33 results on '"Jin, Shouguang"'
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2. Transcript Profiling of Nitroxoline-Treated Biofilms Shows Rapid Up-regulation of Iron Acquisition Gene Clusters.
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Liu, Ke, Abouelhassan, Yasmeen, Zhang, Yanping, Jin, Shouguang, and Huigens III, Robert W.
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- 2022
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3. Modular Synthetic Routes to Fluorine-Containing Halogenated Phenazine and Acridine Agents That Induce Rapid Iron Starvation in Methicillin-Resistant Staphylococcus aureus Biofilms.
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Liu, Ke, Brivio, Massimiliano, Xiao, Tao, Kim, Young S., Jin, Shouguang, Papagni, Antonio, and Vaghi, Luca
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- 2022
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4. A Modular Synthetic Route Involving N‑Aryl-2-nitrosoaniline Intermediates Leads to a New Series of 3‑Substituted Halogenated Phenazine Antibacterial Agents.
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Yang, Hongfen, Kundra, Shivani, Chojnacki, Michaelle, Liu, Ke, Fuse, Marisa A., Abouelhassan, Yasmeen, Kallifidas, Dimitris, Zhang, Peilan, Huang, Guangtao, Jin, Shouguang, Ding, Yousong, Luesch, Hendrik, Rohde, Kyle H., Dunman, Paul M., Lemos, José A., and Huigens III, Robert W.
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- 2021
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5. A Rapid Phenotypic Whole-Cell Screening Approach for the Identification of Small-Molecule Inhibitors That Counter β-Lactamase Resistance in Pseudomonas aeruginosa
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Collia, Deanna, Bannister, Thomas D., Tan, Hao, Jin, Shouguang, Langaee, Taimour, Shumate, Justin, Scampavia, Louis, and Spicer, Timothy P.
- Abstract
Pseudomonas aeruginosais an opportunistic human pathogen that is prevalent in hospitals and continues to develop resistance to multiple classes of antibiotics. Historically, β-lactam antibiotics have been the first line of therapeutic defense. However, the emergence of multidrug-resistant (MDR) strains of P. aeruginosa, such as AmpC β-lactamase overproducing mutants, limits the effectiveness of current antibiotics. Among AmpC hyperproducing clinical isolates, inactivation of AmpG, which is essential for the expression of AmpC, increases bacterial sensitivity to β-lactam antibiotics. We hypothesize that inhibition of AmpG activity will enhance the efficacy of β-lactams against P. aeruginosa. Here, using a highly drug-resistant AmpC-inducible laboratory strain PAO1, we describe an ultra-high-throughput whole-cell turbidity assay designed to identify small-molecule inhibitors of the AmpG. We screened 645,000 compounds to identify compounds with the ability to inhibit bacterial growth in the presence of cefoxitin, an AmpC inducer, and identified 2663 inhibitors that were also tested in the absence of cefoxitin to determine AmpG specificity. The Z′ and signal-to-background ratio were robust at 0.87 ± 0.05 and 2.2 ± 0.2, respectively. Through a series of secondary and tertiary studies, including a novel luciferase-based counterscreen, we ultimately identified eight potential AmpG-specific inhibitors.
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- 2018
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6. A Rapid Phenotypic Whole-Cell Screening Approach for the Identification of Small-Molecule Inhibitors That Counter β-Lactamase Resistance in Pseudomonas aeruginosa
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Collia, Deanna, Bannister, Thomas D., Tan, Hao, Jin, Shouguang, Langaee, Taimour, Shumate, Justin, Scampavia, Louis, and Spicer, Timothy P.
- Abstract
Pseudomonas aeruginosais an opportunistic human pathogen that is prevalent in hospitals and continues to develop resistance to multiple classes of antibiotics. Historically, β-lactam antibiotics have been the first line of therapeutic defense. However, the emergence of multidrug-resistant (MDR) strains of P. aeruginosa, such as AmpC β-lactamase overproducing mutants, limits the effectiveness of current antibiotics. Among AmpC hyperproducing clinical isolates, inactivation of AmpG, which is essential for the expression of AmpC, increases bacterial sensitivity to β-lactam antibiotics. We hypothesize that inhibition of AmpG activity will enhance the efficacy of β-lactams against P. aeruginosa. Here, using a highly drug-resistant AmpC-inducible laboratory strain PAO1, we describe an ultra-high-throughput whole-cell turbidity assay designed to identify small-molecule inhibitors of the AmpG. We screened 645,000 compounds to identify compounds with the ability to inhibit bacterial growth in the presence of cefoxitin, an AmpC inducer, and identified 2663 inhibitors that were also tested in the absence of cefoxitin to determine AmpG specificity. The Z′ and signal-to-background ratio were robust at 0.87 ± 0.05 and 2.2 ± 0.2, respectively. Through a series of secondary and tertiary studies, including a novel luciferase-based counterscreen, we ultimately identified eight potential AmpG-specific inhibitors.
