scholarly journals The Alternative Sigma Factor σE Plays an Important Role in Intestinal Survival and Virulence in Vibrio cholerae

2002 ◽  
Vol 70 (10) ◽  
pp. 5355-5362 ◽  
Author(s):  
Gabriela Kovacikova ◽  
Karen Skorupski
Keyword(s):  
Vibrio Cholerae ◽  
Sigma Factor ◽  
Parental Strain ◽  
Time Course ◽  
Lethal Dose ◽  
Alternative Sigma Factor ◽  
Growth And Survival ◽  
Infant Mouse

ABSTRACT The alternative sigma factor σΕ (RpoE) is involved in the response to extracytoplasmic stress and plays a role in the virulence of a variety of different bacteria. To assess the role of σΕ in Vibrio cholerae pathogenesis, a ΔrpoE mutant was constructed and analyzed using the infant mouse model. The results here show that σΕ contributes significantly to the virulence of V. cholerae. The ΔrpoE mutant was highly attenuated with a 50% lethal dose more than 3 logs higher than that for the parental strain, and its ability to colonize the intestine was reduced approximately 30-fold. A time course of infection revealed that the number of CFU of the ΔrpoE mutant was approximately 1 log lower than that of the parental strain by 12 h postinoculation and decreased further by 24 h. The defect in virulence in the ΔrpoE mutant thus appears to be a diminished ability to survive within the intestinal environment. The results here also show that σΕ is not required for growth and survival of V. cholerae in vitro at high temperatures but is required under other stressful conditions, such as in the presence of 3% ethanol. As in Escherichia coli, the expression of rpoE in V. cholerae is dependent upon two promoters located upstream of the gene, P1 and P2. P1 appears to be σ70 dependent, whereas the downstream promoter, P2, is positively autoregulated by σΕ.

scholarly journals Role of Vibrio cholerae O139 Surface Polysaccharides in Intestinal Colonization

2002 ◽  
Vol 70 (11) ◽  
pp. 5990-5996 ◽  
Author(s):  
Jutta Nesper ◽  
Stefan Schild ◽  
Crystal M. Lauriano ◽  
Anita Kraiss ◽  
Karl E. Klose ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Capsular Polysaccharide ◽  
Genetically Engineered ◽  
Side Chain ◽  
Intestinal Colonization ◽  
Core Oligosaccharide ◽  
Infant Mouse ◽  
Surface Polysaccharides

ABSTRACT Since the first occurrence of O139 Vibrio cholerae as a cause of cholera epidemics, this serogroup has been investigated intensively, and it has been found that its pathogenicity is comparable to that of O1 El Tor strains. O139 isolates express a thin capsule, composed of a polymer of repeating units structurally identical to the lipopolysaccharide (LPS) O side chain. In this study, we investigated the role of LPS O side chain and capsular polysaccharide (CPS) in intestinal colonization by with genetically engineered mutants. We constructed CPS-negative, CPS/LPS O side chain-negative, and CPS-positive/LPS O side chain-negative mutants. Furthermore, we constructed two mutants with defects in LPS core oligosaccharide (OS) assembly. Loss of LPS O side chain or CPS resulted in a ≈30-fold reduction in colonization of the infant mouse small intestine, indicating that the presence of both LPS O side chain and CPS is important during the colonization process. The strain lacking both CPS and LPS O side chain and a CPS-positive, LPS O side chain-negative core OS mutant were both essentially unable to colonize. To characterize the role of surface polysaccharides in survival in the host intestine, resistance to several antimicrobial substances was investigated in vitro. These investigations revealed that the presence of CPS protects the cell against attack of the complement system and that an intact core OS is necessary for survival in the presence of bile.

scholarly journals Role of Cyclic Di-GMP during El Tor Biotype Vibrio cholerae Infection: Characterization of the In Vivo-Induced Cyclic Di-GMP Phosphodiesterase CdpA

2008 ◽  
Vol 76 (4) ◽  
pp. 1617-1627 ◽  
Author(s):  
Rita Tamayo ◽  
Stefan Schild ◽  
Jason T. Pratt ◽  
Andrew Camilli
Keyword(s):  
Small Intestine ◽  
Vibrio Cholerae ◽  
Biofilm Formation ◽  
Virulence Gene ◽  
Virulence Gene Expression ◽  
Infant Mouse

