Response regulator VemR regulates the transcription of flagellar rod gene flgG by interacting with σ54 factor RpoN2 in Xanthomonas citri ssp. citri

Xanthomonas citri ssp. citri, a polar flagellated bacterium, causes citrus canker disease worldwide. In this study, we found that the X. citri ssp. citri response regulator VemR plays a regulatory role in flagellum-derived cell motility. Deletion of the vemR gene resulted in a reduction in cell motility, as well as reductions in virulence and exopolysaccharide production. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that vemR is transcribed in an operon together with rpoN2 and fleQ. In the vemR mutant, the flagellar distal rod gene flgG was significantly down-regulated. Because flgG is also rpoN2 dependent, we speculated that VemR and RpoN2 physically interact, which was confirmed by yeast two-hybrid and maltose-binding protein (MBP) pull-down assays. This suggested that the transcription of flgG is synergistically controlled by VemR and RpoN2. To confirm this, we constructed a vemR and rpoN2 double mutant. In this mutant, the reductions in cell motility and flgG transcription were unable to be restored by the expression of either vemR or rpoN2 alone. In contrast, the expression of both vemR and rpoN2 together in the double mutant restored the wild-type phenotype. Together, our data demonstrate that the response regulator VemR functions as an RpoN2 cognate activator to positively regulate the transcription of the rod gene flgG in X. citri ssp. citri.