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- 2018
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7. Genome Modification Leads to Phenotype Reversal in Human Myotonic Dystrophy Type 1 Induced Pluripotent Stem Cell‐Derived Neural Stem Cells
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Xia, Guangbin, Gao, Yuanzheng, Jin, Shouguang, Subramony, S.H., Terada, Naohiro, Ranum, Laura P.W., Swanson, Maurice S., and Ashizawa, Tetsuo
- Abstract
Myotonic dystrophy type 1 (DM1) is caused by expanded CTG repeats in the 3'‐untranslated region (3′ UTR) of the DMPKgene. Correcting the mutation in DM1 stem cells would be an important step toward autologous stem cell therapy. The objective of this study is to demonstrate in vitro genome editing to prevent production of toxic mutant transcripts and reverse phenotypes in DM1 stem cells. Genome editing was performed in DM1 neural stem cells (NSCs) derived from human DM1 induced pluripotent stem (iPS) cells. An editing cassette containing SV40/bGH polyA signals was integrated upstream of the CTG repeats by TALEN‐mediated homologous recombination (HR). The expression of mutant CUG repeats transcript was monitored by nuclear RNA foci, the molecular hallmarks of DM1, using RNA fluorescence in situ hybridization. Alternative splicing of microtubule‐associated protein tau (MAPT) and muscleblind‐like (MBNL) proteins were analyzed to further monitor the phenotype reversal after genome modification. The cassette was successfully inserted into DMPKintron 9 and this genomic modification led to complete disappearance of nuclear RNA foci. MAPTand MBNL1, 2 aberrant splicing in DM1 NSCs were reversed to normal pattern in genome‐modified NSCs. Genome modification by integration of exogenous polyA signals upstream of the DMPKCTG repeat expansion prevents the production of toxic RNA and leads to phenotype reversal in human DM1 iPS‐cells derived stem cells. Our data provide proof‐of‐principle evidence that genome modification may be used to generate genetically modified progenitor cells as a first step toward autologous cell transfer therapy for DM1. StemCells2015;33:1829–1838
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- 2015
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8. Distinct Roles of Major Peptidoglycan Recycling Enzymes in β-Lactamase Production in Shewanella oneidensis
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Yin, Jianhua, Mao, Yinting, Ju, Lili, Jin, Miao, Sun, Yiyang, Jin, Shouguang, and Gao, Haichun
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ABSTRACTβ-Lactam antibiotics were the earliest discovered and are the most widely used group of antibiotics that work by inactivating penicillin-binding proteins to inhibit peptidoglycan biosynthesis. As one of the most efficient defense strategies, many bacteria produce β-lactam-degrading enzymes, β-lactamases, whose biochemical functions and regulation have been extensively studied. A signal transduction pathway for β-lactamase induction by β-lactam antibiotics, consisting of the major peptidoglycan recycling enzymes and the LysR-type transcriptional regulator, AmpR, has been recently unveiled in some bacteria. Because inactivation of some of these proteins, especially the permease AmpG and the β-hexosaminidase NagZ, results in substantially elevated susceptibility to the antibiotics, these have been recognized as potential therapeutic targets. Here, we show a contrasting scenario in Shewanella oneidensis, in which the homologue of AmpR is absent. Loss of AmpG or NagZ enhances β-lactam resistance drastically, whereas other identified major peptidoglycan recycling enzymes are dispensable. Moreover, our data indicate that there exists a parallel signal transduction pathway for β-lactamase induction, which is independent of either AmpG or NagZ.
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- 2014
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9. Nucleoside Diphosphate Kinase and Flagellin from Pseudomonas aeruginosaInduce Interleukin 1 Expression via the Akt/NF-?B Signaling Pathways
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Kim, Yong-Jae, Lee, Jung-Hoon, Lee, Yeji, Jia, Jingyue, Paek, Se-Hwan, Kim, Hyong-Bai, Jin, Shouguang, and Ha, Un-Hwan
- Abstract
ABSTRACTInflammatory responses are a first line of host defense against a range of invading pathogens, consisting of the release of proinflammatory cytokines followed by attraction of polymorphonuclear neutrophils (PMNs) to the site of inflammation. Among the many virulence factors that contribute to the pathogenesis of infections, nucleoside diphosphate kinase (Ndk) mediates bacterially induced toxicity against eukaryotic cells. However, no study has examined how Ndk affects inflammatory responses. The present study examined the mechanisms by which Pseudomonas aeruginosaactivates inflammatory responses upon infection of cells. The results showed that bacterial Ndk, with the aid of an additional bacterial factor, flagellin, induced expression of the proinflammatory cytokines interleukin-1a (IL-1a) and IL-1ß. Cytokine induction appeared to be dependent on the kinase activity of Ndk and was mediated via the NF-?B signaling pathway. Notably, Ndk activated the Akt signaling pathway, which acts upstream of NF-?B, as well as caspase-1, which is a key component of inflammasome. Thus, this study demonstrated that P. aeruginosa, through the combined effects of Ndk and flagellin, upregulates the expression of proinflammatory cytokines via the Akt/NF-?B signaling pathways.
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- 2014
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10. PrtR Homeostasis Contributes to Pseudomonas aeruginosaPathogenesis and Resistance against Ciprofloxacin
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Sun, Ziyu, Shi, Jing, Liu, Chang, Jin, Yongxin, Li, Kewei, Chen, Ronghao, Jin, Shouguang, and Wu, Weihui
- Abstract
ABSTRACTPseudomonas aeruginosais an opportunistic pathogen that causes acute and chronic infections in humans. Pyocins are bacteriocins produced by P. aeruginosathat are usually released through lysis of the producer strains. Expression of pyocin genes is negatively regulated by PrtR, which gets cleaved under SOS response, leading to upregulation of pyocin synthetic genes. Previously, we demonstrated that PrtR is required for the expression of type III secretion system (T3SS), which is an important virulence component of P. aeruginosa. In this study, we demonstrate that mutation in prtRresults in reduced bacterial colonization in a mouse acute pneumonia model. Examination of bacterial and host cells in the bronchoalveolar lavage fluids from infected mice revealed that expression of PrtR is induced by reactive oxygen species (ROS) released by neutrophils. We further demonstrate that treatment with hydrogen peroxide or ciprofloxacin, known to induce the SOS response and pyocin production, resulted in an elevated PrtR mRNA level. Overexpression of PrtR by a tacpromoter repressed the endogenous prtRpromoter activity, and electrophoretic mobility shift assay revealed that PrtR binds to its own promoter, suggesting an autorepressive mechanism of regulation. A high level of PrtR expressed from a plasmid resulted in increased T3SS gene expression during infection and higher resistance against ciprofloxacin. Overall, our results suggest that the autorepression of PrtR contributes to the maintenance of a relatively stable level of PrtR, which is permissive to T3SS gene expression in the presence of ROS while increasing bacterial tolerance to stresses, such as ciprofloxacin, by limiting pyocin production.
