Roles of RpoS in Yersinia pseudotuberculosis stress survival, motility, biofilm formation and type VI secretion system expression.

RpoS (σ(S)), the stationary phase/stress σ factor, controls the expression of a large number of genes involved in cellular responses to a variety of stresses. However, the role of RpoS appears to differ in different bacteria. While RpoS is an important regulator of flagellum biosynthesis, it is associated with biofilm development in Edwardsiella tarda. Biofilms are dense communities formed by bacteria and are important for microbe survival under unfavorable conditions. The type VI secretion system (T6SS) discovered recently is reportedly associated with several phenotypes, ranging from biofilm formation to stress sensing. For example, Vibrio anguillarum T6SS was proposed to serve as a sensor for extracytoplasmic signals and modulates RpoS expression and stress response. In this study, we investigated the physiological roles of RpoS in Yersinia pseudotuberculosis, including bacterial survival under stress conditions, flagella formation, biofilm development and T6SS expression. We found that RpoS is important in resistance to multiple stressors-including H2O2, acid, osmotic and heat shock-in Y. pseudotuberculosis. In addition, our study showed that RpoS not only modulates the expression of T6SS but also regulates flagellum formation by positively controlling the flagellar master regulatory gene flhDC, and affects the formation of biofilm on Caenorhabditis elegans by regulating the synthesis of exopolysaccharides. Taken together, these results show that RpoS plays a central role in cell fitness under several adverse conditions in Y. pseudotuberculosis.

FliS modulates FlgM activity by acting as a non-canonical chaperone to control late flagellar gene expression, motility and biofilm formation in Yersinia pseudotuberculosis.

The FlgM-FliA regulatory circuit plays a central role in coordinating bacterial flagellar assembly. In this study, we identified multiple novel binding partners of FlgM using bacterial two-hybrid screening. Among these binding partners, FliS, the secretion chaperone of the filament protein FliC, was identified to compete with FliA for the binding of FlgM. We further showed that by binding to FlgM, FliS protects it from secretion and degradation, thus maintaining an intracellular pool of FlgM reserved as the FliS-FlgM complex. Consequently, we found that the flagellar late-class promoter activities are significantly increased in the fliS deletion mutant. The fliS mutant is weakly motile and shows significantly increased biofilm formation on biotic surface. Based on the results obtained, we established for the first time the regulatory role of the flagellin chaperone FliS to fine-tune late flagellar assembly by modulating FlgM activity.

A type VI secretion system regulated by OmpR in Yersinia pseudotuberculosis functions to maintain intracellular pH homeostasis.

Type VI secretion systems (T6SSs) which widely distributed in Gram-negative bacteria have been primarily studied in the context of cell interactions with eukaryotic hosts or other bacteria. We have recently identified a thermoregulated T6SS4 in the enteric pathogen Yersinia pseudotuberculosis. Here we report that OmpR directly binds to the promoter of T6SS4 operon and regulates its expression. Further, we observed that the OmpR-regulated T6SS4 is essential for bacterial survival under acidic conditions and that its expression is induced by low pH. Moreover, we showed that T6SS4 plays a role in pumping H(+) out of the cell to maintain intracellular pH homeostasis. The acid tolerance phenotype of T6SS4 is dependent on the ATPase activity of ClpV4, one of the components of T6SS4. These results not only uncover a novel strategy utilized by Y. pseudotuberculosis for acid resistance, but also reveal that T6SS, a bacteria secretion system known to be functional in protein transportation has an unexpected function in H(+) extrusion under acid conditions.

Modulation of a thermoregulated type VI secretion system by AHL-dependent quorum sensing in Yersinia pseudotuberculosis.

The type VI secretion system (T6SS) is a novel secretion system found in many Gram-negative bacterial pathogens, which appears to be tightly regulated by different regulatory mechanisms. In the present study, we identified 4 T6SS clusters in Yersinia pseudotuberculosis and demonstrated that they were differentially thermoregulated. Among them, T6SS4 was preferentially expressed at 26°C, and its expression was growth phase dependent and subject to quorum sensing regulation. Both YpsI and YtbI AHL synthases contributed to the positive regulation of T6SS4, whereas YpsI synthase played the major role as T6SS4 expression was reduced strongly in the ypsI mutant strain but weakly in the ytbI mutant strain. Moreover, we provided evidence that exogenous addition of different synthetic AHLs complemented T6SS4 expression in different efficiencies in an ypsIytbI double mutant strain, suggesting C6-HSL had an antagonistic effect on T6SS4 expression. This is the first study demonstrating that the expression of T6SS is precisely regulated by temperature, growth phase, and AHL-dependent quorum sensing systems in Y. pseudotuberculosis.

Heterogeneous UV/Fenton catalytic degradation of wastewater containing phenol with Fe-Cu-Mn-Y catalyst.

The heterogeneous UV/Fenton process with the appropriate amount of Fe-Mn-Cu-Y as catalyst was developed and various operation conditions for the degradation of phenol were evaluated. The results indicated that by using the heterogeneous UV/Fenton process, the COD(cr) removal rate reached almost 100% for wastewater containing phenol. Compared with the homogeneous process, the developed catalyst could be used at wider pH range in the UV/Fenton process. Comparison of various heterogeneous process showed that heterogeneous UV/Fenton process was best. The heterogeneous UV/Fenton process with Fe-Mn-Cu-Y catalyst is highly efficient in degrading various organic pollutants.