452 results on '"Jin, Shouguang"'
Search Results
152. aph(3′)-IIb , a Gene Encoding anAminoglycoside-Modifying Enzyme, Is under the Positive Control ofSurrogate RegulatorHpaA
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Zeng, Lin, primary and Jin, Shouguang, additional
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- 2003
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153. c-Jun NH2-Terminal Kinase-Mediated Signaling Is Essential forPseudomonas aeruginosaExoS-Induced Apoptosis
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Jia, Jinghua, primary, Alaoui-El-Azher, Mounia, additional, Chow, Marie, additional, Chambers, Timothy C., additional, Baker, Henry, additional, and Jin, Shouguang, additional
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- 2003
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154. DsbA of Pseudomonas aeruginosa Is Essential for Multiple Virulence Factors
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Ha, Un-Hwan, primary, Wang, Yanping, additional, and Jin, Shouguang, additional
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- 2003
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155. Regulation of Membrane Permeability by a Two-Component Regulatory System in Pseudomonas aeruginosa
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Wang, Yanping, primary, Ha, Unhwan, additional, Zeng, Lin, additional, and Jin, Shouguang, additional
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- 2003
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156. Growth Phase-Dependent Invasion of Pseudomonas aeruginosa and Its Survival within HeLa Cells
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Ha, Unhwan, primary and Jin, Shouguang, additional
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- 2001
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157. migA, a quorum-responsive gene of Pseudomonas aeruginosa, is highly expressed in the cystic fibrosis lung environment and modifies low-molecular-mass lipopolysaccharide
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Yang, Hongjiang, primary, Matewish, Mauricia, additional, Loubens, Isabelle, additional, Storey, Douglas G., additional, Lam, Joseph S., additional, and Jin, Shouguang, additional
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- 2000
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158. Pseudomonas aeruginosa mediated apoptosis requires the ADP-ribosylating activity of ExoS
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Kaufman, Melissa R., primary, Jia, Jinghua, additional, Zeng, Lin, additional, Ha, Unhwan, additional, Chow, Marie, additional, and Jin, Shouguang, additional
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- 2000
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159. Expression of the soxR Gene of Pseudomonas aeruginosa Is Inducible during Infection of Burn Wounds in Mice and Is Required To Cause Efficient Bacteremia
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Ha, Unhwan, primary and Jin, Shouguang, additional
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- 1999
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160. The sensing of plant signal molecules by Agrobacterium: genetic evidence for direct recognition of phenolic inducers by the VirA protein
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Lee, Yong-Woog, primary, Jin, Shouguang, additional, Sim, Woong-Seop, additional, and Nester, Eugene W., additional
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- 1996
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161. An Agrobacterium virulence factor encoded by a Ti plasmid gene or a chromosomal gene is required for T-DNA transfer into plants
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Pan, Shen Q., primary, Jin, Shouguang, additional, Boulton, Margaret I., additional, Hawes, Martha, additional, Gordon, Milton P., additional, and Nester, Eugene W., additional
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- 1995
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162. PilR, a transcriptional regulator of piliation in Pseudomonas aeruginosa, binds to a cis‐acting sequence upstream of the pilin gene promoter
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Jin, Shouguang, primary, Ishimoto, Karyn S., additional, and Lory, Stephen, additional
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- 1994
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163. Characterization of a virG mutation that confers constitutive virulence gene expression in Agrobacterium
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Jin, Shouguang, primary, Song, Yan‐nong, additional, Pan, Shen Q., additional, and Nester, Eugene W., additional
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- 1993
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164. Two-Component Sensory Transduction Systems in Phytobacteria
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Charles, Trevor C., primary, Jin, Shouguang, additional, and Nester, Eugene W., additional
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- 1992
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165. Large-scale production, purification and bioactivity assay of recombinant human interleukin-6 in the methylotrophic yeast Pichia pastoris.
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Li, Hongbo, Wang, Yu, Xu, Aimin, Li, Shiwu, Jin, Shouguang, and Wu, Donghai
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BIOLOGICAL assay ,INTERLEUKIN-6 ,YEAST ,PICHIA pastoris ,DNA ,LYMPHOCYTES ,CELL proliferation - Abstract
A DNA fragment containing the mature human interleukin (IL)-6 sequence was cloned into pPICZαA, generating a fusion protein with the alpha factor from baker's yeast and integrated into the genome of Pichia pastoris strain X-33. Recombinant yeast transformants with high-level rhIL-6 production were identified, secreting as much as 280 mg L rhIL-6 after 4 days of induction by methanol. The rhIL-6 was purified by PEG-8000 precipitation, followed by DEAE anion exchange and Sephadex G-75 gel filtration, yielding over 95% pure rhIL-6 at about 170 mg L. Mass spectrometry analysis showed that the rhIL-6 has a molecular weight of 20 908.85 Da, which is close to the mass calculated from the sequence of the protein. Functional analysis of the purified rhIL-6 using the lymphocyte proliferation assay by an MTT [3-(4,5-dimethylthiazolyl-2)-2,5-diphenyl-tetrazoliumbromide] method demonstrated a specific activity that is at least fivefold higher than the commercial rhIL-6 produced in Escherichia coli. In summary, the experimental procedure we have reported here allows us to obtain a large amount of rhIL-6 from P. pastoris suitable for subsequent biophysical studies. [ABSTRACT FROM AUTHOR]
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- 2011
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166. 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
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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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167. 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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168. Nucleotide sequence of the rpoN gene and the characterization of two downstream open reading...