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- 2014
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11. ampG Gene of Pseudomonas aeruginosa and Its Role in β-Lactamase Expression
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Zhang, Ying, Bao, Qiyu, Gagnon, Luc A., Huletsky, Ann, Oliver, Antonio, Jin, Shouguang, and Langaee, Taimour
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In enterobacteria, the ampG gene encodes a transmembrane protein (permease) that transports 1,6-GlcNAc-anhydro-MurNAc and the 1,6-GlcNAc-anhydro-MurNAc peptide from the periplasm to the cytoplasm, which serve as signal molecules for the induction of ampC β-lactamase. The role of AmpG as a transporter is also essential for cell wall recycling. Pseudomonas aeruginosa carries two AmpG homologues, AmpG (PA4393) and AmpGh1 (PA4218), with 45 and 41% amino acid sequence identity, respectively, to Escherichia coli AmpG, while the two homologues share only 19% amino acid identity. In P. aeruginosa strains PAO1 and PAK, inactivation of ampG drastically repressed the intrinsic β-lactam resistance while ampGh1 deletion had little effect on the resistance. Further, deletion of ampG in an ampD-null mutant abolished the high-level β-lactam resistance that is associated with the loss of AmpD activity. The cloned ampG gene is able to complement both the P. aeruginosa and the E. coli ampG mutants, while that of ampGh1 failed to do so, suggesting that PA4393 encodes the only functional AmpG protein in P. aeruginosa. We also demonstrate that the function of AmpG in laboratory strains of P. aeruginosa can effectively be inhibited by carbonyl cyanide m-chlorophenylhydrazone (CCCP), causing an increased sensitivity to β-lactams among laboratory as well as clinical isolates of P. aeruginosa. Our results suggest that inhibition of the AmpG activity is a potential strategy for enhancing the efficacy of β-lactams against P. aeruginosa, which carries inducible chromosomal ampC, especially in AmpC-hyperproducing clinical isolates.
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- 2010
12. ampGGene of Pseudomonas aeruginosaand Its Role in β-Lactamase Expression
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Zhang, Ying, Bao, Qiyu, Gagnon, Luc A., Huletsky, Ann, Oliver, Antonio, Jin, Shouguang, and Langaee, Taimour
- Abstract
ABSTRACTIn enterobacteria, the ampGgene encodes a transmembrane protein (permease) that transports 1,6-GlcNAc-anhydro-MurNAc and the 1,6-GlcNAc-anhydro-MurNAc peptide from the periplasm to the cytoplasm, which serve as signal molecules for the induction of ampCβ-lactamase. The role of AmpG as a transporter is also essential for cell wall recycling. Pseudomonas aeruginosacarries two AmpG homologues, AmpG (PA4393) and AmpGh1 (PA4218), with 45 and 41% amino acid sequence identity, respectively, to Escherichia coliAmpG, while the two homologues share only 19% amino acid identity. In P. aeruginosastrains PAO1 and PAK, inactivation of ampGdrastically repressed the intrinsic β-lactam resistance while ampGh1deletion had little effect on the resistance. Further, deletion of ampGin an ampD-null mutant abolished the high-level β-lactam resistance that is associated with the loss of AmpD activity. The cloned ampGgene is able to complement both the P. aeruginosaand the E. coli ampGmutants, while that of ampGh1failed to do so, suggesting that PA4393 encodes the only functional AmpG protein in P. aeruginosa. We also demonstrate that the function of AmpG in laboratory strains of P. aeruginosacan effectively be inhibited by carbonyl cyanide m-chlorophenylhydrazone (CCCP), causing an increased sensitivity to β-lactams among laboratory as well as clinical isolates of P. aeruginosa. Our results suggest that inhibition of the AmpG activity is a potential strategy for enhancing the efficacy of β-lactams against P. aeruginosa, which carries inducible chromosomal ampC, especially in AmpC-hyperproducing clinical isolates.
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- 2010
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13. Expression of Pseudomonas aeruginosa Toxin ExoS Effectively Induces Apoptosis in Host Cells
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Jia, Jinghua, Wang, Yanping, Zhou, Lei, and Jin, Shouguang
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Pseudomonas aeruginosa is an opportunistic bacterial pathogen that primarily infects immunocompromised individuals and patients with cystic fibrosis. Invasive strains of P. aeruginosa are known to induce apoptosis at a high frequency in HeLa cells and in many other cell lines, a process that is dependent on the ADP-ribosylation (ADPRT) activity of a type III secreted protein ExoS. In our previous report, it was proposed that P. aeruginosa secreting ExoS, upon infection, shuts down host cell survival signal pathways by inhibiting ERK1/2 and p38 activation, and it activates proapoptotic pathways through activation of JNK1/2, leading ultimately to cytochrome c release and activation of caspases. In this study, we demonstrate that the expression of ExoS in HeLa cells by eukaryotic expression vector effectively caused apoptosis in an ADPRT activity-dependent manner, indicating that ExoS alone is sufficient to trigger apoptotic death of host cells independent of any other bacterial factors. By expressing an EGFP-ExoS fusion protein, we were able to directly correlate the death of HeLa cells with the presence of intracellular ExoS and further proved the dependence of this process on both JNK activation and mitochondrial proapoptotic event. The cellular pathway responsible for the ExoS-induced cytotoxicity appears to be well conserved, since the expression of the ADPRT-competent ExoS also induced rapid cell death in the Drosophila melanogaster S2 cell lines. The presented study not only highlights the ability of ExoS ADPRT to modulate host cell signaling, eventually leading to apoptosis, but also establishes ExoS as a valuable tool, in principle, for the elucidation of apoptosis mechanisms.
