identify new interactions involving this gene. We found that deleting crp had a dramatic and parallel effect on gene expression in two independently evolved populations, but much less effect in their ancestor. An analysis of these changes identified a number of regulatory genes as candidates for harboring beneficial mutations that could account for the parallel changes. These findings indicate that epistasis has played an important role in the evolution of these populations, and they provide insight into the types of genetic changes through which epistasis can evolve.