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Jin, Shouguang and Ishimoto, Karyn
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PSEUDOMONAS , *GENETICS - Abstract
Studies the characteristics of the rpoN gene in Pseudomonas aeroginosa bacterium. P. aeroginosa as a major cause of severe infection in immunocompromised patients; Determination of the nucleotide sequence of the rpoN gene and its nearby regions; Use of mutational analysis in establishing the roles of two proteins encoded by the genes downstream of rpoN.
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- 1994
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169. 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
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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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170. 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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171. 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.
- Abstract
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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172. 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
173. 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
174. 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
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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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175. 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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176. 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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177. 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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178. 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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179. 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
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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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180. 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
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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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181. 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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182. 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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183. Molecular testing devices for on-site detection of E. coli in water samples.
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Manzanas, Carlos, Morrison, Elise, Kim, Young S., Alipanah, Morteza, Adedokun, George, Jin, Shouguang, Osborne, Todd Z., and Fan, Z. Hugh
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ESCHERICHIA coli , *WATER sampling , *COLIFORMS , *WATER quality monitoring , *HALOBACTERIUM , *BACTERIAL DNA , *FECAL contamination - Abstract
Escherichia coli (E. coli) cells are present in fecal materials that can be the main source for disease‐causing agents in water. As a result, E. coli is recommended as a water quality indicator. We have developed an innovative platform to detect E. coli for monitoring water quality on-site by integrating paper-based sample preparation with nucleic acid isothermal amplification. The platform carries out bacterial lysis and DNA enrichment onto a paper pad through ball-based valves for fluid control, with no need of laboratory equipment, followed by loop-mediated isothermal amplification (LAMP) in a battery-operated coffee mug, and colorimetric detection. We have used the platform to detect E. coli in environmental water samples in about 1 h, with a limit of quantitation of 0.2 CFU/mL, and 3 copies per reaction. The platform was confirmed for detecting multiple E. coli strains, and for water samples of different salt concentrations. We validated the functions of the platform by analyzing recreational water samples collected near the Atlantic Ocean that contain different concentrations of salt and bacteria. [ABSTRACT FROM AUTHOR]
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- 2023
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184. Molecular genetic analysis of an XDR Pseudomonas aeruginosa ST664 clone carrying multiple conjugal plasmids.
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Li, Zhenpeng, Cai, Zhao, Cai, Zeqiong, Zhang, Yanhong, Fu, Tongtong, Jin, Yongxin, Cheng, Zhihui, Jin, Shouguang, Wu, Weihui, Yang, Liang, and Bai, Fang
- Subjects
- *
PLASMIDS , *PSEUDOMONAS aeruginosa , *DRUG resistance in microorganisms , *BACTERIAL adaptation , *NUCLEOTIDE sequence , *BACTERIAL evolution , *CARBAPENEMS , *CLINICAL trials , *FUNCTIONAL status , *POST-traumatic stress disorder , *DRUG resistance , *TREATMENT effectiveness , *ECSTASY (Drug) , *PATIENT safety - Abstract
Objectives: A group of ST664 XDR Pseudomonas aeruginosa strains have been isolated from a burn clinic. Here we decipher their resistomes and likely mechanisms of resistance acquisition.Methods: The complete nucleotide sequences of representative isolates were determined, by PacBio and Illumina MiSeq sequencing, and analysed for antimicrobial resistance (AMR) genes as well as sequence variations. S1-PFGE was used to determine the sizes and numbers of plasmids harboured by the isolates. Purified plasmid DNA was further sequenced by PacBio technology, closed manually and annotated by RAST. The mobility of plasmids was determined by conjugation assays.Results: The XDR P. aeruginosa ST664 clone carries 11 AMR genes, including a blaKPC-2 gene that confers resistance to carbapenems. Most of the ST664 isolates carry three coexisting plasmids. blaKPC-2 and a cluster of three AMR genes (aadB-cmlA1-sul1) are encoded on a 475 kb megaplasmid pNK546a, which codes for an IncP-3-like replication and partitioning mechanism, but has lost the conjugative transfer system. Interestingly, however, pNK546a is mobilizable and can be transferred to P. aeruginosa PAO1 with the help of a co-residing IncP-7 conjugative plasmid. The blaKPC-2 gene is carried by an IS6100-ISKpn27-blaKPC-2-ΔISKpn6-Tn1403 mobile element, which might be brought into the ST664 clone by another co-resident IncP-1α plasmid, which is inclined to be lost. Moreover, pNK546a harbours multiple heavy metal (mercury, tellurite and silver) resistance modules.Conclusions: To the best of our knowledge, pNK546a is the first fully sequenced blaKPC-2-carrying megaplasmid from P. aeruginosa. These results give new insights into bacterial adaptation and evolution during nosocomial infections. [ABSTRACT FROM AUTHOR]- Published
- 2020
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185. Combination of Azithromycin and Gentamicin for Efficient Treatment of Pseudomonas aeruginosa Infections.