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- 2006
14. Expression of Pseudomonas aeruginosaToxin ExoS Effectively Induces Apoptosis in Host Cells
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Jia, Jinghua, Wang, Yanping, Zhou, Lei, and Jin, Shouguang
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ABSTRACTPseudomonas aeruginosais an opportunistic bacterial pathogen that primarily infects immunocompromised individuals and patients with cystic fibrosis. Invasive strains of P. aeruginosaare known to induce apoptosis at a high frequency in HeLa cells and in many other cell lines, a process that is dependent on the ADP-ribosylation (ADPRT) activity of a type III secreted protein ExoS. In our previous report, it was proposed that P. aeruginosasecreting ExoS, upon infection, shuts down host cell survival signal pathways by inhibiting ERK1/2 and p38 activation, and it activates proapoptotic pathways through activation of JNK1/2, leading ultimately to cytochrome crelease and activation of caspases. In this study, we demonstrate that the expression of ExoS in HeLa cells by eukaryotic expression vector effectively caused apoptosis in an ADPRT activity-dependent manner, indicating that ExoS alone is sufficient to trigger apoptotic death of host cells independent of any other bacterial factors. By expressing an EGFP-ExoS fusion protein, we were able to directly correlate the death of HeLa cells with the presence of intracellular ExoS and further proved the dependence of this process on both JNK activation and mitochondrial proapoptotic event. The cellular pathway responsible for the ExoS-induced cytotoxicity appears to be well conserved, since the expression of the ADPRT-competent ExoS also induced rapid cell death in the Drosophila melanogasterS2 cell lines. The presented study not only highlights the ability of ExoS ADPRT to modulate host cell signaling, eventually leading to apoptosis, but also establishes ExoS as a valuable tool, in principle, for the elucidation of apoptosis mechanisms.
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- 2006
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15. Use of DNA fingerprinting in decision making for considering closure of neonatal intensive care units because of Pseudomonas aeruginosabloodstream infections
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SCHUTZE, GORDON E., GILLIAM, CRAIG H., JIN, SHOUGUANG, CAVENAUGH, CONNIE K., HALL, R. WHIT, BRADSHER, ROBERT W., and JACOBS, RICHARD F.
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Bloodstream infections with Pseudomonas aeruginosahave been well-described in neonatal intensive care units (NICU) and have resulted in the temporary closure of some nurseries to new admissions. Nosocomial transmission of these infections has been verified by fingerprint analysis of the isolates. We utilized molecular fingerprinting to identify the source of bloodstream infections in an NICU and used this information to apply infection control measures that allowed the nursery to stay open and continue to accept referrals.
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- 2004
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16. aph(3')-IIb, a gene encoding an aminoglycoside-modifying enzyme, is under the positive control of surrogate regulator HpaA.
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Zeng, Lin and Jin, Shouguang
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Pseudomonas aeruginosa harbors a chromosomal aminoglycoside phosphotransferase gene, aph(3')-IIb, which confers P. aeruginosa resistance to several important aminoglycoside antibiotics, including kanamycin A and B, neomycin B and C, butirosin, and seldomycin F5. The aph(3')-IIb gene has been found to be regulated by an AraC-type transcriptional regulator (HpaA) encoded by a gene located upstream of the aph(3')-IIb gene. In the presence of 4-hydroxyphenylacetic acid (4-HPA), HpaA activates the expression of aph(3')-IIb as well as that of the hpa regulon which encodes metabolic enzymes for the utilization of 4-HPA. hpaA and aph(3')-IIb form an operon, and in response to the presence of 4-HPA, the wild-type P. aeruginosa strain PAK (but not its hpaA mutant strain) displays increased resistance to neomycin. A survey of 39 clinical and 19 environmental isolates of P. aeruginosa demonstrated in all of them the presence of an hpaA-aph gene cluster, while 56 out of the 58 isolates are able to utilize the 4-HPA as a sole carbon source, suggesting a feature common to P. aeruginosa strains. Interestingly, a larger portion of clinical isolates than environmental isolates showed 4-HPA-induced resistance to neomycin. The aph(3')-IIb gene product is likely to function as a metabolic enzyme which has a cross-reactivity with aminoglycosides. These findings provide new insight into the possible mechanism of P. aeruginosa antibiotic resistance.
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- 2003
17. aph(3′)-IIb, a Gene Encoding an Aminoglycoside-Modifying Enzyme, Is under the Positive Control of Surrogate Regulator HpaA
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Zeng, Lin and Jin, Shouguang
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ABSTRACTPseudomonas aeruginosaharbors a chromosomal aminoglycoside phosphotransferase gene, aph(3′)-IIb, which confers P. aeruginosaresistance to several important aminoglycoside antibiotics, including kanamycin A and B, neomycin B and C, butirosin, and seldomycin F5. The aph(3′)-IIbgene has been found to be regulated by an AraC-type transcriptional regulator (HpaA) encoded by a gene located upstream of the aph(3′)-IIbgene. In the presence of 4-hydroxyphenylacetic acid (4-HPA), HpaA activates the expression of aph(3′)-IIbas well as that of the hparegulon which encodes metabolic enzymes for the utilization of 4-HPA. hpaAand aph(3′)-IIbform an operon, and in response to the presence of 4-HPA, the wild-type P. aeruginosastrain PAK (but not its hpaAmutant strain) displays increased resistance to neomycin. A survey of 39 clinical and 19 environmental isolates of P. aeruginosademonstrated in all of them the presence of an hpaA-aphgene cluster, while 56 out of the 58 isolates are able to utilize the 4-HPA as a sole carbon source, suggesting a feature common to P. aeruginosastrains. Interestingly, a larger portion of clinical isolates than environmental isolates showed 4-HPA-induced resistance to neomycin. The aph(3′)-IIbgene product is likely to function as a metabolic enzyme which has a cross-reactivity with aminoglycosides. These findings provide new insight into the possible mechanism of P. aeruginosaantibiotic resistance.