ABSTRACT In Vibrio cholerae, the second messenger cyclic di-GMP (c-di-GMP) positively regulates biofilm formation and negatively regulates virulence and is proposed to play an important role in the transition from persistence in the environment to survival in the host. Herein we describe a characterization of the infection-induced gene cdpA, which encodes both GGDEF and EAL domains, which are known to mediate diguanylate cyclase and c-di-GMP phosphodiesterase (PDE) activities, respectively. CdpA is shown to possess PDE activity, and this activity is regulated by its inactive degenerate GGDEF domain. CdpA inhibits biofilm formation but has no effect on colonization of the infant mouse small intestine. Consistent with these observations, cdpA is expressed during in vitro growth in a biofilm but is not expressed in vivo until the late stage of infection, after colonization has occurred. To test for a role of c-di-GMP in the early stages of infection, we artificially increased c-di-GMP and observed reduced colonization. This was attributed to a significant reduction in toxT transcription during infection. Cumulatively, these results support a model of the V. cholerae life cycle in which c-di-GMP must be down-regulated early after entering the small intestine and maintained at a low level to allow virulence gene expression, colonization, and motility at appropriate stages of infection.

scholarly journals A Recombinant Live Attenuated Strain of Vibrio cholerae Induces Immunity against Tetanus Toxin andBordetella pertussis Tracheal Colonization Factor

1998 ◽  
Vol 66 (4) ◽  
pp. 1648-1653 ◽  
Author(s):  
Inês Chen ◽  
Theresa M. Finn ◽  
Liu Yanqing ◽  
Qi Guoming ◽  
Rino Rappuoli ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Tetanus Toxin ◽  
Bacterial Colonization ◽  
Lethal Dose ◽  
Lethal Challenge ◽  
Colonization Factor ◽  
Heterologous Antigens

ABSTRACT An attenuated strain of Vibrio cholerae was used as a carrier for the expression of heterologous antigens such as fragment C from tetanus toxin (TetC) and tracheal colonization factor fromBordetella pertussis (Tcf). In vitro, high levels of protein were obtained when the Escherichia coli nirBpromoter was used and the bacteria were grown with low aeration. Intranasal immunization of mice with IEM101 expressing TetC elicited serum vibriocidal activity and induced antibodies against tetanus toxin which were protective against lethal challenge with 10 times the 50% lethal dose of tetanus toxin. Bacterial viability was essential for the induction of anti-TetC antibodies. Intranasal administration of IEM101 expressing Tcf induced a significant reduction in bacterial colonization of the tracheas of mice challenged with wild-type B. pertussis. These data are in agreement with the putative role of Tcf in Bordetellatracheal colonization. In conclusion, we have demonstrated thatV. cholerae may be used as a live vector to deliver heterologous antigens in vivo and that protection to both systemic and local challenge may be achieved.

scholarly journals The Alternative Sigma Factor, ςE, Is Critically Important for the Virulence of Salmonella typhimurium

1999 ◽  
Vol 67 (4) ◽  
pp. 1560-1568 ◽  
Author(s):  
Sue Humphreys ◽  
Andrew Stevenson ◽  
Andrew Bacon ◽  
A. Barbara Weinhardt ◽  
Mark Roberts
Keyword(s):  
Salmonella Typhimurium ◽  
Cell Lines ◽  
Sigma Factor ◽  
Cell Envelope ◽  
Critical Role ◽  
Lethal Dose ◽  
Carbon Sources ◽  
Alternative Sigma Factor