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Ren, Huan, Liu, Yiwei, Zhou, Jingyi, Long, Yuqing, Liu, Chang, Xia, Bin, Shi, Jing, Fan, Zheng, Liang, Yuying, Chen, Shuiping, Xu, Jun, Wang, Penghua, Zhang, Yanhong, Zhu, Guangbo, Liu, Huimin, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
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- *
PSEUDOMONAS aeruginosa infections , *AZITHROMYCIN , *GENTAMICIN , *PSEUDOMONAS aeruginosa , *ANTIBIOTICS - Abstract
Background: Trans-translation is a ribosome rescue system that plays an important role in bacterial tolerance to environmental stresses. It is absent in animals, making it a potential treatment target. However, its role in antibiotic tolerance in Pseudomonas aeruginosa remains unknown.Methods: The role and activity of trans-translation during antibiotic treatment were examined with a trans-translation-deficient strain and a genetically modified trans-translation component gene, respectively. In vitro assays and murine infection models were used to examine the effects of suppression of trans-translation.Results: We found that the trans-translation system plays an essential role in P. aeruginosa tolerance to azithromycin and multiple aminoglycoside antibiotics. We further demonstrated that gentamicin could suppress the azithromycin-induced activation of trans-translation. Compared with each antibiotic individually, gentamicin and azithromycin combined increased the killing efficacy against planktonic and biofilm-associated P. aeruginosa cells, including a reference strain PA14 and its isogenic carbapenem-resistance oprD mutant, the mucoid strain FRD1, and multiple clinical isolates. Furthermore, the gentamicin-azithromycin resulted in improved bacterial clearance in murine acute pneumonia, biofilm implant, and cutaneous abscess infection models.Conclusions: Combination treatment with gentamicin and azithromycin is a promising strategy in combating P. aeruginosa infections. [ABSTRACT FROM AUTHOR]- Published
- 2019
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186. Identification of novel genes that promote persister formation by repressing transcription and cell division in Pseudomonas aeruginosa.
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Long, Yuqing, Fu, Weixin, Li, Shouyi, Ren, Huan, Li, Mei, Liu, Chang, Zhang, Buyu, Xia, Yushan, Fan, Zheng, Xu, Chang, Liu, Jianfeng, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Liu, Xiaoyun, Jin, Shouguang, and Wu, Weihui
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- *
MULTIDRUG tolerance (Microbiology) , *PSEUDOMONAS aeruginosa , *CELL division , *GENE expression profiling , *RNA sequencing , *PSEUDOMONAS putida - Abstract
Objectives: Bacterial persisters are a small subpopulation of cells that are highly tolerant of antibiotics and contribute to chronic and recalcitrant infections. Global gene expression in Pseudomonas aeruginosa persister cells and genes contributing to persister formation remain largely unknown. The objective of this study was to examine the gene expression profiles of the persister cells and those that regained growth in fresh medium, as well as to identify novel genes related to persister formation.Methods: P. aeruginosa persister cells and those that regrew in fresh medium were collected and subjected to RNA sequencing analysis. Genes up-regulated in the persister cells were overexpressed to evaluate their roles in persister formation. The functions of the persister-contributing genes were assessed with pulse-chase assay, affinity chromatography, fluorescence and electron microscopy, as well as a light-scattering assay.Results: An operon containing PA2282-PA2287 was up-regulated in the persister cells and down-regulated in the regrowing cells. PA2285 and PA2287 play key roles in persister formation. PA2285 and PA2287 were found to bind to RpoC and FtsZ, which are involved in transcription and cell division, respectively. Pulse-chase assays demonstrated inhibitory effects of PA2285 and PA2287 on the overall transcription. Meanwhile, light-scattering and microscopy assays demonstrated that PA2285 and PA2287 interfere with cell division by inhibiting FtsZ aggregation. PA2285 and PA2287 are conserved in pseudomonads and their homologous genes in Pseudomonas putida contribute to persister formation.Conclusions: PA2285 and PA2287 are novel bifunctional proteins that contribute to persister formation in P. aeruginosa. [ABSTRACT FROM AUTHOR]- Published
- 2019
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187. Mechanisms of RsaL mediated tolerance to ciprofloxacin and carbenicillin in Pseudomonas aeruginosa.