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- 2003
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18. c-Jun NH2-terminal kinase-mediated signaling is essential for Pseudomonas aeruginosa ExoS-induced apoptosis.
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Jia, Jinghua, Alaoui-El-Azher, Mounia, Chow, Marie, Chambers, Timothy C, Baker, Henry, and Jin, Shouguang
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As an opportunistic bacterial pathogen, Pseudomonas aeruginosa mainly affects immunocompromised individuals as well as patients with cystic fibrosis. In a previous study, we showed that ExoS of P. aeruginosa, when injected into host cells through a type III secretion apparatus, functions as an effector molecule to trigger apoptosis in various tissue culture cells. Here, we show that injection of the ExoS into HeLa cells activates c-Jun NH(2)-terminal kinase (JNK) phosphorylation while shutting down ERK1/2 and p38 phosphorylation. Inhibiting JNK activation by expression of a dominant negative JNK1 or with a specific JNK inhibitor abolishes ExoS-triggered apoptosis, demonstrating the requirement for JNK-mediated signaling. Following JNK phosphorylation, cytochrome c is released into the cytosol, leading to the activation of caspase 9 and eventually caspase 3. Although c-Jun phosphorylation is also observed as a result of JNK activation, ongoing host protein synthesis is not essential for the apoptotic induction, suggesting that c-Jun- or other AP-1-driven activation of gene expression is dispensable in this process. Therefore, ExoS has opposing effects on different cellular pathways that regulate apoptosis: it shuts down host cell survival signal pathways by inhibiting ERK1/2 and p38 activation, and it activates proapoptotic pathways through activation of JNK1/2 leading ultimately to cytochrome c release and activation of caspases. These results highlight the modulation of host cell signaling by the type III secretion system during interaction between P. aeruginosa and host cells.
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- 2003
19. c-Jun NH2-Terminal Kinase-Mediated Signaling Is Essential for Pseudomonas aeruginosaExoS-Induced Apoptosis
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Jia, Jinghua, Alaoui-El-Azher, Mounia, Chow, Marie, Chambers, Timothy C., Baker, Henry, and Jin, Shouguang
- Abstract
ABSTRACTAs an opportunistic bacterial pathogen, Pseudomonas aeruginosamainly affects immunocompromised individuals as well as patients with cystic fibrosis. In a previous study, we showed that ExoS of P. aeruginosa, when injected into host cells through a type III secretion apparatus, functions as an effector molecule to trigger apoptosis in various tissue culture cells. Here, we show that injection of the ExoS into HeLa cells activates c-Jun NH2-terminal kinase (JNK) phosphorylation while shutting down ERK1/2 and p38 phosphorylation. Inhibiting JNK activation by expression of a dominant negative JNK1 or with a specific JNK inhibitor abolishes ExoS-triggered apoptosis, demonstrating the requirement for JNK-mediated signaling. Following JNK phosphorylation, cytochrome cis released into the cytosol, leading to the activation of caspase 9 and eventually caspase 3. Although c-Jun phosphorylation is also observed as a result of JNK activation, ongoing host protein synthesis is not essential for the apoptotic induction, suggesting that c-Jun- or other AP-1-driven activation of gene expression is dispensable in this process. Therefore, ExoS has opposing effects on different cellular pathways that regulate apoptosis: it shuts down host cell survival signal pathways by inhibiting ERK1/2 and p38 activation, and it activates proapoptotic pathways through activation of JNK1/2 leading ultimately to cytochrome crelease and activation of caspases. These results highlight the modulation of host cell signaling by the type III secretion system during interaction between P. aeruginosaand host cells.
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- 2003
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20. DsbA of Pseudomonas aeruginosa is essential for multiple virulence factors.
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Ha, Un-Hwan, Wang, Yanping, and Jin, Shouguang
- Abstract
DsbA is a periplasmic thiol:disulfide oxidoreductase which contributes to the process of protein folding by catalyzing the formation of disulfide bonds. In this study, we demonstrate that the dsbA gene is required for the expression of the type III secretion system under low-calcium inducing conditions, intracellular survival of P. aeruginosa upon infection of HeLa cells, and twitching motility. The diverse phenotypes of the dsbA mutant are likely due to its defect in the folding of proteins that are involved in various biological processes, such as signal sensing, protein secretion, and defense against host clearing. In light of its effect on various virulence factors, DsbA could be an important target for the control of P. aeruginosa infections.
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- 2003
21. DsbA of Pseudomonas aeruginosaIs Essential for Multiple Virulence Factors
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Ha, Un-Hwan, Wang, Yanping, and Jin, Shouguang
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ABSTRACTDsbA is a periplasmic thiol:disulfide oxidoreductase which contributes to the process of protein folding by catalyzing the formation of disulfide bonds. In this study, we demonstrate that the dsbAgene is required for the expression of the type III secretion system under low-calcium inducing conditions, intracellular survival of P. aeruginosaupon infection of HeLa cells, and twitching motility. The diverse phenotypes of the dsbAmutant are likely due to its defect in the folding of proteins that are involved in various biological processes, such as signal sensing, protein secretion, and defense against host clearing. In light of its effect on various virulence factors, DsbA could be an important target for the control of P. aeruginosainfections.
