scholarly journals The LonA Protease Regulates Biofilm Formation, Motility, Virulence, and the Type VI Secretion System in Vibrio cholerae

2016 ◽  
Vol 198 (6) ◽  
pp. 973-985 ◽  
Author(s):  
Andrew Rogers ◽  
Loni Townsley ◽  
Ana L. Gallego-Hernandez ◽  
Sinem Beyhan ◽  
Laura Kwuan ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Biofilm Formation ◽  
Secretion System ◽  
Lon Protease ◽  
Human Pathogen ◽  
Type Vi Secretion System ◽  
Cyclic Diguanylate ◽  
Content Type ◽  
Infant Mouse ◽  
Type Vi Secretion

ABSTRACTThe presence of the Lon protease in all three domains of life hints at its biological importance. The prokaryotic Lon protease is responsible not only for degrading abnormal proteins but also for carrying out the proteolytic regulation of specific protein targets. Posttranslational regulation by Lon is known to affect a variety of physiological traits in many bacteria, including biofilm formation, motility, and virulence. Here, we identify the regulatory roles of LonA in the human pathogenVibrio cholerae. We determined that the absence of LonA adversely affects biofilm formation, increases swimming motility, and influences intracellular levels of cyclic diguanylate. Whole-genome expression analysis revealed that the message abundance of genes involved in biofilm formation was decreased but that the message abundances of those involved in virulence and the type VI secretion system were increased in alonAmutant compared to the wild type. We further demonstrated that alonAmutant displays an increase in type VI secretion system activity and is markedly defective in colonization of the infant mouse. These findings suggest that LonA plays a critical role in the environmental survival and virulence ofV. cholerae.IMPORTANCEBacteria utilize intracellular proteases to degrade damaged proteins and adapt to changing environments. The Lon protease has been shown to be important for environmental adaptation and plays a crucial role in regulating the motility, biofilm formation, and virulence of numerous plant and animal pathogens. We find that LonA of the human pathogenV. choleraeis in line with this trend, as the deletion of LonA leads to hypermotility and defects in both biofilm formation and colonization of the infant mouse. In addition, we show that LonA regulates levels of cyclic diguanylate and the type VI secretion system. Our observations add to the known regulatory repertoire of the Lon protease and the current understanding ofV. choleraephysiology.

scholarly journals Type VI Secretion System Dynamics Reveals a Novel Secretion Mechanism in Pseudomonas aeruginosa

2018 ◽  
Vol 200 (11) ◽  
Author(s):  
Jacqueline Corbitt ◽  
Jun Seok Yeo ◽  
C. Ian Davis ◽  
Michele LeRoux ◽  
Paul A. Wiggins
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Vibrio Cholerae ◽  
Conformational Change ◽  
Secretion System ◽  
Force Generation ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Evolutionary Diversification ◽  
Type Vi Secretion ◽  
Generation Mechanisms

ABSTRACT The type VI secretion system (T6SS) inhibits the growth of neighboring bacterial cells through a contact-mediated mechanism. Here, we describe a detailed characterization of the protein localization dynamics in the Pseudomonas aeruginosa T6SS. It has been proposed that the type VI secretion process is driven by a conformational-change-induced contraction of the T6SS sheath. However, although the contraction of an optically resolvable TssBC sheath and the subsequent localization of ClpV are observed in Vibrio cholerae , coordinated assembly and disassembly of TssB and ClpV are observed without TssB contraction in P. aeruginosa . These dynamics are inconsistent with the proposed contraction sheath model. Motivated by the phenomenon of dynamic instability, we propose a new model in which ATP hydrolysis, rather than conformational change, generates the force for secretion. IMPORTANCE The type VI secretion system (T6SS) is widely conserved among Gram-negative bacteria and is a central determinant of bacterial fitness in polymicrobial communities. The secretion system targets bacteria and secretes effectors that inhibit the growth of neighboring cells, using a contact-mediated-delivery system. Despite significant homology to the previously characterized Vibrio cholerae T6SS, our analysis reveals that effector secretion is driven by a distinct force generation mechanism in Pseudomonas aeruginosa . The presence of two distinct force generation mechanisms in T6SS represents an example of the evolutionary diversification of force generation mechanisms.

scholarly journals Vibrio cholerae Requires the Type VI Secretion System Virulence Factor VasX To Kill Dictyostelium discoideum

