Abstract
Mounting evidence suggests that d-amino acids play previously underappreciated roles in diverse organisms. In bacteria, even d-amino acids that are absent from canonical peptidoglycan (PG) may act as growth substrates, as signals, or in other functions. Given these proposed roles and the ubiquity of d-amino acids, the paucity of known d-amino-acid-responsive transcriptional control mechanisms in bacteria suggests that such regulation awaits discovery. We found that DarR, a LysR-type transcriptional regulator (LTTR), activates transcription in response to d-Asp. The d-Glu auxotrophy of a Vibrio fischerimurI::Tn mutant was suppressed, with the wild-type PG structure maintained, by a point mutation in darR This darR mutation resulted in the overexpression of an adjacent operon encoding a putative aspartate racemase, RacD, which compensated for the loss of the glutamate racemase encoded by murI Using transcriptional reporters, we found that wild-type DarR activated racD transcription in response to exogenous d-Asp but not upon the addition of l-Asp, l-Glu, or d-Glu. A DNA sequence typical of LTTR-binding sites was identified between darR and the divergently oriented racD operon, and scrambling this sequence eliminated activation of the reporter in response to d-Asp. In several proteobacteria, genes encoding LTTRs similar to DarR are linked to genes with predicted roles in d- and/or l-Asp metabolism. To test the functional similarities in another bacterium, darR and racD mutants were also generated in Acinetobacter baylyi In V. fischeri and A. baylyi, growth on d-Asp required the presence of both darR and racD Our results suggest that multiple bacteria have the ability to sense and respond to d-Asp.IMPORTANCE d-Amino acids are prevalent in the environment and are generated by organisms from all domains of life. Although some biological roles for d-amino acids are understood, in other cases, their functions remain uncertain. Given the ubiquity of d-amino acids, it seems likely that bacteria will initiate transcriptional responses to them. Elucidating d-amino acid-responsive regulators along with the genes they control will help uncover bacterial uses of d-amino acids. Here, we report the discovery of DarR, a novel LTTR in V. fischeri that mediates a transcriptional response to environmental d-Asp and underpins the catabolism of d-Asp. DarR represents the founding member of a group of bacterial homologs that we hypothesize control aspects of aspartate metabolism in response to d-Asp and/or to d-Asp-containing peptides.