scholarly journals Identification of Listeria monocytogenes genes contributing to oxidative stress resistance under conditions relevant to host infection

2021 ◽  
Author(s):  
David R. Mains ◽  
Samuel J. Eallonardo ◽  
Nancy E. Freitag
Keyword(s):  
Oxidative Stress ◽  
Listeria Monocytogenes ◽  
Virulence Factors ◽  
Stress Resistance ◽  
Mammalian Cells ◽  
Bacterial Virulence ◽  
Host Cells ◽  
Physiological Adaptation ◽  
Wild Type ◽  
Oxidative Stress Resistance

The Gram-positive bacterium Listeria monocytogenes survives in environments ranging from the soil to the cytosol of infected host cells. Key to L. monocytogenes intracellular survival is the activation of PrfA, a transcriptional regulator that is required for the expression of multiple bacterial virulence factors. Mutations that constitutively activate prfA (prfA* mutations) result in high-level expression of multiple bacterial virulence factors as well as the physiological adaptation of L. monocytogenes for optimal replication within host cells. Here we demonstrate that L. monocytogenes prfA* mutants exhibit significantly enhanced resistance to oxidative stress in comparison to wild type strains. Transposon mutagenesis of L. monocytogenes prfA* strains resulted in the identification of three novel gene targets required for full oxidative stress resistance only in the context of PrfA activation. One gene, lmo0779, predicted to encode an uncharacterized protein, and two additional genes known as cbpA and ygbB, encoding a cyclic-di-AMP binding protein and a 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase respectively, contribute to the enhanced oxidative stress resistance of prfA* strains while exhibiting no significant contribution in wild type L. monocytogenes. Transposon inactivation of cbpA and lmo0779 in a prfA* background led to reduced virulence in the liver of infected mice. These results indicate that L. monocytogenes calls upon specific bacterial factors for stress resistance in the context of PrfA activation and thus under conditions favorable for bacterial replication within infected mammalian cells.

scholarly journals Proline Metabolism IncreaseskatGExpression and Oxidative Stress Resistance in Escherichia coli

2014 ◽  
Vol 197 (3) ◽  
pp. 431-440 ◽  
Author(s):  
Lu Zhang ◽  
James R. Alfano ◽  
Donald F. Becker
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Hydrogen Peroxide ◽  
Mutant Strain ◽  
Stress Resistance ◽  
Proline Metabolism ◽  
Wild Type ◽  
Content Type ◽  
Oxidative Stress Resistance ◽  
Proline Utilization

The oxidation ofl-proline to glutamate in Gram-negative bacteria is catalyzed by the proline utilization A (PutA) flavoenzyme, which contains proline dehydrogenase (PRODH) and Δ1-pyrroline-5-carboxylate (P5C) dehydrogenase domains in a single polypeptide. Previous studies have suggested that aside from providing energy, proline metabolism influences oxidative stress resistance in different organisms. To explore this potential role and the mechanism, we characterized the oxidative stress resistance of wild-type andputAmutant strains ofEscherichia coli. Initial stress assays revealed that theputAmutant strain was significantly more sensitive to oxidative stress than the parental wild-type strain. Expression of PutA in theputAmutant strain restored oxidative stress resistance, confirming that depletion of PutA was responsible for the oxidative stress phenotype. Treatment of wild-type cells with proline significantly increased hydroperoxidase I (encoded bykatG) expression and activity. Furthermore, the ΔkatGstrain failed to respond to proline, indicating a critical role for hydroperoxidase I in the mechanism of proline protection. The global regulator OxyR activates the expression ofkatGalong with several other genes involved in oxidative stress defense. In addition tokatG, proline increased the expression ofgrxA(glutaredoxin 1) andtrxC(thioredoxin 2) of the OxyR regulon, implicating OxyR in proline protection. Proline oxidative metabolism was shown to generate hydrogen peroxide, indicating that proline increases oxidative stress tolerance inE. colivia a preadaptive effect involving endogenous hydrogen peroxide production and enhanced catalase-peroxidase activity.

