scholarly journals Role for HtrA in Stress Induction and Virulence Potential in Listeria monocytogenes

2005 ◽  
Vol 71 (8) ◽  
pp. 4241-4247 ◽  
Author(s):  
Helena M. Stack ◽  
Roy D. Sleator ◽  
Megan Bowers ◽  
Colin Hill ◽  
Cormac G. M. Gahan
Keyword(s):  
Listeria Monocytogenes ◽  
Heat Shock ◽  
Serine Protease ◽  
Stress Responses ◽  
Signal Sequence ◽  
Transcriptional Analysis ◽  
Start Codon ◽  
Penicillin G ◽  
Insertion Mutant ◽  
Virulence Potential

ABSTRACT In silico analysis of the Listeria monocytogenes genome revealed lmo0292, a gene predicted to encode a HtrA-like serine protease. A stable insertion mutant was constructed, revealing a requirement for htrA in the listerial response to heat, acid, and penicillin stress. Transcriptional analysis revealed that htrA is not induced in response to heat shock but is induced in response to low pH and penicillin G stress. Furthermore, htrA expression was shown to be dependent upon the LisRK two-component sensor-kinase, a system known to respond to changes in integrity of the cell envelope. In addition, we demonstrated that a second in-frame start codon, upstream of that previously annotated for L. monocytogenes htrA, incorporating a putative signal sequence appears to influence virulence potential. Finally, a significant virulence defect was observed for the htrA mutant, indicating that this gene is required for full virulence in mice. Our findings suggest that L. monocytogenes lmo0292 encodes an HtrA-like serine protease that is not part of the classical heat shock response but is involved in stress responses and virulence.

scholarly journals Global Transcriptional Analysis of clpP Mutations of Type 2 Streptococcus pneumoniae and Their Effects on Physiology and Virulence

2002 ◽  
Vol 184 (13) ◽  
pp. 3508-3520 ◽  
Author(s):  
Gregory T. Robertson ◽  
Wai-Leung Ng ◽  
Joseph Foley ◽  
Raymond Gilmour ◽  
Malcolm E. Winkler
Keyword(s):  
Oxidative Stress ◽  
Streptococcus Pneumoniae ◽  
Heat Shock ◽  
Microarray Analysis ◽  
Stress Responses ◽  
Metal Ion ◽  
Transcriptional Analysis ◽  
Exponential Growth Phase ◽  
Dna Uptake

ABSTRACT Streptococcus pneumoniae is an important human pathogen that contains single copies of genes encoding the ClpP and FtsH ATP-dependent proteases but lacks the Lon and HslV proteases. We constructed and characterized the phenotypes of clpP, clpC, and clpX deletion replacement mutants, which lack the ClpP protease subunit or the putative ClpC or ClpX ATPase specificity factor. A ΔclpP mutant, but not a ΔclpC or ΔclpX mutant, of the virulent D39 type 2 strain of S. pneumoniae grew poorly at 30°C and failed to grow at 40°C. Despite this temperature sensitivity, transcription of the heat shock regulon determined by microarray analysis was induced in a ΔclpP mutant, which was also more sensitive to oxidative stress by H2O2 and to puromycin than its clpP + parent strain. A ΔclpP mutant, but not a ΔclpC mutant, was strongly attenuated for virulence in the murine lung and sepsis infection models. All of these phenotypes were complemented in a ΔclpP/clpP + merodiploid strain. Consistent with these complementation patterns, clpP was found to be in a monocistronic operon, whose transcription was induced about fivefold by heat shock in S. pneumoniae as determined by Northern and real-time reverse transcription-PCR analyses. Besides clpP, transcription of clpC, clpE, and clpL, but not clpX or ftsH, was induced by heat shock or entry into late exponential growth phase. Microarray analysis of ΔclpP mutants showed a limited change in transcription pattern (≈80 genes) consistent with these phenotypes, including repression of genes involved in oxidative stress, metal ion transport, and virulence. In addition, transcription of the early and late competence regulon was induced in the ΔclpP mutant, and competence gene expression and DNA uptake seemed to be constitutively induced throughout growth. Together, these results indicate that ClpP-mediated proteolysis plays a complex and central role in numerous pneumococcal stress responses, development of competence, and virulence.

