Identification and Characterization of the HicAB Toxin-Antitoxin System in the Opportunistic Pathogen Pseudomonas aeruginosa
Abstract
:1. Introduction
2. Results
2.1. Identification and Prevalence of the hicAB Locus in P. aeruginosa
2.2. Genetic Organization and Transcriptional Analysis of the hicAB Locus
2.3. Ectopic Production of HicA Induces Growth Arrest of E. coli, which Can Be Alleviated by HicB
2.4. Overexpression of HicA Confers Cell Stasis, which Can Be Rescued by Subsequently Induced HicB
2.5. Deletion of hicAB Has No Effect on the Biofilm Formation and Virulence of P. aeruginosa
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains, Plasmids and Growth Conditions
4.2. Blast Alignment and Bioinformatics Analysis
4.3. DNA Extraction, RNA Purification and RT-PCR
4.4. E. coli Growth Assay
4.5. Construction of hicAB Deletion Mutant
4.6. Biofilm Formation Assay
4.7. Mouse Infection Experiment
4.8. Statistical Analysis
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Strains/plasmids | Characteristics a | Source/reference |
---|---|---|
Strains | ||
P. aeruginosa strains PA1 | A multi-drug-resistant strain isolated from a patient with respiratory tract infection | Lab collection [22] |
PA1∆HicAB | PA1 derivative with the hicAB locus replaced by a Gmr gene cassette; Gmr | This work |
E. coli strains DH5α | Cloning host for maintaining recombinant plasmids | Tiangen |
BL21(DE3)/pLysS | Expression host for exogenous protein production | Tiangen |
Plasmids | ||
pJS298 | A selective expression vector containing the PBAD and PT7 promoters in trans; Kanr | [27] |
pJSHicB | Derivative of pJS298 containing the hicB gene under the PT7 promoter; Kanr | This work |
pJSHicAB | Derivative of pJSHicB containing the hicA gene under the PBAD promoter; Kanr | This work |
pUCP24 | pUC18-based broad host-range vector; Gmr | Lab collection [44] |
pEX18Tc | Gene replacement vector; Tetr | Lab collection [44] |
pEX∆HicAB | Derivative of pEX18Tc designed for knockout of the hicAB locus; Gmr, Tetr | This work |
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Li, G.; Shen, M.; Lu, S.; Le, S.; Tan, Y.; Wang, J.; Zhao, X.; Shen, W.; Guo, K.; Yang, Y.; et al. Identification and Characterization of the HicAB Toxin-Antitoxin System in the Opportunistic Pathogen Pseudomonas aeruginosa. Toxins 2016, 8, 113. https://doi.org/10.3390/toxins8040113
Li G, Shen M, Lu S, Le S, Tan Y, Wang J, Zhao X, Shen W, Guo K, Yang Y, et al. Identification and Characterization of the HicAB Toxin-Antitoxin System in the Opportunistic Pathogen Pseudomonas aeruginosa. Toxins. 2016; 8(4):113. https://doi.org/10.3390/toxins8040113
Chicago/Turabian StyleLi, Gang, Mengyu Shen, Shuguang Lu, Shuai Le, Yinling Tan, Jing Wang, Xia Zhao, Wei Shen, Keke Guo, Yuhui Yang, and et al. 2016. "Identification and Characterization of the HicAB Toxin-Antitoxin System in the Opportunistic Pathogen Pseudomonas aeruginosa" Toxins 8, no. 4: 113. https://doi.org/10.3390/toxins8040113