scholarly journals Manganese regulation of virulence factors and oxidative stress resistance in Neisseria gonorrhoeae

2010 ◽  
Vol 73 (5) ◽  
pp. 899-916 ◽  
Author(s):  
Hsing-Ju Wu ◽  
Kate L. Seib ◽  
Yogitha N. Srikhanta ◽  
Jennifer Edwards ◽  
Stephen P. Kidd ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Neisseria Gonorrhoeae ◽  
Virulence Factors ◽  
Stress Resistance ◽  
Oxidative Stress Resistance ◽  
And Oxidative Stress ◽  
Manganese Regulation

scholarly journals Intersection of phosphate transport, oxidative stress and TOR signalling inCandida albicansvirulence

2018 ◽  
Author(s):  
Ning-Ning Liu ◽  
Priya Uppuluri ◽  
Achille Broggi ◽  
Angelique Besold ◽  
Kicki Ryman ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stress Management ◽  
Stress Resistance ◽  
Ex Vivo ◽  
Growth Defect ◽  
Invasive Infection ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
Ros Accumulation

AbstractPhosphate is an essential macronutrient required for cell growth and division. Pho84 is the major high-affinity cell-surface phosphate importer ofSaccharomyces cerevisiaeand a crucial element in the phosphate homeostatic system of this model yeast. We found that loss ofCandida albicansPho84 attenuated virulence inDrosophilaand murine oropharyngeal and disseminated models of invasive infection, and conferred hypersensitivity to neutrophil killing. Susceptibility of cells lacking Pho84 to neutrophil attack depended on reactive oxygen species (ROS):pho84-/-cells were no more susceptible than wild typeC. albicansto neutrophils from a patient with chronic granulomatous disease, or to those whose oxidative burst was pharmacologically inhibited or neutralized.pho84-/-mutants hyperactivated oxidative stress signalling. They accumulated intracellular ROS in the absence of extrinsic oxidative stress, in high as well as low ambient phosphate conditions. ROS accumulation correlated with diminished levels of the unique superoxide dismutase Sod3 inpho84-/-cells, whileSOD3overexpression from a conditional promoter substantially restored these cells’ oxidative stress resistance in vitro. Repression ofSOD3expression sharply increased their oxidative stress hypersensitivity. Neither of these oxidative stress management effects of manipulatingSOD3transcription was observed inPHO84wild type cells. Sod3 levels were not the only factor driving oxidative stress effects onpho84-/-cells, though, because overexpressingSOD3did not ameliorate these cells’ hypersensitivity to neutrophil killing ex vivo, indicating Pho84 has further roles in oxidative stress resistance and virulence. Measurement of cellular metal concentrations demonstrated that diminished Sod3 expression was not due to decreased import of its metal cofactor manganese, as predicted from the function ofS. cerevisiaePho84 as a low-affinity manganese transporter. Instead of a role of Pho84 in metal transport, we found its role in TORC1 activation to impact oxidative stress management: overexpression of the TORC1-activating GTPase Gtr1 relieved the Sod3 deficit and ROS excess inpho84-/-null mutant cells, though it did not suppress their hypersensitivity to neutrophil killing or hyphal growth defect. Pharmacologic inhibition of Pho84 by small molecules including the FDA-approved drug foscarnet also induced ROS accumulation. Inhibiting Pho84 could hence support host defenses by sensitizingC. albicansto oxidative stress.

scholarly journals Role of ςB in Heat, Ethanol, Acid, and Oxidative Stress Resistance and during Carbon Starvation inListeria monocytogenes