scholarly journals Contribution of Three Bile-Associated Loci, bsh, pva, and btlB, to Gastrointestinal Persistence and Bile Tolerance of Listeria monocytogenes

2005 ◽  
Vol 73 (2) ◽  
pp. 894-904 ◽  
Author(s):  
Máire Begley ◽  
Roy D. Sleator ◽  
Cormac G. M. Gahan ◽  
Colin Hill
Keyword(s):  
Listeria Monocytogenes ◽  
Bile Salt ◽  
Stress Responses ◽  
Bile Salts ◽  
Molecular Mechanisms ◽  
Penicillin G ◽  
Bile Salt Hydrolase ◽  
Penicillin V ◽  
Resistance To Penicillin ◽  
Intestinal Persistence

ABSTRACT Listeria monocytogenes must resist the deleterious actions of bile in order to infect and subsequently colonize the human gastrointestinal tract. The molecular mechanisms used by the bacterium to resist bile and the influence of bile on pathogenesis are as yet largely unexplored. This study describes the analysis of three genes—bsh, pva, and btlB—previously annotated as bile-associated loci in the sequenced L. monocytogenes EGDe genome (lmo2067, lmo0446, and lmo0754, respectively). Analysis of deletion mutants revealed a role for all three genes in resisting the acute toxicity of bile and bile salts, particularly glycoconjugated bile salts at low pH. Mutants were unaffected in the other stress responses examined (acid, salt, and detergents). Bile hydrolysis assays demonstrate that L. monocytogenes possesses only one bile salt hydrolase gene, namely, bsh. Transcriptional analyses and activity assays revealed that, although it is regulated by both PrfA and σB, the latter appears to play the greater role in modulating bsh expression. In addition to being incapable of bile hydrolysis, a sigB mutant was shown to be exquisitely sensitive to bile salts. Furthermore, increased expression of sigB was detected under anaerobic conditions and during murine infection. A gene previously annotated as a possible penicillin V amidase (pva) or bile salt hydrolase was shown to be required for resistance to penicillin V but not penicillin G but did not demonstrate a role in bile hydrolysis. Finally, animal (murine) studies revealed an important role for both bsh and btlB in the intestinal persistence of L. monocytogenes.

scholarly journals clpB, a Novel Member of the Listeria monocytogenes CtsR Regulon, Is Involved in Virulence but Not in General Stress Tolerance

2004 ◽  
Vol 186 (4) ◽  
pp. 1165-1174 ◽  
Author(s):  
Arnaud Chastanet ◽  
Isabelle Derre ◽  
Shamila Nair ◽  
Tarek Msadek
Keyword(s):  
Listeria Monocytogenes ◽  
Heat Shock ◽  
Model Organism ◽  
Transcriptional Analysis ◽  
Gram Positive ◽  
General Stress ◽  
Potential Binding ◽  
Induced Thermotolerance

ABSTRACT Clp-HSP100 ATPases are a widespread family of ubiquitous proteins that occur in both prokaryotes and eukaryotes and play important roles in the folding of newly synthesized proteins and refolding of aggregated proteins. They have also been shown to participate in the virulence of several pathogens, including Listeria monocytogenes. Here, we describe a member of the Clp-HSP100 family of L. monocytogenes that harbors all the characteristics of the ClpB subclass, which is absent in the closely related gram-positive model organism, Bacillus subtilis. Transcriptional analysis of clpB revealed a heat shock-inducible σA-type promoter. Potential binding sites for the CtsR regulator of stress response were identified in the promoter region. In vivo and in vitro approaches were used to show that expression of clpB is repressed by CtsR, a finding indicating that clpB is a novel member of the L. monocytogenes CtsR regulon. We showed that ClpB is involved in the pathogenicity of L. monocytogenes since the ΔclpB mutant is significantly affected by virulence in a murine model of infection; we also demonstrate that this effect is apparently not due to a defect in general stress resistance. Indeed, ClpB is not involved in tolerance to heat, salt, detergent, puromycin, or cold stress, even though its synthesis is inducible by heat shock. However, ClpB was shown to play a role in induced thermotolerance, allowing increased resistance of L. monocytogenes to lethal temperatures. This work gives the first example of a clpB gene directly controlled by CtsR and describes the first role for a ClpB protein in induced thermotolerance and virulence in a gram-positive organism.

