Abstract
<jats:p> Many bacteria engage in quorum sensing (QS), a cell-cell communication system used to coordinate group behaviors. In one type of QS, acyl-homoserine lactone signals generated by LuxI homologs bind to LuxR homolog transcription factors, usually resulting in gene activation. The genome of <jats:italic>Pseudomonas aeruginosa</jats:italic> encodes three such LuxR homologs: LasR, RhlR, and QscR. Of these, LasR regulates the most genes, including that encoding RhlR. There is strong evidence that, during chronic infections, <jats:italic>lasR</jats:italic> and other genes encoding LuxR-type regulators are under strong selective pressure for mutations that both inactivate and modulate their function. Thus, we wondered if some mutations in the <jats:italic>lasR</jats:italic> gene might result in a protein with affinity for promoters usually regulated by the other LuxR homologs; to do so, we investigated the DNA-binding domain (DBD) of LasR through alanine substitution. As expected, we found that most alanine substitutions across the LasR DBD led to loss of function, as did previously identified clinical LasR DBD variants. Additionally, some alanine mutants were indistinguishable from the wild type. We describe a handful of variant LasR polypeptides that unexpectedly exhibit enhanced regulation on a RhlR-regulated gene, which conferred a fitness defect when competed against the wild type. Most other LasR variants had a competitive advantage. Our results suggest a pathway for expansion of the regulon of LuxR-homolog transcription factors, but also that such mutations may be disfavored due to the incurred metabolic burden. </jats:p>