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Fan, Zheng, Xu, Chang, Pan, Xiaolei, Dong, Yuanyuan, Ren, Huan, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
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- *
PSEUDOMONAS aeruginosa , *CIPROFLOXACIN , *CARBENICILLIN , *GENE expression in bacteria , *BACTERIAL operons - Abstract
The Pseudomonas aeruginosa RsaL is a negative regulator of the quorum sensing signal synthesis gene lasI. The expression of RsaL is directly activated by the LasI cognate regulator LasR. Thus, RsaL and LasI-LasR (LasI/R) form a regulatory loop. Further studies revealed that RsaL is a global regulator which controls the expression of numerous genes through quorum sensing system dependent and independent pathways. However, whether RsaL is involved in antibiotic tolerance remains elusive. In this study, we found that the mutation of rsaL increased bacterial tolerance to ciprofloxacin and carbenicillin. Through motif search, gene expression analyses and electrophoretic mobility shift assays, we found that RsaL directly represses the expression of the narK1K2GHJI operon, which is involved in the tolerance to ciprofloxacin. We further demonstrated that the narK1K2GHJI operon is directly regulated by LasR. In combination, our study revealed a novel operon under the control of the RsaL, LasI/R regulatory loop. [ABSTRACT FROM AUTHOR]
- Published
- 2019
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188. Halogenated quinolines bearing polar functionality at the 2-position: Identification of new antibacterial agents with enhanced activity against Staphylococcus epidermidis.
- Author
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Basak, Akash, Abouelhassan, Yasmeen, Kim, Young S., IVNorwood, Verrill M., Jin, Shouguang, and IIIHuigens, Robert W.
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STAPHYLOCOCCUS epidermidis , *BIOFILMS , *QUINOLINE , *ANTIBACTERIAL agents , *STAPHYLOCOCCUS aureus , *ANTI-infective agents - Abstract
Antibiotic-resistant bacteria and surface-attached biofilms continue to play a significant role in human health and disease. Innovative strategies are needed to identify new therapeutic leads to tackle infections of drug-resistant and tolerant bacteria. We synthesized a focused library of 14 new halogenated quinolines to investigate the impact of ClogP values on antibacterial and biofilm-eradication activities. During these investigations, we found select polar appendages at the 2-position of the HQ scaffold were more well-tolerated than others. We were delighted to see multiple compounds display enhanced activities against the major human pathogen S. epidermidis. In particular, HQ 2 (ClogP = 3.44) demonstrated enhanced activities against MRSE 35984 planktonic cells (MIC = 0.59 μM) compared to MRSA and VRE strains in addition to potent MRSE biofilm eradication activities (MBEC = 2.35 μM). Several of the halogenated quinolines identified here reported low cytotoxicity against HeLa cells with minimal hemolytic activity against red blood cells. We believe that halogenated quinoline small molecules could play an important role in the development of next-generation antibacterial therapeutics capable of targeting and eradicating biofilm-associated infections. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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189. Enhanced differentiation of human pluripotent stem cells into cardiomyocytes by bacteria-mediated transcription factors delivery.
- Author
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Jin, Yongxin, Liu, Ying, Li, Zhenpeng, Santostefano, Katherine, Shi, Jing, Zhang, Xinwen, Wu, Donghai, Cheng, Zhihui, Wu, Weihui, Terada, Naohiro, Jin, Shouguang, and Bai, Fang
- Subjects
- *
PLURIPOTENT stem cells , *CELL differentiation , *HEART cells , *BACTERIAL genetics , *GENE expression , *GENETIC transcription - Abstract
Virus-mediated expression of defined transcription factor (TF) genes can effectively induce cellular reprogramming. However, sustained expression of the TFs often hinders pluripotent stem cell (PSC) differentiation into specific cell types, as each TF exerts its effect on PSCs for a defined period of time during differentiation. Here, we applied a bacterial type III secretion system (T3SS)-based protein delivery tool to directly translocate TFs in the form of protein into human PSCs. This transient protein delivery technique showed high delivery efficiency for hPSCs, and it avoids potential genetic alterations caused by the introduction of transgenes. In an established cardiomyocyte de novo differentiation procedure, five transcriptional factors, namely GATA4, MEF2C, TBX5, ESRRG and MESP1 (abbreviated as GMTEM), were translocated at various time points. By detecting the expression of cardiac marker genes (Nkx2.5 and cTnT), we found that GMTEM proteins delivered on mesoderm stage of the cardiomyocytes lineage differentiation significantly enhanced both the human ESC and iPSC differentiation into cardiomyocytes, while earlier or later delivery diminished the enhancing effect. Furthermore, all of the five factors were required to enhance the cardiac differentiation. This work provides a virus-free strategy of transient transcription factors delivery for directing human stem cell fate without jeopardizing genome integrity, thus safe for biomedical applications. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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190. Bacterial type III secretion system as a protein delivery tool for a broad range of biomedical applications.
