The Regulation Mechanism Of Glutathione On The Type ? Secretion System In Pseudomonas Aeruginosa

Pseudomonas aeruginosa is a human opportunistic pathogen which causes serious infection.Glutathione is one of the most important antioxidants in living cells.Previously,we found that the deletion of glutathione(GSH)synthase gene gsh B in P.aeruginosa resulted in the decrease of the expression of type III secretion system(T3SS),but the specific mechanism is not clear.In order to study the role of glutathione in P.aeruginosa further,we constructed glutamate-cysteine ligase synthesis gene deletion mutant(35)gsh A,glutathione synthase gene deletion mutant(35)gsh B,and mutants(35)gsh A(35)gsh B with simultaneous deletion of two genes.RNA-seq transcriptome analysis showed that not only the expression of type III secretion system,but also the expression of type VI secretion system(T6SS)in(35)gsh A(35)gsh B was significantly reduced.Acute infection of lung in mice showed cytotoxicity decreased in(35)gsh A(35)gsh B,suggesting that glutathione played an important role in the pathogenicity of P.aeruginosa.Exs A is the key regulatory protein of T3 SS.Vfr is able to regulate the secretory system of type III in Pseudomonas aeruginosa by activating the transcription of exs A.The expression of exo S in P.aeruginosa increased significantly when exogenous glutathione was added to the medium which T3 SS is induced,while the expression of T3 SS in ?vfr was not regulated by glutathione.The complementary strain ?vfr/vfr could restore the induction of glutathione to T3 SS.Therefore,these data suggest that glutathione regulates T3 SS through Vfr.There are five cysteine residues in Vfr.When all cysteine residues are replaced by alanine through site-directed mutation,the ability of glutathione to induce T3 SS expression is disappeared.In vitro experiments showed that glutathione is able to restore oxidized Vfr to reduced state.These results suggest that glutathione,an intracellular redox signaling molecule,regulates the expression of T3 SS in P.aeruginosa by altering the redox state of Vfr.The results of this study elucidate the role of glutathione in P.aeruginosa further.