Regulation of Denitrification and Quorum Sensing in Sinorhizobium meliloti

The overall goal of this study was to identify and discern the functional role of a quorum sensing-like transcriptional regulator in Sinorhizobium meliloti. This aerobic soil-dwelling bacterium forms a symbiotic relationship Medicago sativa and is an excellent model for hostsymbiont interactions. Quorum sensing, bacterial cell-cell communication, allows S. meliloti to coordinate its gene expression in a population density dependent manner. The process of symbiosis is dependent on the SinRI/ExpR quorum sensing system, which coordinates the expression of genes responsible for exopolysaccharide production and motility. Quorum sensing-like regulators, or orphan regulators, play an important role in S. meliloti but may not be involved in the process of cell-cell communication. For example, the orphan regulator NesR is involved in stress response, but does not respond to known quorum sensing signal molecules. The SMc00658 locus encodes another such orphan regulator that we have named Adr (aerobic denitrification regulator). The protein sequence of Adr has some similarity to typical LuxR-like proteins, including a predicted N-terminal signal binding domain and a Cterminal DNA binding domain. To determine the role of Adr, the transcriptomic profile of an adr mutant was compared to wildtype S. meliloti. The results of this analysis indicated that Adr is involved in regulating denitrification and microoxic respiration in S. meliloti. Since the transcriptomic study was performed under aerobic conditions, it is likely that Adr is involved in regulating aerobic denitrification or detoxification of nitrogen oxides. To test this, assays designed to determine the denitrification ability of the adr mutant were performed. Additionally, plant symbiosis and competition assays were performed to determine the symbiotic capability of the adr mutant compared to wild-type S. meliloti. From these studies we demonstrate that the adr mutant is a less efficient denitrifier than its wildtype counterpart. We also found that while Adr is not required for nitrogen fixation, strains deficient in this regulator will not be as competitive as wild-type S. meliloti when invading M. sativa. The results obtained in this study enhance the current understanding of denitrification regulation in S. meliloti and will be essential to further understanding aerobic denitrification in this and other denitrifying bacteria.