2001 ◽  
Vol 67 (10) ◽  
pp. 4454-4457 ◽  
Author(s):  
Adriana Ferreira ◽  
Conor P. O'Byrne ◽  
Kathryn J. Boor
Keyword(s):  
Oxidative Stress ◽  
Stationary Phase ◽  
Stress Resistance ◽  
Parent Strain ◽  
Acid Resistance ◽  
Carbon Starvation ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
And Oxidative Stress ◽  
Dependent Mechanism

ABSTRACT To determine the contribution of sigma B (ςB) to survival of stationary-phase Listeria monocytogenescells following exposure to environmental stresses, we compared the viability of strain 10403S with that of an isogenic nonpolarsigB null mutant strain after exposure to heat (50°C), ethanol (16.5%), or acid (pH 2.5). Strain viabilities were also determined under the same conditions in cultures that had been previously exposed to sublethal levels of the same stresses (45°C, 5% ethanol, or pH 4.5). The ΔsigB and wild-type strains had similar viabilities following exposure to ethanol and heat, but the ΔsigB strain was almost 10,000-fold more susceptible to lethal acid stress than its parent strain. However, a 1-h preexposure to pH 4.5 yielded a 1,000-fold improvement in viability for the ΔsigB strain. These results suggest the existence in L. monocytogenes of both a ςB-dependent mechanism and a pH-dependent mechanism for acid resistance in the stationary phase. ςB contributed to resistance to both oxidative stress and carbon starvation inL. monocytogenes. The ΔsigB strain was 100-fold more sensitive to 13.8 mM cumene hydroperoxide than the wild-type strain. Following glucose depletion, the ΔsigB strain lost viability more rapidly than the parent strain. ςB contributions to viability during carbon starvation and to acid resistance and oxidative stress resistance support the hypothesis that ςB plays a role in protecting L. monocytogenes against environmental adversities.

scholarly journals Contribution of the Helicobacter pylori Thiol Peroxidase Bacterioferritin Comigratory Protein to Oxidative Stress Resistance and Host Colonization

2005 ◽  
Vol 73 (1) ◽  
pp. 378-384 ◽  
Author(s):  
Ge Wang ◽  
Adriana A. Olczak ◽  
James P. Walton ◽  
Robert J. Maier
Keyword(s):  
Oxidative Stress ◽  
Mutant Strain ◽  
Stress Resistance ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
Organic Hydroperoxides ◽  
H Pylori ◽  
Thiol Peroxidase

ABSTRACT Peroxiredoxins, the enzymes that catalyze the reduction of hydrogen peroxide and organic hydroperoxides, are ubiquitous proteins that protect organisms from damage by reactive oxygen species. Helicobacter pylori contains three members of the peroxiredoxin family: AhpC (alkyl hydroperoxide reductase), Tpx (thiol-specific peroxidase), and bacterioferritin comigratory protein (BCP). In this study, we characterized H. pylori bcp mutant strains and wild-type BCP. Compared to the parent strain and the ahpC mutant strain, the bcp mutant showed moderate sensitivity to the superoxide-generating agent paraquat and to organic hydroperoxides. Upon exposure of 108 cells to air for 10 h, 106 wild-type cells survived but none of the 108 bcp mutant cells were recovered. Introduction of an intact bcp gene at an unrelated locus in the bcp strain restored the wild-type-like oxidative stress resistance phenotype. Purified BCP was shown to be a thiol peroxidase that depends on the reducing activity of thioredoxin and thioredoxin reductase. Among a series of peroxides tested, linoleic acid hydroperoxide was the preferred substrate of BCP. By examining the profiles of protein expression within H. pylori cells, we confirmed that AhpC is much more abundant than BCP. The overlapping functions and activities of BCP and AhpC probably explain why the bcp mutant displayed a relatively weak oxidative stress resistance phenotype. The bcp mutant strain could colonize mouse stomachs, although colonization by the wild-type strain was slightly better than that by the mutant strain at 1 week after host inoculation. However, at 3 weeks after inoculation, the colonization ability of the wild type was significantly greater than that of the bcp mutant; for example, H. pylori was recovered from 10 of 11 mouse stomachs inoculated with the wild-type strain but from only 4 of 12 mice that were inoculated with the bcp mutant strain. This indicates that H. pylori BCP plays a significant role in efficient host colonization.