ABSTRACT In Escherichia coli, extracytoplasmic stress is partially controlled by the alternative sigma factor, RpoE (ςE). In response to environmental stress or alteration in the protein content of the cell envelope, ςEupregulates the expression of a number of genes, includinghtrA. It has been shown that htrA is required for intramacrophage survival and virulence in Salmonella typhimurium. To investigate whether ςE-regulated genes other than htrA are involved in salmonella virulence, we inactivated the rpoE gene of S. typhimurium SL1344 by allelic exchange and compared the phenotype of the mutant (GVB311) in vitro and in vivo with its parent and an isogenic htrA mutant (BRD915). UnlikeE. coli, ςE is not required for the growth and survival of S. typhimurium at high temperatures. However, GVB311 did display a defect in its ability to utilize carbon sources other than glucose. GVB311 was more sensitive to hydrogen peroxide, superoxide, and antimicrobial peptides than SL1344 and BRD915. Although able to invade both macrophage and epithelial cell lines normally, the rpoE mutant was defective in its ability to survive and proliferate in both cell lines. The effect of the rpoE mutation on the intracellular behavior of S. typhimurium was greater than that of the htrA mutation. Both GVB311 and BRD915 were highly attenuated in mice. Neither strain was able to kill mice via the oral route, and the 50% lethal dose (LD50) for both strains via the intravenous (i.v.) route was very high. The i.v. LD50s for SL1344, BRD915, and GVB311 were <10, 5.5 × 105, and 1.24 × 107 CFU, respectively. Growth in murine tissues after oral and i.v. inoculation was impaired for both thehtrA and rpoE mutant, with the latter mutant being more severely affected. Neither mutant was able to translocate successfully from the Peyer’s patches to other organs after oral infection or to proliferate in the liver and spleen after i.v. inoculation. However, the htrA mutant efficiently colonized the livers and spleens of mice infected i.v., but the rpoE mutant did not. Previous studies have shown that salmonella htrA mutants are excellent live vaccines. In contrast, oral immunization of mice with GVB311 was unable to protect any of the mice from oral challenge with SL1344. Furthermore, i.v. immunization with a large dose (∼106 CFU) of GVB311 protected less than half of the orally challenged mice. Thus, our results indicate that genes in the ςE regulon other than htrA play a critical role in the virulence and immunogenicity of S. typhimurium.

scholarly journals Enhanced phosphorylation of Na+–Cl− co-transporter in experimental metabolic syndrome: role of insulin

2012 ◽  
Vol 123 (11) ◽  
pp. 635-647 ◽  
Author(s):  
Radko Komers ◽  
Shaunessy Rogers ◽  
Terry T. Oyama ◽  
Bei Xu ◽  
Chao-Ling Yang ◽  
...  
Keyword(s):  
Metabolic Syndrome ◽  
Time Course ◽  
Kinase Inhibitor ◽  
Endogenous Expression ◽  
The Metabolic Syndrome ◽  
Mammalian Cell Lines ◽  
Phosphoinositide 3 Kinase

In the present study, we investigated the activity of the thiazide-sensitive NCC (Na+–Cl− co-transporter) in experimental metabolic syndrome and the role of insulin in NCC activation. Renal responses to the NCC inhibitor HCTZ (hydrochlorothiazide), as a measure of NCC activity in vivo, were studied in 12-week-old ZO (Zucker obese) rats, a model of the metabolic syndrome, and in ZL (Zucker lean) control animals, together with renal NCC expression and molecular markers of NCC activity, such as localization and phosphorylation. Effects of insulin were studied further in mammalian cell lines with inducible and endogenous expression of this molecule. ZO rats displayed marked hyperinsulinaemia, but no differences in plasma aldosterone, compared with ZL rats. In ZO rats, natriuretic and diuretic responses to NCC inhibition with HCTZ were enhanced compared with ZL rats, and were associated with a decrease in BP (blood pressure). ZO rats displayed enhanced Thr53 NCC phosphorylation and predominant membrane localization of both total and phosphorylated NCC, together with a different profile in expression of SPAK (Ste20-related proline/alanine-rich kinase) isoforms, and lower expression of WNK4. In vitro, insulin induced NCC phosphorylation, which was blocked by a PI3K (phosphoinositide 3-kinase) inhibitor. Insulin-induced reduction in WNK4 expression was also observed, but delayed compared with the time course of NCC phosphorylation. In summary, we report increased NCC activity in hyperinsulinaemic rodents in conjunction with the SPAK expression profile consistent with NCC activation and reduced WNK4, as well as an ability of insulin to induce NCC stimulatory phosphorylation in vitro. Together, these findings indicate that hyperinsulinaemia is an important driving force of NCC activity in the metabolic syndrome with possible consequences for BP regulation.

scholarly journals Alternative Sigma Factor SigK Has a Role in Stress Tolerance of Group I Clostridium botulinum Strain ATCC 3502

2013 ◽  
Vol 79 (12) ◽  
pp. 3867-3869 ◽  
Author(s):  
Elias Dahlsten ◽  
David Kirk ◽  
Miia Lindström ◽  
Hannu Korkeala
Keyword(s):  
Stress Tolerance ◽  
Sigma Factor ◽  
Clostridium Botulinum ◽  
Alternative Sigma Factor ◽  
Strain Atcc ◽  
Content Type ◽  
Osmotic Stress Tolerance ◽  
Group I ◽  
Temperature Downshift