Extracellular Proteins as Enterobacterial Thermometers

2003 ◽  
Vol 86 (1-2) ◽  
pp. 139-156 ◽  
Author(s):  
Robin J. Rowbury
Keyword(s):  
Thermal Stress ◽  
Heat Shock ◽  
Stress Responses ◽  
Direct Evidence ◽  
Experimental Studies ◽  
Extracellular Proteins ◽  
Temperature Changes ◽  
Induction Components ◽  
Cellular Components ◽  
Thermal Stimuli

Biological thermometers are cellular components or structures which sense increasing temperatures, interaction of the thermometer and the thermal stress bringing about the switching-on of inducible responses, with gradually enhanced levels of response induction following gradually increasing temperatures. In enterobacteria, for studies of such thermometers, generally induction of heat shock protein (HSP) synthesis has been examined, with experimental studies aiming to establish (often indirectly) how the temperature changes which initiate HSP synthesis are sensed; numerous other processes and responses show graded induction as temperature is increased, and how the temperature changes which induce these are sensed is also of interest. Several classes of intracellular component and structure have been proposed as enterobacterial thermometers, with the ribosome and the DnaK chaperone being the most favoured, although for many of the proposed intracellular thermometers, most of the evidence for their functioning in this way is indirect. In contrast to the above, the studies reviewed here firmly establish that for four distinct stress responses, which are switched-on gradually as temperature increases, temperature changes are sensed by extracellular components (extracellular sensing components, ESCs) i.e. there is firm and direct evidence for the occurrence of extracellular thermometers. All four thermometers described here are proteins, which appear to be distinct and different from each other, and on sensing thermal stress are activated by it to four distinct extracellular induction components (EICs), which interact with receptors on the surface of organisms to induce the appropriate responses. It is predicted that many other temperature-induced processes, including the synthesis of HSPs, will be switched-on following the activation of similar extracellular thermometers by thermal stimuli.

scholarly journals Arabidopsis heat shock transcription factor HSFA7b positively mediates salt stress tolerance by binding to an E-box-like motif to regulate gene expression

2019 ◽  
Vol 70 (19) ◽  
pp. 5355-5374 ◽  
Author(s):  
Dandan Zang ◽  
Jingxin Wang ◽  
Xin Zhang ◽  
Zhujun Liu ◽  
Yucheng Wang
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Heat Shock ◽  
Salt Tolerance ◽  
Stress Responses ◽  
Ion Homeostasis ◽  
Cellulose Synthase ◽  
Regulate Gene Expression ◽  
E Box ◽  
Regulate Gene

Abstract Plant heat shock transcription factors (HSFs) are involved in heat and other abiotic stress responses. However, their functions in salt tolerance are little known. In this study, we characterized the function of a HSF from Arabidopsis, AtHSFA7b, in salt tolerance. AtHSFA7b is a nuclear protein with transactivation activity. ChIP-seq combined with an RNA-seq assay indicated that AtHSFA7b preferentially binds to a novel cis-acting element, termed the E-box-like motif, to regulate gene expression; it also binds to the heat shock element motif. Under salt conditions, AtHSFA7b regulates its target genes to mediate serial physiological changes, including maintaining cellular ion homeostasis, reducing water loss rate, decreasing reactive oxygen species accumulation, and adjusting osmotic potential, which ultimately leads to improved salt tolerance. Additionally, most cellulose synthase-like (CSL) and cellulose synthase (CESA) family genes were inhibited by AtHSFA7b; some of them were randomly selected for salt tolerance characterization, and they were mainly found to negatively modulate salt tolerance. By contrast, some transcription factors (TFs) were induced by AtHSFA7b; among them, we randomly identified six TFs that positively regulate salt tolerance. Thus, AtHSFA7b serves as a transactivator that positively mediates salinity tolerance mainly through binding to the E-box-like motif to regulate gene expression.

scholarly journals Characterisation of Listeria monocytogenes food-associated isolates to assess environmental fitness and virulence potential

2021 ◽  
pp. 109247
Author(s):  
Jessica A. Gray ◽  
P. Scott Chandry ◽  
Mandeep Kaur ◽  
Chawalit Kocharunchitt ◽  
John P. Bowman ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Virulence Potential ◽  
Environmental Fitness

scholarly journals Detection and Potential Virulence of Viable but Non-Culturable (VBNC) Listeria monocytogenes: A Review