scholarly journals Oxidative-Stress Resistance Mutants of Helicobacter pylori

2002 ◽  
Vol 184 (12) ◽  
pp. 3186-3193 ◽  
Author(s):  
Adriana A. Olczak ◽  
Jonathan W. Olson ◽  
Robert J. Maier
Keyword(s):  
Oxidative Stress ◽  
Stress Resistance ◽  
Double Mutant ◽  
Mutation Frequency ◽  
Spontaneous Mutation ◽  
Type I ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
Spontaneous Mutation Frequency ◽  
Resistance Mutants

ABSTRACT Within a large family of peroxidases, one member that catalyzes the reduction of organic peroxides to alcohols is known as alkyl hydroperoxide reductase, or AhpC. Gene disruption mutations in the gene encoding AhpC of Helicobacter pylori (ahpC) were generated by screening transformants under low-oxygen conditions. Two classes of mutants were obtained. Both types lack AhpC protein, but the major class (type I) isolated was found to synthesize increased levels (five times more than the wild type) of another proposed antioxidant protein, an iron-binding, neutrophil-activating protein (NapA). The other class of mutants, the minor class (type II), produced wild-type levels of NapA. The two types of AhpC mutants differed in their frequencies of spontaneous mutation to rifampin resistance and in their sensitivities to oxidative-stress chemicals, with the type I mutants exhibiting less sensitivity to organic hydroperoxides as well as having a lower mutation frequency. The napA promoter regions of the two types of AhpC mutants were identical, and primer extension analysis revealed their transcription start site to be the same as for the wild type. Gene disruption mutations were obtained in napA alone, and a double mutant strain (ahpC napA) was also created. All four of the oxidative-stress resistance mutants could be distinguished from the wild type in oxygen sensitivity or in some other oxidative-stress resistance phenotype (i.e., in sensitivity to stress-related chemicals and spontaneous mutation frequency). For example, growth of the NapA mutant was more sensitive to oxygen than that of the wild-type strain and both of the AhpC-type mutants were highly sensitive to paraquat and to cumene hydroperoxide. Of the four types of mutants, the double mutant was the most sensitive to growth inhibition by oxygen and by organic peroxides and it had the highest spontaneous mutation frequency. Notably, two-dimensional gel electrophoresis combined with protein sequence analysis identified another possible oxidative-stress resistance protein (HP0630) that was up-regulated in the double mutant. However, the transcription start site of the HP0630 gene was the same for the double mutant as for the wild type. It appears that H. pylori can readily modulate the expression of other resistance factors as a compensatory response to loss of a major oxidative-stress resistance component.

scholarly journals Gengnianchun, a Traditional Chinese Medicine, Enhances Oxidative Stress Resistance and Lifespan in Caenorhabditis elegans by Modulating daf-16/FOXO

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
Author(s):  
Fanhui Meng ◽  
Jun Li ◽  
Wenjun Wang ◽  
Yan Fu
Keyword(s):  
Oxidative Stress ◽  
Caenorhabditis Elegans ◽  
Chinese Medicine ◽  
Traditional Chinese Medicine ◽  
Stress Resistance ◽  
Transgenic Animals ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
C Elegans ◽  
And Oxidative Stress

