Vibrio cholerae is a waterborne bacterium responsible for worldwide outbreaks of acute and fatal cholera. Recently, small regulatory RNAs (sRNAs) have become increasingly recognized as important regulators of virulence gene expression in response to environmental signals. In this study, we determined that two-component system EnvZ/OmpR was required for intestinal colonization in V. cholerae O1 EI Tor strain E12382. Analysis of the characteristics of OmpR revealed a potential binding site in the intergenic region between vc1470 and vc1471, and qRT-PCR showed that expression of the intergenic region increased 5.3-fold in the small intestine compared to LB medium. Race and northern blot assays were performed and demonstrated a new sRNA, coaR (cholerae osmolarity and acidity related regulatory RNA). A ΔcoaR mutant showed a deficient colonization ability in small intestine with CI of 0.15. We identified a target of coaR, tcpI, a negative regulator of the major pilin subunit of TcpA. The ΔtcpI mutant has an increased colonization with CI of 3.16. The expression of coaR increased 2.8-fold and 3.3-fold under relative acidic and hypertonic condition. In summary, coaR was induced under the condition of high osmolarity and acid stress via EnvZ/OmpR and explained that tcpI relieves pH-mediated repression of toxin coregulated pilus synthesis.
Background
Vibrio cholerae, a Gram-negative bacterium, is highly motile owing to the presence of a single polar flagellum. The global anaerobiosis response regulator, ArcA regulates the expression of virulence factors and enhance biofilm formation in V. cholerae. However, the function of ArcA for the motility of V. cholerae is yet to be elucidated. CytR, which represses nucleoside uptake and catabolism, is known to play a chief role in V. cholerae pathogenesis and flagellar synthesis but the mechanism that CytR influences motility is unclear.
Results
In this study, we found that the ΔarcA mutant strain exhibited higher motility than the WT strain due to ArcA directly repressed flrA expression. We further discovered that CytR directly enhanced fliK expression, which explained why the ΔcytR mutant strain was retarded in motility. On the other hand, cytR was a direct ArcA target and cytR expression was directly repressed by ArcA. As expected, cytR expression was down-regulated.
Conclusions
Overall, ArcA plays a critical role in V. cholerae motility by regulating flrA expression directly and fliK indirectly in the manner of cytR.
Background
The anoxic redox control binary system plays an important role in the response to oxygen as a signal in the environment. In particular, phosphorylated ArcA, as a global transcription factor, binds to the promoter regions of its target genes to regulate the expression of aerobic and anaerobic metabolism genes. However, the function of ArcA in Plesiomonas shigelloides is unknown.
Results
In the present study, P. shigelloides was used as the research object. The differences in growth, motility, biofilm formation, and virulence between the WT strain and the ΔarcA isogenic deletion mutant strain were compared. The data showed that the absence of arcA not only caused growth retardation of P. shigelloides in the log phase, but also greatly reduced the glucose utilization in M9 medium before the stationary phase. The motility of the ΔarcA mutant strain was either greatly reduced when grown in swim agar, or basically lost when grown in swarm agar. The electrophoretic mobility shift assay results showed that ArcA bound to the promoter regions of the flaK, rpoN, and cheV genes, indicating that ArcA directly regulates the expression of these three motility-related genes in P. shigelloides. Meanwhile, the ability of the ΔarcA strain to infect Caco-2 cells was reduced by 40%; on the contrary, its biofilm formation was enhanced. Furthermore, the complementation of the WT arcA gene from pBAD33-arcA+ was constructed and all of the above features of the pBAD33-arcA+ complemented strain were restored to the WT level.
Conclusions
We showed the effect of ArcA on the growth, motility, biofilm formation, and virulence of Plesiomonas shigelloides, and demonstrated that ArcA functions as a positive regulator controls the motility of P. shigelloides by directly regulating the expression of flaK, rpoN and cheV genes.
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