scholarly journals TetR-Type Transcriptional Regulator VtpR Functions as a Global Regulator in Vibrio tubiashii

2009 ◽  
Vol 75 (24) ◽  
pp. 7602-7609 ◽  
Author(s):  
Hiroaki Hasegawa ◽  
Claudia C. Häse
Keyword(s):  
Virulence Factors ◽  
Elevated Temperatures ◽  
Extracellular Proteins ◽  
Protease Production ◽  
High Homology ◽  
Global Regulator ◽  
Extracellular Proteases ◽  
Negative Effects ◽  
Vibrio Tubiashii ◽  
Protein Functions

ABSTRACT Vibrio tubiashii, a causative agent of severe shellfish larval disease, produces multiple extracellular proteins, including a metalloprotease (VtpA), as potential virulence factors. We previously reported that VtpA is toxic for Pacific oyster (Crassostrea gigas) larvae. In this study, we show that extracellular protease production by V. tubiashii was much reduced by elevated salt concentrations, as well as by elevated temperatures. In addition, V. tubiashii produced dramatically less protease in minimal salts medium supplemented with glucose or sucrose as the sole carbon source than with succinate. We identified a protein that belongs to the TetR family of transcriptional regulators, VtpR, which showed high homology with V. cholerae HapR. We conclude that VtpR activates VtpA production based on the following: (i) a VtpR-deficient V. tubiashii mutant did not produce extracellular proteases, (ii) the mutant showed reduced expression of a vtpA-lacZ fusion, and (iii) VtpR activated vtpA-lacZ in a V. cholerae heterologous background. Moreover, we show that VtpR activated the expression of an additional metalloprotease gene (vtpB). The deduced VtpB sequence showed high homology with a metalloprotease, VhpA, from V. harveyi. Furthermore, the vtpR mutant strain produced reduced levels of extracellular hemolysin, which is attributed to the lower expression of the V. tubiashii hemolysin genes (vthAB). The VtpR-deficient mutant also had negative effects on bacterial motility and did not demonstrate toxicity to oyster larvae. Together, these findings establish that the V. tubiashii VtpR protein functions as a global regulator controlling an array of potential virulence factors.

scholarly journals The Increased Accumulation of Staphylococcus aureus Virulence Factors Is Maximized in a purR Mutant by the Increased Production of SarA and Decreased Production of Extracellular Proteases

2021 ◽  
Vol 89 (4) ◽  
Author(s):  
Duah Alkam ◽  
Piroon Jenjaroenpun ◽  
Aura M. Ramirez ◽  
Karen E. Beenken ◽  
Horace J. Spencer ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Binding Proteins ◽  
Protease Production ◽  
Alpha Toxin ◽  
Extracellular Proteases ◽  
Content Type ◽  
Enhanced Production ◽  
Fibronectin Binding ◽  
The Impact

ABSTRACT Mutation of purR was previously shown to enhance the virulence of Staphylococcus aureus in a murine sepsis model, and this cannot be fully explained by increased expression of genes within the purine biosynthesis pathway. Rather, the increased production of specific S. aureus virulence factors, including alpha toxin and the fibronectin-binding proteins, was shown to play an important role. Mutation of purR was also shown previously to result in increased abundance of SarA. Here, we demonstrate by transposon sequencing that mutation of purR in the USA300 strain LAC increases fitness in a biofilm while mutation of sarA has the opposite effect. Therefore, we assessed the impact of sarA on reported purR-associated phenotypes by characterizing isogenic purR, sarA, and sarA/purR mutants. The results confirmed that mutation of purR results in increased abundance of alpha toxin, protein A, the fibronectin-binding proteins, and SarA, decreased production of extracellular proteases, an increased capacity to form a biofilm, and increased virulence in an osteomyelitis model. Mutation of sarA had the opposite effects on all of these phenotypes and, other than bacterial burdens in the bone, all of the phenotypes of sarA/purR mutants were comparable to those of sarA mutants. Limiting the production of extracellular proteases reversed all of the phenotypes of sarA mutants and most of those of sarA/purR mutants. We conclude that a critical component defining the virulence of a purR mutant is the enhanced production of SarA, which limits protease production to an extent that promotes the accumulation of critical S. aureus virulence factors.

scholarly journals Phosphorylation of MgrA and Its Effect on Expression of the NorA and NorB Efflux Pumps of Staphylococcus aureus