Objective. Gengnianchun (GNC), a traditional Chinese medicine (TCM), is primarily used to improve declining functions related to aging. In this study, we investigated its prolongevity and stress resistance properties and explored the associated regulatory mechanism using a Caenorhabditis elegans model. Methods. Wild-type C. elegans N2 was used for lifespan analysis and oxidative stress resistance assays. Transgenic animals were used to investigate pathways associated with antioxidative stress activity. The effects of GNC on levels of reactive oxygen species (ROS) and expression of specific genes were examined. Results. GNC-treated wild-type worms showed an increase in survival time under both normal and oxidative stress conditions. GNC decreased intracellular ROS levels by 67.95%. GNC significantly enhanced the oxidative stress resistance of several mutant strains, suggesting that the protective effect of GNC is independent of the function of these genes. However, the oxidative stress resistance effect of GNC was absent in worms with daf-16 mutation. We also found upregulation of daf-16 downstream targets including sod-3 and mtl-1. Conclusions. Our findings suggest that GNC extends the lifespan of C. elegans and enhances its resistance to oxidative stress via a daf-16/FOXO-dependent pathway. This study also provides a feasible method for screening the biological mechanisms of TCMs.

scholarly journals Loss of SigB in Listeria monocytogenes Strains EGD-e and 10403S Confers Hyperresistance to Hydrogen Peroxide in Stationary Phase under Aerobic Conditions

2016 ◽  
Vol 82 (15) ◽  
pp. 4584-4591 ◽  
Author(s):  
Marcia Boura ◽  
Ciara Keating ◽  
Kevin Royet ◽  
Ranju Paudyal ◽  
Beth O'Donoghue ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Listeria Monocytogenes ◽  
Stationary Phase ◽  
Stress Resistance ◽  
Wild Type ◽  
Content Type ◽  
Stress Genes ◽  
Reverse Transcription Pcr ◽  
Stress Gene

ABSTRACTSigB is the main stress gene regulator inListeria monocytogenesaffecting the expression of more than 150 genes and thus contributing to multiple-stress resistance. Despite its clear role in most stresses, its role in oxidative stress is uncertain, as results accompanying the loss ofsigBrange from hyperresistance to hypersensitivity. Previously, these differences have been attributed to strain variation. In this study, we show conclusively that unlike for all other stresses, loss ofsigBresults in hyperresistance to H2O2(more than 8 log CFU ml−1compared to the wild type) in aerobically grown stationary-phase cultures ofL. monocytogenesstrains 10403S and EGD-e. Furthermore, growth at 30°C resulted in higher resistance to oxidative stress than that at 37°C. Oxidative stress resistance seemed to be higher with higher levels of oxygen. Under anaerobic conditions, the loss of SigB in 10403S did not affect survival against H2O2, while in EGD-e, it resulted in a sensitive phenotype. During exponential phase, minor differences occurred, and this result was expected due to the absence ofsigBtranscription. Catalase tests were performed under all conditions, and stronger catalase results corresponded well with a higher survival rate, underpinning the important role of catalase in this phenotype. Furthermore, we assessed the catalase activity in protein lysates, which corresponded with the catalase tests and survival. In addition, reverse transcription-PCR (RT-PCR) showed no differences in transcription between the wild type and the ΔsigBmutant in various oxidative stress genes. Further investigation of the molecular mechanism behind this phenotype and its possible consequences for the overall phenotype ofL. monocytogenesare under way.IMPORTANCESigB is the most important stress gene regulator inL. monocytogenesand other Gram-positive bacteria. Its increased expression during stationary phase results in resistance to multiple stresses. However, despite its important role in general stress resistance, its expression is detrimental for the cell in the presence of oxidative stress, as it promotes hypersensitivity against hydrogen peroxide. This peculiar phenotype is an important element of the physiology ofL. monocytogenes, and it might help us explain the behavior of this organism in environments where oxidative stress is present.

scholarly journals An NADPH Quinone Reductase of Helicobacter pylori Plays an Important Role in Oxidative Stress Resistance and Host Colonization

2004 ◽  
Vol 72 (3) ◽  
pp. 1391-1396 ◽  
Author(s):  
Ge Wang ◽  
Robert J. Maier
Keyword(s):  
Oxidative Stress ◽  
Helicobacter Pylori ◽  
Mutant Strain ◽  
Stress Resistance ◽  
Type Strain ◽  
Wild Type Strain ◽  
Quinone Reductase ◽  
Wild Type ◽  
Oxidative Stress Resistance ◽  
H Pylori