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Bai, Fang, Li, Zhenpeng, Umezawa, Akihiro, Terada, Naohiro, and Jin, Shouguang
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- *
BACTERIAL secretions , *CHROMOSOMAL translocation , *PROTEIN fractionation , *ANTIGENS , *GENOME editing - Abstract
A protein delivery tool based on bacterial type III secretion system (T3SS) has been broadly applied in biomedical researches. In this review, we summarize various applications of the T3SS-mediate protein delivery which enables translocation of proteins directly into mammalian cells without protein purification. Some of the remarkable advancements include delivery of antigens for therapeutic vaccines, nucleases for genome editing, transcription factors for cellular reprogramming and stem cells differentiation, and signaling molecules for post-translational proteomics studies. With continued improvement of the T3SS-mediated protein delivery tools, even wider application of the technology is anticipated. [ABSTRACT FROM AUTHOR]
- Published
- 2018
- Full Text
- View/download PDF
191. Epidemiological characterization of Acinetobacter baumannii bloodstream isolates from a Chinese Burn Institute: A three-year study.
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Huang, Guangtao, Yin, Supeng, Xiang, Lijuan, Gong, Yali, Sun, Kedai, Luo, Xiaoqiang, Zhang, Cheng, Yang, Zichen, Deng, Liuyang, Jiang, Bei, Jin, Shouguang, Chen, Jing, and Peng, Yizhi
- Subjects
- *
BACTERIAL disease risk factors , *BURN patients , *EPIDEMIOLOGY , *ACINETOBACTER baumannii , *DRUG resistance in bacteria , *NUCLEOTIDE sequencing , *PUBLIC health , *ACINETOBACTER infections , *ANTIBIOTICS , *BACTEREMIA , *BACTERIAL proteins , *BACTERIOPHAGE typing , *BURNS & scalds , *CROSS infection , *DRUG resistance in microorganisms , *HYDROLASES , *MICROBIAL sensitivity tests , *MOLECULAR epidemiology , *POLYMERASE chain reaction , *RNA , *RETROSPECTIVE studies , *GRAM-negative aerobic bacteria , *SEQUENCE analysis , *PHARMACODYNAMICS - Abstract
Acinetobacter baumannii infection is a serious threat to burn patients. Bacteremia due to A. baumannii is becoming the most common cause of mortality following burn. However, the epidemiology of A. baumannii causing burn-related bloodstream infections has rarely been reported. We retrospectively collected 81 A. baumannii isolates from the bloodstream of burn patients over a three-year period. Antibiotic susceptibility tests, the prevalence of antibiotic-resistant genes and sequence typing (ST) were conducted to characterize these strains. Most of the isolates showed an extensive drug-resistant phenotype. The resistance frequencies to imipenem and meropenem were 94% and 91%, respectively. The blaOXA-23-like gene, AmpC, IS-AmpC, PER and SIM are the five most prevalent resistant genes, and their prevalence rates are 93% (75/81), 86% (70/81), 73% (59/81), 73% (59/81) and 52% (42/81), respectively. The 81 isolates were grouped into 10 known and 18 unknown ST types, with ST368 (38%) being the most prevalent. Except for ST457 and four new types (STn2, STn6, STn11 and STn14), the remaining 23 ST types belonged to one clonal complex 92, which is most common among clinical isolate in China. The above results indicated that ST368 isolates possessing both the blaOXA-23-like gene and ampC gene were the main culprits of the increasing nosocomial A. baumannii infection in this study. More attention should be paid to monitoring the molecular epidemiology of A. baumannii isolates from burn patients to prevent further distribution. Such information may help clinicians with therapeutic decisions and infection control in the Burns Institute. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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192. Expression and efficient purification of tag-cleaved active recombinant human insulin-like growth factor-II from Escherichia coli.
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Li, Hongbo, Hui, Xiaoyan, Li, Peng, Xu, Aimin, Li, Shiwu, Jin, Shouguang, and Wu, Donghai
- Subjects
- *
SOMATOMEDIN A , *CELL proliferation , *APOPTOSIS , *THIOREDOXIN , *PROTEINS , *BACTERIAL cultures - Abstract
Insulin-like growth factor-II (IGF2) is a growth factor for the control of cell proliferation and apoptosis. To explore the clinical use of human IGF2, an efficient method for production of a large amount of active recombinant hIGF2 is necessary. Human IGF2 cDNA was cloned into pET32 vector where it is under the control of an IPTGinducible T7 promoter. High level soluble thioredoxin (Trx)-hIGF2 fusion protein was produced at room temperature following IPTG induction, amounting up to 20% of the total soluble bacterial proteins. The recombinant Trx-hIGF2 fusion protein was purified to an approximate 95% purity using Ni-NTA affinity chromatography with an overall yield of 120 mg protein per liter of bacterial culture. After cleavage of the Trx fusion fragment by recombinant enterokinase, the tag-free recombinant hIGF2 protein (rhIGF2) was purified by passage through the Ni+-NTA affinity column again. Biological activity of the purified hIGF2 was determined by its ability to support NIH/3T3 cells proliferation and to activate AKT signaling pathways. Our results demonstrate that tag-free active rhIGF2 can easily be obtained for various applications from E. coli using the procedure described in this report. [ABSTRACT FROM AUTHOR]
- Published
- 2015
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193. MKP1 regulates the induction of MCP1 by Streptococcus pneumoniae pneumolysin in human epithelial cells.