2010 ◽  
Vol 192 (10) ◽  
pp. 2525-2534 ◽  
Author(s):  
Que Chi Truong-Bolduc ◽  
David C. Hooper
Keyword(s):  
Staphylococcus Aureus ◽  
Multidrug Resistance ◽  
Virulence Factors ◽  
Double Mutant ◽  
Efflux Pumps ◽  
Efflux Transporters ◽  
Global Regulator ◽  
Binding Assays ◽  
Genes Encoding ◽  
Gel Shift

ABSTRACT MgrA is a global regulator in Staphylococcus aureus that controls the expression of diverse genes encoding virulence factors and multidrug resistance (MDR) efflux transporters. We identified pknB, which encodes the (Ser/Thr) kinase PknB, in the S. aureus genome. PknB was able to autophosphorylate as well as phosphorylate purified MgrA. We demonstrated that rsbU, which encodes a Ser/Thr phosphatase and is involved in the activation of the SigB regulon, was able to dephosphorylate MgrA-P but not PknB-P. Serines 110 and 113 of MgrA were found to be phosphorylated, and Ala substitutions at these positions resulted in reductions in the level of phosphorylation of MgrA. DNA gel shift binding assays using norA and norB promoters showed that MgrA-P was able to bind the norB promoter but not the norA promoter, a pattern which was the reverse of that for unphosphorylated MgrA. The double mutant MgrAS110A-S113A bound to the norA promoter but not the norB promoter. The double mutant led to a 2-fold decrease in norA transcripts and a 2-fold decrease in the MICs of norfloxacin and ciprofloxacin in strain RN6390. Thus, phosphorylation of MgrA results in loss of binding to the norA promoter, but with a gain of the ability to bind the norB promoter. Loss of the ability to phosphorylate MgrA by Ala substitution resulted in increased repression of norA expression and in reductions in susceptibilities to NorA substrates.

scholarly journals Nutritional Regulation of the Sae Two-Component System by CodY inStaphylococcus aureus

2018 ◽  
Vol 200 (8) ◽  
Author(s):  
Kevin D. Mlynek ◽  
William E. Sause ◽  
Derek E. Moormeier ◽  
Marat R. Sadykov ◽  
Kurt R. Hill ◽  
...  
Keyword(s):  
Gene Expression ◽  
Virulence Factors ◽  
Virulence Gene ◽  
Nutrient Depletion ◽  
Global Regulator ◽  
Two Component System ◽  
Component System ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Two Component

ABSTRACTStaphylococcus aureussubverts innate defenses during infection in part by killing host immune cells to exacerbate disease. This human pathogen intercepts host cues and activates a transcriptional response via theS. aureusexoprotein expression (SaeR/SaeS [SaeR/S]) two-component system to secrete virulence factors critical for pathogenesis. We recently showed that the transcriptional repressor CodY adjusts nuclease (nuc) gene expression via SaeR/S, but the mechanism remained unknown. Here, we identified two CodY binding motifs upstream of thesaeP1 promoter, which suggested direct regulation by this global regulator. We show that CodY shares a binding site with the positive activator SaeR and that alleviating direct CodY repression at this site is sufficient to abrogate stochastic expression, suggesting that CodY repressessaeexpression by blocking SaeR binding. Epistasis experiments support a model that CodY also controlssaeindirectly through Agr and Rot-mediated repression of thesaeP1 promoter. We also demonstrate that CodY repression ofsaerestrains production of secreted cytotoxins that kill human neutrophils. We conclude that CodY plays a previously unrecognized role in controlling virulence gene expression via SaeR/S and suggest a mechanism by which CodY acts as a master regulator of pathogenesis by tying nutrient availability to virulence gene expression.IMPORTANCEBacterial mechanisms that mediate the switch from a commensal to pathogenic lifestyle are among the biggest unanswered questions in infectious disease research. Since the expression of most virulence genes is often correlated with nutrient depletion, this implies that virulence is a response to the lack of nourishment in host tissues and that pathogens likeS. aureusproduce virulence factors in order to gain access to nutrients in the host. Here, we show that specific nutrient depletion signals appear to be funneled to the SaeR/S system through the global regulator CodY. Our findings reveal a strategy by whichS. aureusdelays the production of immune evasion and immune-cell-killing proteins until key nutrients are depleted.