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- 2003
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22. Regulation of Membrane Permeability by a Two-Component Regulatory System in Pseudomonas aeruginosa
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Wang, Yanping, Ha, Unhwan, Zeng, Lin, and Jin, Shouguang
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ABSTRACTMembrane impermeability is the major contributing factor to multidrug resistance in clinical isolates of Pseudomonas aeruginosa. By using laboratory strain PAK, a spontaneous P. aeruginosamutant (mutant PAK1-3) whose membrane had reduced permeability and which displayed increased levels of resistance to various antibiotics, especially aminoglycosides, was isolated. By complementation of the mutant with a genomic clone library derived from wild-type strain PAK, a novel two-component regulatory system (PprA and PprB) was identified and was found to be able to increase the permeability of the bacterial membrane and render PAK1-3 sensitive to antibiotics. Furthermore, specific phosphorylation of the response regulator (PprB) by histidine kinase (PprA) was observed in vitro, demonstrating that they are cognate two-component regulatory genes. Introduction of a plasmid expressing the pprBgene into randomly chosen clinical isolates (n= 17) resulted in increased sensitivity to aminoglycosides in the majority of isolates (n= 13) tested. This is the first demonstration that P. aeruginosamembrane permeability can be regulated, providing an important clue in the understanding of the mechanism of membrane impermeability-mediated multidrug resistance in P. aeruginosa.
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- 2003
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23. Growth Phase-Dependent Invasion ofPseudomonas aeruginosaand Its Survival within HeLa Cells
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Ha, Unhwan and Jin, Shouguang
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ABSTRACTClinical isolates of Pseudomonas aeruginosaare classified into invasive and noninvasive (cytolytic) strains. In a noninvasive PA103 background, ExoS and ExoT have recently been shown to function as anti-internalization factors. However, these two factors seemed not to have such a function in an invasive strain PAK background. In this study, using HeLa tissue culture cells, we observed that the internalization of invasive strain PAK is dependent on its growth phases, with the stationary-phase cells internalized about 100-fold more efficiently than the exponential-phase cells. This growth phase-dependent internalization was not observed in the noninvasive PA103 strain. Further analysis of various mutant derivatives of the invasive PAK and the noninvasive PA103 strains demonstrated that ExoS or ExoT that is injected into host cells by a type III secretion machinery functions as an anti-internalization factor in both types of strains. In correlation with the growth phase-dependent internalization, the invasive strain PAK translocates much higher amount of ExoS and ExoT into HeLa cells when it is in an exponential-growth phase than when it is in a stationary-growth phase, whereas the translocation of ExoT by the noninvasive strain PA103 is consistently high regardless of the growth phases, suggesting a difference in the regulatory mechanism of type III secretion between the two types of strains. Consistent with the invasive phenotype of the parent strain, an internalized PAK derivative survived well within the HeLa cells, whereas the viability of internalized PA103 derivative was dramatically decreased and completely cleared within 48 h. These results indicate that the invasive strains of P. aeruginosahave evolved the mechanism of intracellular survival, whereas the noninvasive P. aeruginosastrains have lost or not acquired the ability to survive within the epithelial cells.
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- 2001
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24. Expression of the soxRGene ofPseudomonas aeruginosaIs Inducible during Infection of Burn Wounds in Mice and Is Required To Cause Efficient Bacteremia
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Ha, Unhwan and Jin, Shouguang
- Abstract
ABSTRACTBurn wounds are prone to infection by Pseudomonas aeruginosa, which is an opportunistic pathogen causing various human diseases. During infection, the bacterium senses environmental changes and regulates the expression of genes appropriate for survival. A purine-auxotrophic mutant of P. aeruginosawas unable to replicate efficiently on burn wounds, suggesting that burn wounds are purine-deficient environments. An in vivo expression technology based on purEKgene expression was applied to the burned mouse infection model to isolate P. aeruginosagenes that are specifically induced during infection. Four such in vivo-inducible (ivi) genetic loci were identified, including the gene for a superoxide response regulator (soxR), the gene for a malate synthase G homologue (glcG), an antisense transcript of a putative regulator responding to copper (copR), and an uncharacterized genetic locus. SoxR of Escherichia coliis known to regulate genes involved in protecting the bacterium against oxidative stress. The expression of soxRwas proven to be highly inducible during the infection of burned mice and also inducible by treatment with paraquat, which is a redox-cycling reagent generating intracellular superoxide. The SoxR protein functions as an autorepressor in the absence of paraquat, whereas in the presence of paraquat, this autorepression is diminished. Furthermore, asoxRnull mutant was shown to be much more sensitive than wild-type P. aeruginosato macrophage-mediated killing. In support of this observation, a soxRnull mutant exhibited a significant delay in causing systemic infections in the burned mice. Since most mortality in burn patients is caused by systemic infection, the defect in the ability to cause efficient bacteremia in burned mice suggests an important role of the soxRgene in the infection of burn wounds.
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- 1999
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25. A Modular Synthetic Route Involving N-Aryl-2-nitrosoaniline Intermediates Leads to a New Series of 3-Substituted Halogenated Phenazine Antibacterial Agents
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Yang, Hongfen, Kundra, Shivani, Chojnacki, Michaelle, Liu, Ke, Fuse, Marisa A., Abouelhassan, Yasmeen, Kallifidas, Dimitris, Zhang, Peilan, Huang, Guangtao, Jin, Shouguang, Ding, Yousong, Luesch, Hendrik, Rohde, Kyle H., Dunman, Paul M., Lemos, José A., and Huigens, Robert W.