ABSTRACTThe role of the alternative sigma factor SigK in cold and osmotic stress tolerance ofClostridium botulinumATCC 3502 was demonstrated by induction ofsigKafter temperature downshift and exposure to hyperosmotic conditions and by impaired growth of thesigKmutants under the respective conditions.

scholarly journals Role of INSL4 Signaling in Sustaining the Growth and Viability of LKB1-Inactivated Lung Cancer

2018 ◽  
Vol 111 (7) ◽  
pp. 664-674 ◽  
Author(s):  
Rongqiang Yang ◽  
Steven W Li ◽  
Zirong Chen ◽  
Xin Zhou ◽  
Wei Ni ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Transcriptome Profiling ◽  
Cancer Genome ◽  
The Cancer Genome Atlas ◽  
Growth And Survival ◽  
Cancer Genome Atlas ◽  
Lung Adenocarcinomas ◽  
Genome Atlas

Abstract Background The LKB1 tumor suppressor gene is commonly inactivated in non-small cell lung carcinomas (NSCLC), a major form of lung cancer. Targeted therapies for LKB1-inactivated lung cancer are currently unavailable. Identification of critical signaling components downstream of LKB1 inactivation has the potential to uncover rational therapeutic targets. Here we investigated the role of INSL4, a member of the insulin/IGF/relaxin superfamily, in LKB1-inactivated NSCLCs. Methods INSL4 expression was analyzed using global transcriptome profiling, quantitative reverse transcription PCR, western blotting, enzyme-linked immunosorbent assay, and RNA in situ hybridization in human NSCLC cell lines and tumor specimens. INSL4 gene expression and clinical data from The Cancer Genome Atlas lung adenocarcinomas (n = 515) were analyzed using log-rank and Fisher exact tests. INSL4 functions were studied using short hairpin RNA (shRNA) knockdown, overexpression, transcriptome profiling, cell growth, and survival assays in vitro and in vivo. All statistical tests were two-sided. Results INSL4 was identified as a novel downstream target of LKB1 deficiency and its expression was induced through aberrant CRTC-CREB activation. INSL4 was highly induced in LKB1-deficient NSCLC cells (up to 543-fold) and 9 of 41 primary tumors, although undetectable in all normal tissues except the placenta. Lung adenocarcinomas from The Cancer Genome Atlas with high and low INSL4 expression (with the top 10th percentile as cutoff) showed statistically significant differences for advanced tumor stage (P < .001), lymph node metastasis (P = .001), and tumor size (P = .01). The INSL4-high group showed worse survival than the INSL4-low group (P < .001). Sustained INSL4 expression was required for the growth and viability of LKB1-inactivated NSCLC cells in vitro and in a mouse xenograft model (n = 5 mice per group). Expression profiling revealed INSL4 as a critical regulator of cell cycle, growth, and survival. Conclusions LKB1 deficiency induces an autocrine INSL4 signaling that critically supports the growth and survival of lung cancer cells. Therefore, aberrant INSL4 signaling is a promising therapeutic target for LKB1-deficient lung cancers.

scholarly journals Identification of the Vibrio cholerae Enterobactin Receptors VctA and IrgA: IrgA Is Not Required for Virulence

2002 ◽  
Vol 70 (7) ◽  
pp. 3419-3426 ◽  
Author(s):  
Alexandra R. Mey ◽  
Elizabeth E. Wyckoff ◽  
Amanda G. Oglesby ◽  
Eva Rab ◽  
Ronald K. Taylor ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Mutant Strain ◽  
Genomic Sequence ◽  
Iron Acquisition ◽  
Lethal Dose ◽  
Single Mutation ◽  
Periplasmic Binding Protein ◽  
Infant Mouse ◽  
Abc Transport ◽  
Important Virulence Factor