Extracellular Heat-Resistant Proteases of Psychrotrophic Pseudomonads

1983 ◽  
Vol 46 (2) ◽  
pp. 90-94 ◽  
Author(s):  
THAKOR R. PATEL ◽  
FRANCIS M. BARTLETT ◽  
JAWED HAMID
Keyword(s):  
Protease Activity ◽  
Milk Powder ◽  
Milk Proteins ◽  
Raw Milk ◽  
Enzyme Synthesis ◽  
Maximum Activity ◽  
Protease Production ◽  
Divalent Metal Ions ◽  
Extracellular Proteases ◽  
Heat Resistant

Several bacterial isolates from raw milk produced proteases. Most of such 28 isolates were gram-negative rods which were oxidase- and catalase-positive. All the isolates grew at temperatures in the range of 0–35°C, but failed to grow at 37°C. Nineteen of these isolates were tentatively assigned to genus Pseudomonas, and were used in the present investigation. Extracellular proteases from these psychrotrophic pseudomonads were heat-resistant, being able to retain partial activity even after heat-treatment at 120°C for 10 min. Milk proteins were preferred substrates by these proteases although some also hydrolysed bovine serum albumin, hemoglobin and ovalbumin. The optimum pH for the maximum activity was between pH 7.2 and 7.4. Divalent metal ions like Cu2+, Co2+, Hg2+, and Zn2+ were inhibitory to protease activity while Ca2+, Mg2+, and Mn2+ had little or no inhibitory effect on the proteases. Induced levels of protease production were observed when cultures were grown in minimal media containing either casein or nonfat dried milk powder. Glucose, citrate and lactose repressed enzyme synthesis in a minmal salts medium containing either casein or nonfat dried milk powder. Protease activity was also detected in cultures grown in minimal medium containing glutamine. Proteases from different isolates varied in their molecular weights.

scholarly journals The Extracellular Metalloprotease of Vibrio tubiashii Is a Major Virulence Factor for Pacific Oyster (Crassostrea gigas) Larvae

2008 ◽  
Vol 74 (13) ◽  
pp. 4101-4110 ◽  
Author(s):  
Hiroaki Hasegawa ◽  
Erin J. Lind ◽  
Markus A. Boin ◽  
Claudia C. Häse
Keyword(s):  
Crassostrea Gigas ◽  
Pacific Oyster ◽  
Vibrio Vulnificus ◽  
Invasive Disease ◽  
High Homology ◽  
Structural Genes ◽  
Vibrio Tubiashii ◽  
Extracellular Products ◽  
Larval Toxicity ◽  
Culture Supernatants

ABSTRACT Vibrio tubiashii is a recently reemerging pathogen of larval bivalve mollusks, causing both toxigenic and invasive disease. Marine Vibrio spp. produce an array of extracellular products as potential pathogenicity factors. Culture supernatants of V. tubiashii have been shown to be toxic to oyster larvae and were reported to contain a metalloprotease and a cytolysin/hemolysin. However, the structural genes responsible for these proteins have yet to be identified, and it is uncertain which extracellular products play a role in pathogenicity. We investigated the effects of the metalloprotease and hemolysin secreted by V. tubiashii on its ability to kill Pacific oyster (Crassostrea gigas) larvae. While V. tubiashii supernatants treated with metalloprotease inhibitors severely reduced the toxicity to oyster larvae, inhibition of the hemolytic activity did not affect larval toxicity. We identified structural genes of V. tubiashii encoding a metalloprotease (vtpA) and a hemolysin (vthA). Sequence analyses revealed that VtpA shared high homology with metalloproteases from a variety of Vibrio species, while VthA showed high homology only to the cytolysin/hemolysin of Vibrio vulnificus. Compared to the wild-type strain, a VtpA mutant of V. tubiashii not only produced reduced amounts of protease but also showed decreased toxicity to C. gigas larvae. Vibrio cholerae strains carrying the vtpA or vthA gene successfully secreted the heterologous protein. Culture supernatants of V. cholerae carrying vtpA but not vthA were highly toxic to Pacific oyster larvae. Together, these results suggest that the V. tubiashii extracellular metalloprotease is important in its pathogenicity to C. gigas larvae.

scholarly journals Transcriptional Profiling of Target of RNAIII-Activating Protein, a Master Regulator of Staphylococcal Virulence

2005 ◽  
Vol 73 (10) ◽  
pp. 6220-6228 ◽  
Author(s):  
Moshe Korem ◽  
Yael Gov ◽  
Madanahally D. Kiran ◽  
Naomi Balaban
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Transcriptional Profiling ◽  
Protein A ◽  
Cell Communication ◽  
Global Regulator ◽  
Positive Bacterium ◽  
Master Regulator ◽  
Expression Of Genes ◽  
Histidine Phosphorylation