2011 ◽  
Vol 79 (7) ◽  
pp. 2941-2949 ◽  
Author(s):  
Sarah T. Miyata ◽  
Maya Kitaoka ◽  
Teresa M. Brooks ◽  
Steven B. McAuley ◽  
Stefan Pukatzki
Keyword(s):  
Vibrio Cholerae ◽  
Dictyostelium Discoideum ◽  
Virulence Factor ◽  
Membrane Lipids ◽  
Gene Clusters ◽  
Secretion System ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Enzymatic Function ◽  
Type Vi Secretion

ABSTRACTThe type VI secretion system (T6SS) is recognized as an important virulence mechanism in several Gram-negative pathogens. InVibrio cholerae, the causative agent of the diarrheal disease cholera, a minimum of three gene clusters—one main cluster and two auxiliary clusters—are required to form a functional T6SS apparatus capable of conferring virulence toward eukaryotic and prokaryotic hosts. Despite an increasing understanding of the components that make up the T6SS apparatus, little is known about the regulation of these genes and the gene products delivered by this nanomachine. VasH is an important regulator of theV. choleraeT6SS. Here, we present evidence that VasH regulates the production of a newly identified protein, VasX, which in turn requires a functional T6SS for secretion. Deletion ofvasXdoes not affect export or enzymatic function of the structural T6SS proteins Hcp and VgrG-1, suggesting that VasX is dispensable for the assembly of the physical translocon complex. VasX localizes to the bacterial membrane and interacts with membrane lipids. We present VasX as a novel virulence factor of the T6SS, as aV. choleraemutant lackingvasXexhibits a phenotype of attenuated virulence towardDictyostelium discoideum.

scholarly journals Screening for Inhibition of Vibrio cholerae VipA-VipB Interaction Identifies Small-Molecule Compounds Active against Type VI Secretion

2014 ◽  
Vol 58 (7) ◽  
pp. 4123-4130 ◽  
Author(s):  
Kun Sun ◽  
Jeanette Bröms ◽  
Moa Lavander ◽  
Bharat Kumar Gurram ◽  
Per-Anders Enquist ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Small Molecule ◽  
Secretion System ◽  
Eukaryotic Cells ◽  
Minimal Risk ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Development Of Resistance ◽  
Type Vi Secretion ◽  
Bacterial Replication

ABSTRACTThe type VI secretion system (T6SS) is the most prevalent bacterial secretion system and an important virulence mechanism utilized by Gram-negative bacteria, either to target eukaryotic cells or to combat other microbes. The components show much variability, but some appear essential for the function, and two homologues, denoted VipA and VipB inVibrio cholerae, have been identified in all T6SSs described so far. Secretion is dependent on binding of an α-helical region of VipA to VipB, and in the absence of this binding, both components are degraded within minutes and secretion is ceased. The aim of the study was to investigate if this interaction could be blocked, and we hypothesized that such inhibition would lead to abrogation of T6S. A library of 9,600 small-molecule compounds was screened for their ability to block the binding of VipA-VipB in a bacterial two-hybrid system (B2H). After excluding compounds that showed cytotoxicity toward eukaryotic cells, that inhibited growth ofVibrio, or that inhibited an unrelated B2H interaction, 34 compounds were further investigated for effects on the T6SS-dependent secretion of hemolysin-coregulated protein (Hcp) or of phospholipase A1activity. Two compounds, KS100 and KS200, showed intermediate or strong effects in both assays. Analogues were obtained, and compounds with potent inhibitory effects in the assays and desirable physicochemical properties as predicted byin silicoanalysis were identified. Since the compounds specifically target a virulence mechanism without affecting bacterial replication, they have the potential to mitigate the virulence with minimal risk for development of resistance.

scholarly journals Light Modulates Metabolic Pathways and Other Novel Physiological Traits in the Human Pathogen Acinetobacter baumannii

2017 ◽  
Vol 199 (10) ◽  
Author(s):  
Gabriela L. Müller ◽  
Marisel Tuttobene ◽  
Matías Altilio ◽  
Maitena Martínez Amezaga ◽  
Meaghan Nguyen ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Metabolic Pathways ◽  
Secretion System ◽  
Human Pathogen ◽  
Type Vi Secretion System ◽  
Bacterial Persistence ◽  
Content Type ◽  
Light Sensing ◽  
Type Vi Secretion ◽  
Novel Concept