- Abstract
Pathogenic bacteria demonstrate incredible abilities to evade conventional antibiotics through the development of resistance and formation of dormant, surface-attached biofilms. Therefore, agents that target and eradicate planktonic and biofilm bacteria are of significant interest. We explored a new series of halogenated phenazines (HP) through the use of N-aryl-2-nitrosoaniline synthetic intermediates that enabled functionalization of the 3-position of this scaffold. Several HPs demonstrated potent antibacterial and biofilm-killing activities (e.g., HP 29, against methicillin-resistant Staphylococcus aureus: MIC = 0.075 μM; MBEC = 2.35 μM), and transcriptional analysis revealed that HPs 3, 28, and 29induce rapid iron starvation in MRSA biofilms. Several HPs demonstrated excellent activities against Mycobacterium tuberculosis(HP 34, MIC = 0.80 μM against CDC1551). This work established new SAR insights, and HP 29demonstrated efficacy in dorsal wound infection models in mice. Encouraged by these findings, we believe that HPs could lead to significant advances in the treatment of challenging infections.
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- 2021
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26. TpiA is a Key Metabolic Enzyme That Affects Virulence and Resistance to Aminoglycoside Antibiotics through CrcZ in Pseudomonas aeruginosa
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Xia, Yushan, Wang, Dan, Pan, Xiaolei, Xia, Bin, Weng, Yuding, Long, Yuqing, Ren, Huan, Zhou, Jingyi, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
- Abstract
The increase in bacterial resistance against antibiotics imposes a severe threat to public health. It is urgent to identify new drug targets and develop novel antimicrobials. Metabolic homeostasis of bacteria plays an essential role in their virulence and resistance to antibiotics. Recent studies demonstrated that antibiotic efficacies can be improved by modulating the bacterial metabolism. Pseudomonas aeruginosais an important opportunistic human pathogen that causes various infections. The bacterium is intrinsically resistant to antibiotics. In this study, we provide clear evidence that TpiA (triosephosphate isomerase) plays an essential role in the metabolism of P. aeruginosaand influences bacterial virulence and antibiotic resistance. The significance of this work is in identifying a key enzyme in the metabolic network, which will provide clues as to the development of novel treatment strategies against infections caused by P. aeruginosa.
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- 2020
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27. Pseudomonas aeruginosaExsA Regulates a Metalloprotease, ImpA, That Inhibits Phagocytosis of Macrophages
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Tian, Zhenyang, Cheng, Sen, Xia, Bin, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, Liu, Xiaoyun, and Wu, Weihui
- Abstract
Pseudomonas aeruginosais an opportunistic pathogenic bacterium whose type III secretion system (T3SS) plays a critical role in acute infections. Translocation of the T3SS effectors into host cells induces cytotoxicity. In addition, the T3SS promotes the intracellular growth of P. aeruginosaduring host infections. The T3SS regulon genes are regulated by an AraC-type regulator, ExsA. In this study, we found that an extracellular metalloprotease encoded by impA(PA0572) is under the regulation of ExsA.
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- 2019
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28. Oligoribonuclease Contributes to Tolerance to Aminoglycoside and β-Lactam Antibiotics by Regulating KatA in Pseudomonas aeruginosa
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Xia, Bin, Li, Mei, Tian, Zhenyang, Chen, Gukui, Liu, Chang, Xia, Yushan, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
- Abstract
Pseudomonas aeruginosais an opportunistic bacterial pathogen and is intrinsically resistant to a variety of antibiotics. Oligoribonuclease (Orn) is a 3′-to-5′ exonuclease that degrades nanoRNAs.
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- 2019
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29. PA5470 Counteracts Antimicrobial Effect of Azithromycin by Releasing Stalled Ribosome in Pseudomonas aeruginosa
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Shi, Jing, Liu, Yiwei, Zhang, Yueying, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
- Abstract
ABSTRACTPseudomonas aeruginosacauses various acute and chronic infections in humans. Treatment with azithromycin (AZM) has been shown to benefit patients with chronic P. aeruginosainfections. By binding to the exit tunnel of the 50S ribosome, AZM causes ribosome stalling and depletion of the intracellular tRNA pool. It has been shown that AZM is able to kill stationary-phase P. aeruginosacells and repress quorum sensing-regulated virulence factors as well as swarming motility. In P. aeruginosa, the PA5470 gene encodes a putative peptide chain release factor whose expression is highly induced by macrolide antibiotics. However, its function remains unknown. Here, we found that overexpression of PA5470 increased bacterial tolerance against AZM and alleviated the repression of swarming motility. Ribosome pulldown assays revealed that PA5470 contributes to the release of ribosome stalled by AZM. We further demonstrate that overexpression of PA5470 counteracts AZM-mediated repression of the translation of the quorum sensing regulator RhlR. Overall, our results revealed a novel role of PA5470 in the bacterial response to AZM.
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- 2017
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30. Bacterial Nucleotidyl Cyclase Inhibits the Host Innate Immune Response by Suppressing TAK1 Activation
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He, Chenxi, Zhou, Yilong, Liu, Feng, Liu, Haipeng, Tan, Hao, Jin, Shouguang, Wu, Weihui, and Ge, Baoxue
- Abstract
ABSTRACTExoenzyme Y (ExoY) is a type III secretion system effector found in 90% of the Pseudomonas aeruginosaisolates. Although it is known that ExoY is a soluble nucleotidyl cyclase that increases the cytoplasmic levels of nucleoside 3′,5′-cyclic monophosphates (cNMPs) to mediate endothelial Tau phosphorylation and permeability, its functional role in the innate immune response is still poorly understood. Transforming growth factor β-activated kinase 1 (TAK1) is critical for mediating Toll-like receptor (TLR) signaling and subsequent activation of NF-κB and AP-1, which are transcriptional activators of innate immunity. Here, we report that ExoY inhibits proinflammatory cytokine production through suppressing the activation of TAK1 as well as downstream NF-κB and mitogen-activated protein (MAP) kinases. Mice infected with ExoY-deficient P. aeruginosahad higher levels of tumor necrosis factor (TNF) and interleukin-6 (IL-6), more neutrophil recruitment, and a lower bacterial load in lung tissue than mice infected with wild-type P. aeruginosa. Taken together, our findings identify a previously unknown mechanism by which P. aeruginosaExoY inhibits the host innate immune response.