ABSTRACT Staphylococcus aureus is a gram-positive bacterium that is part of the normal healthy flora but that can become virulent and cause infections by producing biofilms and toxins. The production of virulence factors is regulated by cell-cell communication (quorum sensing) through the histidine phosphorylation of target of RNAIII-activating protein (TRAP), which is a 21-kDa protein that is highly conserved among staphylococci. Using microarray analysis, we show here that the expression and phosphorylation of TRAP upregulate the expression of most, if not all, toxins known to date, as well as their global regulator agr. In addition, we show here that the expression and phosphorylation of TRAP are also necessary for the expression of genes known to be necessary for the survival of the bacteria in a biofilm, like arc, pyr, and ure. TRAP is thus demonstrated to be a master regulator of staphylococcal pathogenesis.

scholarly journals Elevated Temperature Enhances Virulence of Erwinia carotovora subsp. carotovora Strain EC153 to Plants and Stimulates Production of the Quorum Sensing Signal, N-Acyl Homoserine Lactone, and Extracellular Proteins

2005 ◽  
Vol 71 (8) ◽  
pp. 4655-4663 ◽  
Author(s):  
H. Hasegawa ◽  
A. Chatterjee ◽  
Y. Cui ◽  
A. K. Chatterjee
Keyword(s):  
Quorum Sensing ◽  
Extracellular Enzymes ◽  
Rna Binding ◽  
Elevated Temperatures ◽  
Pectate Lyase ◽  
Erwinia Carotovora ◽  
Homoserine Lactone ◽  
Extracellular Proteins ◽  
Acyl Homoserine Lactone ◽  
Tissue Maceration

ABSTRACT Erwinia carotovora subsp. atroseptica, E. carotovora subsp. betavasculorum, and E. carotovora subsp. carotovora produce high levels of extracellular enzymes, such as pectate lyase (Pel), polygalacturonase (Peh), cellulase (Cel), and protease (Prt), and the quorum-sensing signal N-acyl-homoserine lactone (AHL) at 28°C. However, the production of these enzymes and AHL by these bacteria is severely inhibited during growth at elevated temperatures (31.2°C for E. carotovora subsp. atroseptica and 34.5°C for E. carotovora subsp. betavasculorum and most E. carotovora subsp. carotovora strains). At elevated temperatures these bacteria produce high levels of RsmA, an RNA binding protein that promotes RNA decay. E. carotovora subsp. carotovora strain EC153 is an exception in that it produces higher levels of Pel, Peh, Cel, and Prt at 34.5°C than at 28°C. EC153 also causes extensive maceration of celery petioles and Chinese cabbage leaves at 34.5°C, which correlates with a higher growth rate and higher levels of rRNA and AHL. The lack of pectinase production by E. carotovora subsp. carotovora strain Ecc71 at 34.5°C limits the growth of this organism in plant tissues and consequently impairs its ability to cause tissue maceration. Comparative studies with ahlI (the gene encoding a putative AHL synthase), pel-1, and peh-1 transcripts documented that at 34.5°C the RNAs are more stable in EC153 than in Ecc71. Our data reveal that overall metabolic activity, AHL levels, and mRNA stability are responsible for the higher levels of extracellular protein production and the enhanced virulence of EC153 at 34.5°C compared to 28°C.

A Review of the Implications of Silica in Solid Oxide Fuel Cells

2011 ◽  
Vol 8 (5) ◽  
Author(s):  
Michael Lankin ◽  
Yanhai Du ◽  
Caine Finnerty
Keyword(s):  
Fuel Cell ◽  
Solid Oxide Fuel Cells ◽  
Solid Oxide Fuel Cell ◽  
Elevated Temperatures ◽  
Raw Materials ◽  
Solid Oxide ◽  
Oxide Fuel ◽  
Negative Effects ◽  
Long Term Stability

Silica is a well-known impurity in solid oxide fuel cell raw materials, namely NiO and yttria-stabilized zirconia (YSZ). At elevated temperatures silica will migrate to the grain boundaries, form insulating siliceous phases, and lead to a decrease in the ionic conductivity of the electrolyte. Furthermore, silica impurities have been shown to damage the anode/electrolyte interface, such that an overall decrease in cell performance and long-term stability is observed. Despite the fact that silica is ubiquitous in commercial-grade raw materials and can be incorporated from several extrinsic sources, it has negative effects on the solid oxide fuel cell, such that any further contamination should be avoided to prevent performance degradation and eventual cell failure. This paper reviews and outlines the sources and effects of silica on the solid oxide fuel cell, and attempts to determine a guideline for acceptable levels of silica contamination.