- Author
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Shin, Hee-Sung, Yoo, In-Hwa, Kim, Yong-Jae, Lee, Ji-Yun, Kim, Hyong-Bai, Jin, Shouguang, and Ha, Un-Hwan
- Abstract
Epithelial cells act as the first line of host defense against microbes by producing a range of different molecules for clearance. Chemokines facilitate the clearance of invaders through the recruitment of leukocytes. Thus, upregulation of chemokine expression represents an important innate host defense response against invading microbes such as Streptococcus pneumoniae. In this study, we report that the expression of Monocyte Chemotactic Protein 1 (MCP1) was highly induced in response to S. pneumoniae in vitro and in vivo. Among numerous virulence factors, pneumococcal pneumolysin was found to be the major factor responsible for this induction. Furthermore, MCP1 induction was mediated by the p38 mitogen-activated protein kinase (MAPK) whose activation was controlled by MAPK phosphatase 1 (MKP1). Therefore, this study reveals novel roles of pneumolysin in mediating MKP1 expression for the regulation of MCP1 expression in human epithelial cells. [ABSTRACT FROM AUTHOR]
- Published
- 2010
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194. Sensitive detection of multiplex toxins using antibody microarray
- Author
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Lian, Wei, Wu, Donghai, Lim, Daniel V., and Jin, Shouguang
- Subjects
- *
NANOPARTICLES , *FLUORESCENT lighting , *BIOTERRORISM , *MONOCLONAL antibodies , *CHOLERA toxin , *ENZYME-linked immunosorbent assay - Abstract
Abstract: Using a newly developed fluorescent nanoparticle (NP) that gives rise to a high-intensity and stable fluorescent light, a sensitive antibody (Ab) microarray assay system has been developed for specific detection of bioterrorism agents, as exemplified by ricin, cholera toxin (CT), and staphylococcal enterotoxin B (SEB). The Ab microarray uses a sandwich format that consists of capture Abs, analytes (toxins), biotinylated detection Abs, and avidin-conjugated NP. In all three cases, polyclonal Abs (pAbs) displayed superiority over monoclonal antibodies (mAbs) in capturing toxins on microarray slides even when the pAbs and mAbs had similar affinity as determined by enzyme-linked immunosorbent assay (ELISA). The detection system was successfully used to detect toxins spiked in milk, apple cider, and blood samples. We were able to detect ricin at 100pg/ml in buffer and at 1ng/ml in spiked apple cider or milk, whereas CT and SEB were detected at 10pg/ml in buffer and 100pg/ml in spiked apple cider or milk. High specificities were also demonstrated in the detection of mixed toxin samples with similar sensitivities. The matrix effect of blood samples on the detection of mixed toxins seems to be minimal when the toxin concentration is at or above 100ng/ml. The current study highlights the significant role of pAb and NP in increasing selectivity and sensitivity of toxin detection in a microarray format. [Copyright &y& Elsevier]
- Published
- 2010
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- View/download PDF
195. Ultrasensitive detection of biomolecules with fluorescent dye-doped nanoparticles
- Author
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Lian, Wei, Litherland, Sally A., Badrane, Hassan, Tan, Weihong, Wu, Donghai, Baker, Henry V., Gulig, Paul A., Lim, Daniel V., and Jin, Shouguang
- Subjects
- *
BIOMOLECULES , *NANOPARTICLES , *IMMUNOCYTOCHEMISTRY , *IMMUNOFLUORESCENCE - Abstract
Fluorescent-labeled molecules have been used extensively for a wide range of applications in biological detection and diagnosis. A new form of highly luminescent and photostable nanoparticles was generated by doping the fluorescent dye tris(2′2-bipyridyl)dichlororuthenium(II)hexahydrate (Rubpy) inside silica material. Because thousands of fluorescent dye molecules are encapsulated in the silica matrix that also serves to protect Rubpy dye from photodamaging oxidation, the Rubpy-dye-doped nanoparticles are extremely bright and photostable. We have used these nanoparticles successfully in various fluorescence labeling techniques, including fluorescent-linked immunosorbent assay, immunocytochemistry, immunohistochemistry, DNA microarray, and protein microarray. By combining the high-intensity luminescent nanoparticles with the specificity of antibody-mediated recognition, ultrasensitive target detection has been achieved. In all cases, assay results clearly demonstrated the superiority of the nanoparticles over organic fluorescent dye molecules and quantum dots in probe labeling for sensitive target detection. These results demonstrate the potential to apply these newly developed fluorescent nanoparticles in various biodetection systems. [Copyright &y& Elsevier]
- Published
- 2004
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196. A rapid bioassay for single bacterial cell quantitation using bioconjugated nanoparticles.