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- 2017
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31. Pseudomonas aeruginosaOligoribonuclease Contributes to Tolerance to Ciprofloxacin by Regulating Pyocin Biosynthesis
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Chen, Fei, Chen, Gukui, Liu, Yiwei, Jin, Yongxin, Cheng, Zhihui, Liu, Yang, Yang, Liang, Jin, Shouguang, and Wu, Weihui
- Abstract
ABSTRACTBacterial oligoribonuclease (Orn) is a conserved 3′-to-5′ exonuclease. In Pseudomonas aeruginosa, it has been demonstrated that Orn plays a major role in the hydrolysis of pGpG, which is required for cyclic-di-GMP homeostasis. Meanwhile, Orn is involved in the degradation of nanoRNAs, which can alter global gene expression by serving as transcription initiation primers. Previously, we found that Orn is required for the type III secretion system and pathogenesis of P. aeruginosa, indicating a role of Orn in the bacterial response to environmental stimuli. Here we report that Orn is required for the tolerance of P. aeruginosato ciprofloxacin. Transcriptome analysis of an ornmutant revealed the upregulation of pyocin biosynthesis genes. Mutation of genes involved in pyocin biosynthesis in the background of an ornmutant restored bacterial tolerance to ciprofloxacin. We further demonstrate that the upregulation of pyocin biosynthesis genes is due to RecA-mediated autoproteolysis of PrtR, which is the major negative regulator of pyocin biosynthesis genes. In addition, the SOS response genes were upregulated in the ornmutant, indicating a DNA damage stress. Therefore, our results revealed a novel role of Orn in bacterial tolerance to ciprofloxacin.
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- 2017
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32. Pseudomonas aeruginosaGroEL Stimulates Production of PTX3 by Activating the NF-κB Pathway and Simultaneously Downregulating MicroRNA-9
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Shin, Heesung, Jeon, Jisu, Lee, Jung-Hoon, Jin, Shouguang, and Ha, Un-Hwan
- Abstract
ABSTRACTAs one of the first lines of host defense, monocytes play important roles in clearing infected microbes. The defensive response is triggered by recognition of diverse microbial moieties, including released factors, which modulate host immune responses to establish a harsh environment for clinically important bacterial pathogens. In this study, we found that the expression of PTX3, a soluble form of pattern recognition receptor, was induced by infection with live Pseudomonas aeruginosaor treatment of cells with its supernatant. P. aeruginosaGroEL, a homolog of heat shock protein 60, was identified as one of the factors responsible for inducing the expression of PTX3in host cells. GroEL induced PTX3expression by activating the Toll-like receptor 4 (TLR4)-dependent pathway via nuclear factor-kappa B (NF-κB), while simultaneously inhibiting expression of microRNA-9, which targets the PTX3transcript. Finally, by acting as an opsonin, GroEL-induced PTX3 promoted the association and phagocytosis of Staphylococcus aureusinto macrophages. These data suggest that the host defensive environment is supported by the production of PTX3 in response to GroEL, which thus has therapeutic potential for clearance of bacterial infections.
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- 2016
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33. SuhB Is a Regulator of Multiple Virulence Genes and Essential for Pathogenesis of Pseudomonas aeruginosa
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Li, Kewei, Xu, Chang, Jin, Yongxin, Sun, Ziyu, Liu, Chang, Shi, Jing, Chen, Gukui, Chen, Ronghao, Jin, Shouguang, and Wu, Weihui
- Abstract
ABSTRACTDuring initial colonization and chronic infection, pathogenic bacteria encounter distinct host environments. Adjusting gene expression accordingly is essential for the pathogenesis. Pseudomonas aeruginosahas evolved complicated regulatory networks to regulate different sets of virulence factors to facilitate colonization and persistence. The type III secretion system (T3SS) and motility are associated with acute infections, while biofilm formation and the type VI secretion system (T6SS) are associated with chronic persistence. To identify novel regulatory genes required for pathogenesis, we screened a P. aeruginosatransposon (Tn) insertion library and found suhBto be an essential gene for the T3SS gene expression. The expression of suhBwas upregulated in a mouse acute lung infection model, and loss of suhBresulted in avirulence. Suppression of T3SS gene expression in the suhBmutant is linked to a defective translation of the T3SS master regulator, ExsA. Further studies demonstrated that suhBmutation led to the upregulation of GacA and its downstream small RNAs, RsmY and RsmZ, triggering T6SS expression and biofilm formation while inhibiting the T3SS. Our results demonstrate that an in vivo-inducible gene, suhB, reciprocally regulates genes associated with acute and chronic infections and plays an essential role in the pathogenesis of P. aeruginosa.IMPORTANCEA variety of bacterial pathogens, such as Pseudomonas aeruginosa, cause acute and chronic infections in humans. During infections, pathogens produce different sets of virulence genes for colonization, tissue damage, and dissemination and for countering host immune responses. Complex regulatory networks control the delicate tuning of gene expression in response to host environments to enable the survival and growth of invading pathogens. Here we identified suhBas a critical gene for the regulation of virulence factors in P. aeruginosa. The expression of suhBwas upregulated during acute infection in an animal model, and mutation of suhBrendered P. aeruginosaavirulent. Moreover, we demonstrate that SuhB is required for the activation of virulence factors associated with acute infections while suppressing virulence factors associated with chronic infections. Our report provides new insights into the multilayered regulatory network of virulence genes in P. aeruginosa.
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- 2013
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