scholarly journals Mapping the Global Network of Extracellular Protease Regulation in Staphylococcus aureus

2019 ◽  
Author(s):  
Brittney D. Gimza ◽  
Maria I. Larias ◽  
Bridget G. Budny ◽  
Lindsey N. Shaw
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Global Network ◽  
Protease Production ◽  
Secondary Circuit ◽  
Pathogenic Potential ◽  
Regulatory Circuit ◽  
Primary Network ◽  
The Impact ◽  
Protease Expression

AbstractA primary function of the extracellular proteases of Staphylococcus aureus is to control the progression of infection by selectively modulating the stability of virulence factors. Consequently, a regulatory network exists to titrate protease abundance/activity, to influence accumulation, or lack thereof, of individual virulence factors. Herein, we comprehensively map this system, exploring regulation of the four protease loci by known and novel factors. In so doing, we determine that seven major elements (SarS, SarR, Rot, MgrA, CodY, SaeR, and SarA) form the primary network of control, with the latter three being the most powerful. We note that expression of aureolysin is largely repressed by these factors, whilst the spl operon is subject to the strongest upregulation of any protease loci, particularly by SarR and SaeR. Furthermore, when exploring scpA expression, we find it to be profoundly influenced in opposing fashions by SarA (repressor) and SarR (activator). We also present the screening of >100 regulator mutants of S. aureus, identifying 7 additional factors (ArgR2, AtlR, MntR, Rex, XdrA, Rbf, and SarU) that form a secondary circuit of protease control. Primarily these elements serve as activators, although we reveal XdrA as a new repressor of protease expression. With the exception or ArgR2, each of the new effectors appear to work through the primary network of regulation to influence protease production. Collectively, we present a comprehensive regulatory circuit that emphasizes the complexity of protease regulation and suggest that its existence speaks to the importance of these enzymes to S. aureus physiology and pathogenic potential.ImportanceThe complex regulatory role of the proteases necessitates very tight coordination and control of their expression. Whilst this process has been well studied, a major oversight has been the consideration of proteases as a single entity, rather than 10 enzymes produced from four different promoters. As such, in this study we comprehensively characterized the regulation of each protease promoter, discovering vast differences in the way each protease operon is controlled. Additionally, we broaden the picture of protease regulation using a global screen to identify novel loci controlling protease activity, uncovering a cadre of new effectors of protease expression. The impact of these elements on the activity of proteases and known regulators was characterized producing a comprehensive regulatory circuit that emphasizes the complexity of protease regulation in Staphylococcus aureus.

scholarly journals Ephemeral hypoxia reduces oxygen consumption in the Caribbean coral Orbicella faveolata

Coral Reefs ◽  
2021 ◽  
Author(s):  
Philip M. Gravinese ◽  
Alex Douwes ◽  
Katherine R. Eaton ◽  
Erinn M. Muller
Keyword(s):  
Oxygen Consumption ◽  
Elevated Temperature ◽  
Coastal Waters ◽  
Elevated Temperatures ◽  
Wet Season ◽  
Negative Effects ◽  
Hypoxic Conditions ◽  
Orbicella Faveolata ◽  
The Caribbean ◽  
Solubility Of Oxygen

AbstractOxygen concentrations in coastal waters have declined globally by 10% since the mid-twentieth century, and ocean warming will further reduce the solubility of oxygen in coastal habitats. Some nearshore reefs experience periodic hypoxic conditions due to eutrophication, especially during the wet season. Here, we determined the combined impacts of hypoxia and elevated temperature on the reef-building coral, Orbicella faveolata, by exposing corals to normoxic or hypoxic conditions and ambient or elevated temperatures. Oxygen consumption was monitored using closed-system respirometry. Corals within hypoxic conditions consumed 34% less oxygen relative to corals in normoxic conditions. Corals in the elevated temperature normoxic treatment experienced a 10% increase in oxygen consumption relative to the control. Corals exposed to both stressors simultaneously experienced a 62% reduction in oxygen consumption. These results suggest that increased temperature may exacerbate the negative effects of hypoxia on O. faveolata.

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