Abstract
The evolution of bacterial regulatory networks has largely been explained at macroevolutionary scales through lateral gene transfer and gene duplication. Transcription factors (TF) have been found to be less conserved across species than their target genes (TG). This would be expected if TFs accumulate mutations faster than TGs. This hypothesis is supported by several lab evolution studies which found TFs to be frequently mutated. Despite these studies, the contribution of point mutations in TFs to the evolution of regulatory network, especially at microevolutionary scales is poorly understood. We tested if TFs show greater sequence diversity than their TGs using whole-genome sequencing data on thousands of clinical and environmental isolates of E coli. We found that TFs were less diverse in sequence than their TGs, and that their diversity was constrained by their regulatory roles. Over longer time-scales, the conservation of TFs, other than global regulators (GR), was low across species. Over very short time-scales represented by lab evolution studies, we confirmed an excess of beneficial mutations in TFs. TFs accumulated mutations much faster than TGs in the first 10,000 generations of a long-term evolution experiment. However, as evolution proceeded, mutations appeared in TFs at rates similar to or lower than those in TGs. Our results suggest that point mutations, conferring large-scale expression changes, may drive the early stages of adaptation but gene regulation is subjected to stronger purifying selection post adaptation.