- Author
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zhao, Xiaojun, Hilliard, Lisa R., Mechery, Shelly John, wang, Yanping, Bagwe, Rahul P., Jin, Shouguang, and Tan, Weihong
- Subjects
- *
PATHOGENIC bacteria , *MEDICAL bacteriology , *BACTERIAL diseases , *BIOTECHNOLOGY , *NANOPARTICLES , *BIOTERRORISM - Abstract
The rapid and sensitive determination of pathogenic bacteria is extremely important in biotechnology, medical diagnosis, and the current fight against bioterrorism. Current methods either lack ultra sensitivity or take a long time for analysis. Here, we report a bio conjugated nanoparticle-based bioassay for in situ pathogen quantification down to single bacterium within 20 min. The bios conjugated nanoparticle provides an extremely high fluorescent signal for bioanalysis and can be easily incorporated with biorecognition molecules, such nobody. The antibody-conjugated nanoparticles can readily and specifically identify variety of bacterium, such as Escherkhia coil 0157 H7, through antibody-antigen inter. The single-bacterium-detection capability within 20 min has been confirmed by the plate-counting method and realized by using two independent optical techniques. The two detection methods correlated extremely well. Furthermore, we were able to detect multiple bacterial samples with high through- put by using a 384-well microplate format. ,o show the usefulness of this assay, we have accurately detected 1-400 E coIl 0157 bacterial cells in spiked ground beef samples. Our results demons strate the potential for a broad application of bioconjugated nanoparticles in practical biotechnological and medical applications in various biodetection systems. The ultimate power of integrating bionano technology into complex biological systems will emerge as a revolutionary tool for ultra sensitive detection of disease markers and infectious agents. [ABSTRACT FROM AUTHOR]
- Published
- 2004
- Full Text
- View/download PDF
197. The truA gene of Pseudomonas aeruginosa is required for the expression of type III secretory genes.
- Author
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Ahn, Kyung-Seop, Ha, Unhwan, Jia, Jinghua, Wu, Donghai, and Jin, Shouguang
- Subjects
- *
PSEUDOMONAS aeruginosa , *GENE expression , *GENES , *GENETIC mutation , *PROTEINS , *TRANSFER RNA - Abstract
Invasive strains of Pseudomonas aeruginosa can cause rapid host cell apoptosis by injecting the type III effector molecule ExoS. A transposon insertional mutant bank of P. aeruginosa was screened to identify P. aeruginosa genes that contribute to the ability of the bacteria to trigger host cell apoptosis. Several isolated mutants had disruptions in the fimV gene. A fimV mutant was unable to induce the expression of exoS, exoT and exsA genes under type III inducing conditions, thus exhibiting a defect in type III protein secretion. Furthermore, this mutant was defective in twitching motility, although type IV pili were present on the bacterial surface. Complementation by a fimV-containing cosmid clone restored both phenotypes to the wild-type levels. However, expression of the type III genes in the fimV mutant was not restored by the introduction of a fimV gene alone, although it restored the twitching motility. A gene downstream of fimV, encoding a tRNA pseudouridine synthase (truA) homologue, was able to complement the type III gene expression defect of the fimV mutant. Thus firnV and truA form an operon and fimV mutation has a polar effect on truA. Indeed, a truA mutant is defective in type III gene expression while its twitching motility is unaffected, and a truA clone is able to complement the type III secretion defect. Pseudouridination of tRNAs is important for tRNA structure, thereby improving the fidelity of protein synthesis and helping to maintain the proper reading frame; thus the results imply that truA controls tRNAs that are critical for the translation of type III genes or their regulators. [ABSTRACT FROM AUTHOR]
- Published
- 2004
- Full Text
- View/download PDF
198. The Pseudomonas aeruginosa HSP70-like protein DnaK induces IL-1β expression via TLR4-dependent activation of the NF-κB and JNK signaling pathways.
