Gain-of-function experiments with bacteriophage lambda uncover residues under diversifying selection in nature

Maddamsetti, Rohan; Johnson, Daniel T.; Spielman, Stephanie J.; Petrie, Katherine L.; Marks, Debora S.; Meyer, Justin R.

To verify that the J and specificity region phylogenies are significantly different from one another, we compared their phylogenies to phylogenies constructed from all λ proteins. Our hypothesis is supported if the J specificity region phylogeny is more similar to other λ proteins than the phylogeny constructed from the adjacent J sequence. We computed phylogenies for all proteins in the λ genome, and compared them using Robinson-Foulds distance (which measures the topological distance between phylogenies) and geodesic distance (which takes both topology and branch lengths into account). When comparing phylogenies by Robinson-Foulds distance, many distances could not be computed because some proteins were missing in many of the genomes. In fact, the failure of this calculation demonstrates a high rate of gene loss, duplication, and horizontal transfer in lambdoid viruses. When comparing phylogenies by geodesic distance, four distinct clusters of phylogenies are apparent, showing which proteins tend to be coinherited. Importantly, the J specificity region does not cluster with the phylogeny estimated from the remaining J sequence that encompasses structural domains (Fig. S5) . From these analyses, as well as our visual observation of recombination in the J protein alignment (Data S1), we conclude that the specificity region is an evolutionary module that forms an independent functional unit distinct from the rest of J.