Identification and functional analysis of Tdrxoo, the member of TonB-dependent-receptor family proteins in Xanthomonas oryzae pv. Oryzae

Abstract:

Abstract: [Objective] To demonstrate the novel regulatory pathways mediated in bacterial pathogenicity and motility in Xanthomonas oryzae pv. oryzae (Xoo), the casual agent of bacterial blight in rice. [Methods] Molecular identification and functional characterization of Tdrxoo, which interacts with GacAxoo of the two-component regulatory system (GacSxoo/GacAxoo) in Xoo, were performed through gene cloning, sequencing and disrupt analysis. [Results] tdrxoo was successfully cloned from the genomic DNA of wild-type PXO99A by using polymerase chain reactions with the degenerated primers tdrxooF/R. The tdrxoo gene was found to be highly conserved in the plant-pathogenic Xanthomonas spp. Sequence analysis showed that Tdrxoo was homological to a protein with the TonB-Dependent-Receptor (TDR) domain. Tdrxoo is probably localized in the outer membrane of bacterial cells, recognizing the signals from extracellular environment, and inducing the intracellular signal transduction. △tdrxoo, the disrupted mutant, was obtained after a single cross-over recombination event between tdrxoo and the plasmid pK-tdr with the tdrxoo segment. The mutant lost the ability of causing the disease, and was affected in growth in vitro compared to PXO99A. In addition, the motility and the extracellular enzymes production of △tdrxoo were reduced, which can be restored through complementation of the △tdrxoo mutant by introduction of tdrxoo. tdrxoo deficiency didn’t affect siderophore production. [Conclusion] According to the existence of tdrxoo in Xoo genome and phenotype of △tdrxoo, Tdrxoo, as the outer membrance protein, is proposed to be involved in regulation of pathogenicity, extracellular enzyme production, the growth and motility of Xoo.