- Author
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Lee, Jung-Hoon, Jeon, Jisu, Bai, Fang, Jin, Shouguang, Wu, Weihui, and Ha, Un-Hwan
- Subjects
- *
PSEUDOMONAS aeruginosa , *HEAT shock proteins , *EMERGING infectious diseases , *NOSOCOMIAL infections ,MECHANICAL shock measurement - Abstract
• P. aeruginosa is recognized as an emerging nosocomial infection in veterinary. • IL-1β mediates acute inflammatory responses to P. aeruginosa infections. • P. aeruginosa HSP 70-like protein DnaK induces the expression of IL-1β. • DnaK-mediated IL-1β expression is driven by TLR4-dependent NF-κB and JNK pathways. IL-1β expression is increased in response to P. aeruginosa infection, but the responsible proteins have not been clearly elucidated. Here, we demonstrate for the first time that IL-1β expression is induced in response to the heat shock protein 70-like protein DnaK. Treatment with recombinant DnaK (rDnaK) increased IL-1β expression in a dose- and time-dependent manner, and the release of mature IL-1β in response to rDnaK was detected to an extent similar to that stimulated by the well-known agonists, lipopolysaccharide and nigericin. rDnaK-mediated IL-1β expression was driven by the NF-κB signaling pathway. In addition, expression was controlled by the JNK signaling pathway, although these two signaling cascades act independently upon rDnaK stimulation. Finally, rDnaK-induced IL-1β expression was initiated via the action of TLR4. Taken together, the data reveal that P. aeruginosa -derived DnaK induces expression of IL-1β via TLR4-dependent activation of the NF-κB and JNK signaling pathways. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF
199. HigB Reciprocally Controls Biofilm Formation and the Expression of Type III Secretion System Genes through Influencing the Intracellular c-di-GMP Level in Pseudomonas aeruginosa.
- Author
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Zhang, Yueying, Xia, Bin, Li, Mei, Shi, Jing, Long, Yuqing, Jin, Yongxin, Bai, Fang, Cheng, Zhihui, Jin, Shouguang, and Wu, Weihui
- Subjects
- *
PSEUDOMONAS aeruginosa , *ANTITOXINS , *GENE expression , *MICROBIAL virulence , *BIOFILMS - Abstract
Toxin-antitoxin (TA) systems play important roles in bacteria persister formation. Increasing evidence demonstrate the roles of TA systems in regulating virulence factors in pathogenic bacteria. The toxin HigB in Pseudomonas aeruginosa contributes to persister formation and regulates the expression of multiple virulence factors and biofilm formation. However, the regulatory mechanism remains elusive. In this study, we explored the HigB mediated regulatory pathways. We demonstrate that HigB decreases the intracellular level of c-di-GMP, which is responsible for the increased expression of the type III secretion system (T3SS) genes and repression of biofilm formation. By analyzing the expression levels of the known c-di-GMP metabolism genes, we find that three c-di-GMP hydrolysis genes are up regulated by HigB, namely PA2133, PA2200 and PA3825. Deletion of the three genes individually or simultaneously diminishes the HigB mediated regulation on the expression of T3SS genes and biofilm formation. Therefore, our results reveal novel functions of HigB in P. aeruginosa. [ABSTRACT FROM AUTHOR]
- Published
- 2018
- Full Text
- View/download PDF
200. Host Cells Upregulate Phosphate Transporter PIT1 to Inhibit Ehrlichia chaffeensis Intracellular Growth.
- Author
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Li M, Yang N, Li X, Duan N, Qin S, Wang M, Zhou Y, Jin Y, Wu W, Jin S, and Cheng Z
- Subjects
- Humans, THP-1 Cells, Up-Regulation, Ehrlichiosis microbiology, Ehrlichiosis metabolism, Host-Pathogen Interactions, Macrophages metabolism, Macrophages microbiology, Macrophages immunology, Phosphates metabolism, NF-kappa B metabolism, Monocytes metabolism, Monocytes microbiology, Ehrlichia chaffeensis metabolism, Ehrlichia chaffeensis genetics
- Abstract
Ehrlichia chaffeensis infects and proliferates inside monocytes or macrophages and causes human monocytic ehrlichiosis (HME), an emerging life-threatening tick-borne zoonosis. After internalization, E. chaffeensis resides in specialized membrane-bound inclusions, E. chaffeensis -containing vesicles (ECVs), to evade from host cell innate immune responses and obtain nutrients. However, mechanisms exploited by host cells to inhibit E. chaffeensis growth in ECVs are still largely unknown. Here we demonstrate that host cells recognize E. chaffeensis Ech_1067, a penicillin-binding protein, and then upregulate the expression of PIT1, which is a phosphate transporter and transports phosphate from ECVs to the cytosol to inhibit bacterial growth. We found that host cells upregulate the PIT1 expression upon E. chaffeensis infection using transcriptome sequencing, qRT-PCR and Western blotting, and PIT1 is localized on the ECV membrane in infected THP-1 cells using confocal microscopy. Silence of PIT1 using shRNA enhances E. chaffeensis intracellular growth. Finally, we found that E. chaffeensis Ech_1067 induces the upregulation of PIT1 expression through the MyD88-NF-κB pathway using recombinant protein for stimulation and siRNA for silence. Our findings deepen the understanding of the innate immune responses of host cells to inhibit bacterial intracellular growth and facilitate the development of new therapeutics for HME.
- Published
- 2024
- Full Text
- View/download PDF
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