Mechanisms Of The Toxin/Antitoxin System HigB/HigA Mediated Regulation On Biofilm Formation And The Type Ⅲ Secretion System In Pseudomonas Aeruginosa

Pseudomonas aeruginosa is an important conditional pathogen,burn victims and immunocompromised patients are susceptible to infection.Toxin-antitoxin system(TA)usually consists of two co-expressed genes,one coding the toxin and the other coding the antitoxin.Previous studies revealed that Pseudomonas aeruginosa toxin HigB is involved in the persister formation as well as the regulation of biofilm formation and the expression of the type Ⅲ secretion system(T3SS)genes.Biofilm is a kind of bacterial aggregation formed by bacteria adhering to the solid surface,which is usually found during the chnic infection of P.aeruginosa.The T3 SS is a needle-like complex composed of multiple proteins.Upon contact with host cells,the T3 SS effector factors can be injected into host cells through the needle structure,causing cell death.The T3 SS plays an important role in the acute infection caused by P.aeruginosa.C-di-GMP,known as cyclic guanosine diphosphate,is an important secondary regulatory signal in bacteria.It is synthesized by two GTP molecules catalyzed by guanosine cyclase.The level of c-di-GMP in bacteria is regulated by synthetases and degrading enzymes.In this study,we explored the mechanism of the toxin HigB mediated regulation on biofilm and the T3 SS.Through molecular biology and genetics,we found that the toxin HigB reduced the level of c-di-GMP in cells,thus inhibiting the formation of biofilm and increasing the expression of T3 SS genes..By analyzing the expression levels of the known genes related to c-di-GMP metabolism,we found that three genes responsible for c-di-GMP degradation were up-regulated by the toxin HigB,namely PA2133,PA2200 and PA3825.Knock-out of these three genes individually or simultaneously diminished the HigB caused reduction of the intracellular c-di-GMP level and biofilm formation as well as the up regulation of the T3 SS genes induced by toxin HigB.In conclusion,our study shows that HigB plays an important role in biofilm formation and T3 SS of P.aeruginosa through the regulation of c-di-GMP.