0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not
- 0 International license peer-reviewed) is the…
19 20. CC-BY-NC-ND 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not. Abstract 21 Species achieve evolutionary innovations through two major genetic 22 mechanisms, namely regulatory-and structural-level mutations. The ability of 23 populations to evolve involves a balance between selection, genetic drift, epistasis, 24 biochemical and biophysical requirements, thermodynamic properties and other 25 factors. This adaptive diversity begs the question as to whether a restricted pathway 26 governs adaptations or whether multiple pathways are possible to achieve an adaptive 27 response. By combining a unique set of tools drawn from synthetic biology, 28 evolutionary biology and genomics, we experimentally evolved and then characterized 29 the adaptive properties of a modern E. coli strain containing a 700 million-year-old 30 reconstructed ancestral Elongation Factor Tu (EF-Tu) gene inserted into its genome for 31 the first time. We then tracked the evolutionary steps taken by the ancient-modern 32 hybrid microorganism through laboratory evolution by monitoring genomic mutations. 33 This study reveals that lineages respond to the ancient gene by increasing the 34 expression levels of the maladapted protein, rather than through direct accumulation of 35 mutations in the open reading frame. In particular, these findings show that the general 36 strategy for the bacteria to adapt to the ancient protein is to accumulate mutations in 37 the cis-regulatory region; gene-coding mutations appear to preclude rapid adaptation 38 upon integration of the ancient gene for our system.