2021 ◽  
Vol 9 (1) ◽  
pp. 194
Author(s):  
Nathan E. Wideman ◽  
James D. Oliver ◽  
Philip Glen Crandall ◽  
Nathan A. Jarvis
Keyword(s):  
Public Health ◽  
Listeria Monocytogenes ◽  
Viable Count ◽  
Vbnc State ◽  
Testing Methods ◽  
Tetrazolium Chloride ◽  
Definitive Evidence ◽  
Virulence Potential ◽  
Vbnc Bacteria ◽  
The Impact

The detection, enumeration, and virulence potential of viable but non-culturable (VBNC) pathogens continues to be a topic of discussion. While there is a lack of definitive evidence that VBNC Listeria monocytogenes (Lm) pose a public health risk, recent studies suggest that Lm in its VBNC state remains virulent. VBNC bacteria cannot be enumerated by traditional plating methods, so the results from routine Lm testing may not demonstrate a sample’s true hazard to public health. We suggest that supplementing routine Lm testing methods with methods designed to enumerate VBNC cells may more accurately represent the true level of risk. This review summarizes five methods for enumerating VNBC Lm: Live/Dead BacLightTM staining, ethidium monoazide and propidium monoazide-stained real-time polymerase chain reaction (EMA- and PMA-PCR), direct viable count (DVC), 5-cyano-2,3-ditolyl tetrazolium chloride-4′,6-diamidino-2-phenylindole (CTC-DAPI) double staining, and carboxy-fluorescein diacetate (CDFA) staining. Of these five supplementary methods, the Live/Dead BacLightTM staining and CFDA-DVC staining currently appear to be the most accurate for VBNC Lm enumeration. In addition, the impact of the VBNC state on the virulence of Lm is reviewed. Widespread use of these supplemental methods would provide supporting data to identify the conditions under which Lm can revert from its VBNC state into an actively multiplying state and help identify the environmental triggers that can cause Lm to become virulent. Highlights: Rationale for testing for all viable Listeria (Lm) is presented. Routine environmental sampling and plating methods may miss viable Lm cells. An overview and comparison of available VBNC testing methods is given. There is a need for resuscitation techniques to recover Lm from VBNC. A review of testing results for post VBNC virulence is compared

scholarly journals Transcriptional Analysis of the Adeno-Associated Virus Integration Site

2009 ◽  
Vol 83 (23) ◽  
pp. 12512-12525 ◽  
Author(s):  
Nathalie Dutheil ◽  
Els Henckaerts ◽  
Erik Kohlbrenner ◽  
R. Michael Linden
Keyword(s):  
Transcriptional Activity ◽  
Integration Site ◽  
Regulatory Elements ◽  
Transcriptional Analysis ◽  
Start Codon ◽  
Adeno Associated Virus ◽  
Site Specific ◽  
Complex Interactions ◽  
Site Specific Integration ◽  
Virus Integration

ABSTRACT The nonpathogenic human adeno-associated virus type 2 (AAV-2) has adopted a unique mechanism to site-specifically integrate its genome into the human MBS85 gene, which is embedded in AAVS1 on chromosome 19. The fact that AAV has evolved to integrate into this ubiquitously transcribed region and that the chromosomal motifs required for integration are located a few nucleotides upstream of the translation initiation start codon of MBS85 suggests that the transcriptional activity of MBS85 might influence site-specific integration and thus might be involved in the evolution of this mechanism. In order to begin addressing this question, we initiated the characterization of the human MBS85 promoter region and compared its transcriptional activity to that of the AAV-2 p5 promoter. Our results clearly indicate that AAVS1 is defined by a complex transcriptional environment and that the MBS85 promoter shares key regulatory elements with the viral p5 promoter. Furthermore, we provide evidence for bidirectional MBS85 promoter activity and demonstrate that the minimal motifs required for AAV site-specific integration are present in the 5′ untranslated region of the gene and play a posttranscriptional role in the regulation of MBS85 expression. These findings should provide a framework to further elucidate the complex interactions between the virus and its cellular host in this unique pathway to latency.

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