The Regulation Of Pathogenicity By Two Potential Regulators In Pseudomonas Aerugionsa

The Gram-negative bacterium Pseudomonas aerugionsa is a prevalent and dangerous pathogen which causes different infections in humans. It persists in different environments including in animal hosts mainly due to its tremendous metabolic diversity and abundant regulatory networks. The small protein RsmA in the Gac/Rsm regulatory pathway is a post-transcriptional RNA-binding protein that acts as a pleiotropic global regulator in P.aeruginosa. The RsmA regulon includes numerous genes, many of which are important in P. aeruginosa pathogenicity. In our previous search for regulatory genes of rsmA, the gene of PA4664 (prmC) was isolated in which mutant the expression of rsmA was greatly reduced. The homologous gene of PA4664 in E.coli, named prmC, is an S-adenosyl-L-methionine (AdoMet)-dependent methyltransferase of peptide chain release factors (RFs)The aim of this study was to investigate the mechanism of the regulation of rsmA by PA4664 and the effects of PA4664 on other virulence factors. The results obtained show that the motility including swimming?swarming and twitching was defective in ?PA4664 mutant. The motility could be restored to the wild type level when rsmA was over expressed in the ?PA4664 mutant background. The structures of flagellum and fimbriae showed no much difference between the wild type and the ?PA4664 mutant, the regulation of motility by PA4664 was not by affecting the structure of flagellum or fimbriae but via affecting of RsmA.In addition to mobility, the ?PA4664 mutant also had major effect on the expression of quorum-sensing system (QS) and the formation of biofilm. Both of QS and the formation of biofilm were significantly reduced in comparison with the wild type. Intracellular c-di-GMP signal in the mutant was no more than a half of that in the wild type strain. The production of pyocyanin was hardly detectable in the ?PA4664 mutant. Using fruit fly (Drosophila melanogaster Canton S) and Chinese cabbage (Brassica pekinensis) infection models, ?PA4664 mutant showed significantly reduced pathogenicity compared with the wild type. Complementation with ?PA4664 carried on a plasmid restored the motility, pyocyanin production and the level of c-di-GMP to the wild type. In conclusion, these results indicate that the PA4664 is a potential regulator of rsmA, and it plays an important role in modulating pathogenicity and cellular processes in P.aeruginosa.Meanwhile, the cellular localization and function of another potential regulator PA4595 were also determined. PA4595 protein belongs to the REG sub-family of ATP-binding cassette (ABC) family. Western blot result showed that the PA4595 protein was localized in the cytoplasm. Unlike the common ABC locating on the membrane, the PA4595 protein is probably not involved in transportation. The results also indicate ?PA4595 mutant caused increased expression of T3SS secretory proteins, however, the effect of PA4595 on the T3SS was not through the GacSA-RsmY/Z-RsmA-ExsA or vfr pathways.