ABSTRACT Oxidative stress resistance is one of the key properties that enable pathogenic bacteria to survive the toxic reactive oxygen species released by the host. In a previous study characterizing oxidative stress resistance mutants of Helicobacter pylori, a novel potential antioxidant protein (MdaB) was identified by the observation that the expression of this protein was significantly upregulated to compensate for the loss of other major antioxidant components. In this study, we characterized an H. pylori mdaB mutant and the MdaB protein. While the wild-type strain can tolerate 10% oxygen for growth, the growth of the mdaB mutant was significantly inhibited by this oxygen condition. The mdaB mutant is also more sensitive to H2O2, organic hydroperoxides, and the superoxide-generating agent paraquat. Although the wild-type strain can survive more than 10 h of air exposure, exposure of the mutant strain to air for 8 h resulted in recovery of no viable cells. The oxidative stress sensitivity of the mdaB mutant resulted in a deficiency in the ability to colonize mouse stomachs. H. pylori was recovered from 10 of 11 mouse stomachs inoculated with the wild-type strain, with about 5,000 to 45,000 CFU/g of stomach. However, only 3 of 12 mice that were inoculated with the mdaB mutant strain were found to harbor any H. pylori, and these 3 contained less than 2,000 CFU/g of stomach. A His-tagged MdaB protein was purified and characterized. It was shown to be a flavoprotein that catalyzes two-electron transfer from NAD(P)H to quinones. It reduces both ubiquinones and menaquinones with similar efficiencies and preferably uses NADPH as an electron donor. We propose that the physiological function of the H. pylori MdaB protein is that of an NADPH quinone reductase that plays an important role in managing oxidative stress and contributes to successful colonization of the host.

scholarly journals Structural and functional analysis of the Francisella lysine decarboxylase as a key actor in oxidative stress resistance

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jan Felix ◽  
Claire Siebert ◽  
Julia Novion Ducassou ◽  
Jérôme Nigou ◽  
Pierre Simon Garcia ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stress Response ◽  
Stress Resistance ◽  
Pathogenic Bacteria ◽  
Host Cells ◽  
Acid Decarboxylase ◽  
Growth Media ◽  
Lysine Decarboxylase ◽  
Oxidative Stress Resistance

AbstractFrancisella tularensis is one of the most virulent pathogenic bacteria causing the acute human respiratory disease tularemia. While the mechanisms underlying F. tularensis pathogenesis are largely unknown, previous studies have shown that a F. novicida transposon mutant with insertions in a gene coding for a putative lysine decarboxylase was attenuated in mouse spleen, suggesting a possible role of its protein product as a virulence factor. Therefore, we set out to structurally and functionally characterize the F. novicida lysine decarboxylase, which we termed LdcF. Here, we investigate the genetic environment of ldcF as well as its evolutionary relationships with other basic AAT-fold amino acid decarboxylase superfamily members, known as key actors in bacterial adaptative stress response and polyamine biosynthesis. We determine the crystal structure of LdcF and compare it with the most thoroughly studied lysine decarboxylase, E. coli LdcI. We analyze the influence of ldcF deletion on bacterial growth under different stress conditions in dedicated growth media, as well as in infected macrophages, and demonstrate its involvement in oxidative stress resistance. Finally, our mass spectrometry-based quantitative proteomic analysis enables identification of 80 proteins with expression levels significantly affected by ldcF deletion, including several DNA repair proteins potentially involved in the diminished capacity of the F. novicida mutant to deal with oxidative stress. Taken together, we uncover an important role of LdcF in F. novicida survival in host cells through participation in oxidative stress response, thereby singling out this previously uncharacterized protein as a potential drug target.

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