ABSTRACT The gram-negative enteric pathogen Vibrio cholerae requires iron for growth. V. cholerae has multiple iron acquisition systems, including utilization of heme and hemoglobin, synthesis and transport of the catechol siderophore vibriobactin, and transport of several siderophores that it does not itself make. One siderophore that V. cholerae transports, but does not make, is enterobactin. Enterobactin transport requires TonB and is independent of the vibriobactin receptor ViuA. In this study, two candidate enterobactin receptor genes, irgA (VC0475) and vctA (VCA0232), were identified by analysis of the V. cholerae genomic sequence. A single mutation in either of these genes did not significantly impair enterobactin utilization, but a strain defective in both genes did not use enterobactin. When either irgA or vctA was supplied on a plasmid, the ability of the irgA vctA double mutant to use enterobactin was restored. This indicates that both VctA and IrgA transport enterobactin. We also identify the genes vctPDGC, which are linked to vctA and encode a periplasmic binding protein-dependent ABC transport system that functions in the utilization of both enterobactin and vibriobactin (VCA0227-0230). An irgA::TnphoA mutant strain, MBG40, was shown in a previous study to be highly attenuated and to have a strong colonization defect in an infant mouse model of V. cholerae infection (M. B. Goldberg, V. J. DiRita, and S. B. Calderwood, Infect. Immun. 58:55-60, 1990). In this work, a new irgA mutation was constructed, and this mutant strain was not significantly impaired in its ability to compete with the parental strain in infant mice and was not attenuated for virulence in an assay of 50% lethal dose. These data indicate that the virulence defect in MBG40 is not due to the loss of irgA function and that irgA is unlikely to be an important virulence factor.

scholarly journals Role of mechanical cues and hypoxia on the growth of tumor cells in strong and weak confinement: A dual in vitro–in silico approach

2020 ◽  
Vol 6 (13) ◽  
pp. eaaz7130 ◽  
Author(s):  
V. Le Maout ◽  
K. Alessandri ◽  
B. Gurchenkov ◽  
H. Bertin ◽  
P. Nassoy ◽  
...  
Keyword(s):  
In Silico ◽  
Time Course ◽  
Growth Dynamics ◽  
Tumor Model ◽  
Mechanistic Modeling ◽  
Physical Factors ◽  
Factors Affecting ◽  
Numerical Studies

Characterization of tumor growth dynamics is of major importance for cancer understanding. By contrast with phenomenological approaches, mechanistic modeling can facilitate disclosing underlying tumor mechanisms and lead to identification of physical factors affecting proliferation and invasive behavior. Current mathematical models are often formulated at the tissue or organ scale with the scope of a direct clinical usefulness. Consequently, these approaches remain empirical and do not allow gaining insight into the tumor properties at the scale of small cell aggregates. Here, experimental and numerical studies of the dynamics of tumor aggregates are performed to propose a physics-based mathematical model as a general framework to investigate tumor microenvironment. The quantitative data extracted from the cellular capsule technology microfluidic experiments allow a thorough quantitative comparison with in silico experiments. This dual approach demonstrates the relative impact of oxygen and external mechanical forces during the time course of tumor model progression.

Rv2223c, an acid inducible carboxyl-esterase of Mycobacterium tuberculosis enhanced the growth and survival of Mycobacterium smegmatis

2019 ◽  
Vol 14 (16) ◽  
pp. 1397-1415
Author(s):  
Pratibha Maan ◽  
Jagdeep Kaur
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mycobacterium Smegmatis ◽  
Intracellular Survival ◽  
Growth And Survival ◽  
Enhanced Expression ◽  
Stress Environment ◽  
In Vitro Stress ◽  
Carboxyl Esterase

Aim: To elucidate the role of Rv2223c in Mycobacterium tuberculosis. Methods: Purified recombinant Rv2223c protein was characterized. Expression of rv2223c in the presence of different stress environment and subcellular localization were performed in M. tuberculosis H37Ra and Mycobacterium smegmatis ( MS_2223c). Effect of its overexpression on growth rate, infection and intracellular survival in THP-1/PBMC cells were studied. Results: rRv2223c demonstrated esterase activity with preference for pNP-octanoate and hydrolyzed trioctanoate to di- and mono-octanoate. Expression of rv2223c was upregulated in acidic and nutritive stress conditions. rRv2223c was identified in extracellular and cell wall fractions. MS_2223c exhibited enhanced growth, survival during in vitro stress, infection and intracellular survival. Conclusions: Rv2223c is a secretary, carboxyl-esterase, with enhanced expression under acidic and nutritive stress condition and might help in intracellular survival of bacteria.

Sign in / Sign up

Export Citation Format

Share Document