ABSTRACT Light sensing in chemotrophic bacteria has been relatively recently ascertained. In the human pathogen Acinetobacter baumannii, light modulates motility, biofilm formation, and virulence through the blue-light-sensing-using flavin (BLUF) photoreceptor BlsA. In addition, light can induce a reduction in susceptibility to certain antibiotics, such as minocycline and tigecycline, in a photoreceptor-independent manner. In this work, we identified new traits whose expression levels are modulated by light in this pathogen, which comprise not only important determinants related to pathogenicity and antibiotic resistance but also metabolic pathways, which represents a novel concept for chemotrophic bacteria. Indeed, the phenylacetic acid catabolic pathway and trehalose biosynthesis were modulated by light, responses that completely depend on BlsA. We further show that tolerance to some antibiotics and modulation of antioxidant enzyme levels are also influenced by light, likely contributing to bacterial persistence in adverse environments. Also, we present evidence indicating that surfactant production is modulated by light. Finally, the expression of whole pathways and gene clusters, such as genes involved in lipid metabolism and genes encoding components of the type VI secretion system, as well as efflux pumps related to antibiotic resistance, was differentially induced by light. Overall, our results indicate that light modulates global features of the A. baumannii lifestyle. IMPORTANCE The discovery that nonphototrophic bacteria respond to light constituted a novel concept in microbiology. In this context, we demonstrated that light could modulate aspects related to bacterial virulence, persistence, and resistance to antibiotics in the human pathogen Acinetobacter baumannii. In this work, we present the novel finding that light directly regulates metabolism in this chemotrophic bacterium. Insights into the mechanism show the involvement of the photoreceptor BlsA. In addition, tolerance to antibiotics and catalase levels are also influenced by light, likely contributing to bacterial persistence in adverse environments, as is the expression of the type VI secretion system and efflux pumps. Overall, a profound influence of light on the lifestyle of A. baumannii is suggested to occur.

scholarly journals Pathoadaptive Conditional Regulation of the Type VI Secretion System in Vibrio cholerae O1 Strains

2011 ◽  
Vol 80 (2) ◽  
pp. 575-584 ◽  
Author(s):  
Takahiko Ishikawa ◽  
Dharmesh Sabharwal ◽  
Jeanette Bröms ◽  
Debra L. Milton ◽  
Anders Sjöstedt ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Secretion System ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Content Type ◽  
High Osmolarity ◽  
Secretion Pathway ◽  
Type Vi Secretion ◽  
Vibrio Cholerae O1 ◽  
K 12

ABSTRACTThe most recently discovered secretion pathway in Gram-negative bacteria, the type VI secretion system (T6SS), is present in many species and is considered important for the survival of non-O1 non-O139Vibrio choleraein aquatic environments. Until now, it was not known whether there is a functionally active T6SS in wild-typeV. choleraeO1 strains, the cause of cholera disease in humans. Here, we demonstrate the presence of a functionally active T6SS in wild-typeV. choleraeO1 strains, as evidenced by the secretion of the T6SS substrate Hcp, which required several gene products encoded within the putativevasgene cluster. Our analyses showed that the T6SS of wild-typeV. choleraeO1 strain A1552 was functionally activated when the bacteria were grown under high-osmolarity conditions. The T6SS was also active when the bacteria were grown under low temperature (23°C), suggesting that the system may be important for the survival of the bacterium in the environment. A test of the interbacterial virulence ofV. choleraestrain A1552 against anEscherichia coliK-12 strain showed that it was strongly enhanced under high osmolarity and that it depended on thehcpgenes. Interestingly, we found that the newly recognized osmoregulatory protein OscR plays a role in the regulation of T6SS gene expression and secretion of Hcp fromV. choleraeO1 strains.

scholarly journals Diversity of Clinical and Environmental Isolates of Vibrio cholerae in Natural Transformation and Contact-Dependent Bacterial Killing Indicative of Type VI Secretion System Activity

2016 ◽  
Vol 82 (9) ◽  
pp. 2833-2842 ◽  
Author(s):  
Eryn E. Bernardy ◽  
Maryann A. Turnsek ◽  
Sarah K. Wilson ◽  
Cheryl L. Tarr ◽  
Brian K. Hammer
Keyword(s):  
Vibrio Cholerae ◽  
Natural Transformation ◽  
Secretion System ◽  
Past Century ◽  
Dna Uptake ◽  
Type Vi Secretion System ◽  
Environmental Isolates ◽  
Bacterial Killing ◽  
Content Type ◽  
Type Vi Secretion

ABSTRACTThe bacterial pathogenVibrio choleraecan occupy both the human gut and aquatic reservoirs, where it may colonize chitinous surfaces that induce the expression of factors for three phenotypes: chitin utilization, DNA uptake by natural transformation, and contact-dependent bacterial killing via a type VI secretion system (T6SS). In this study, we surveyed a diverse set of 53 isolates from different geographic locales collected over the past century from human clinical and environmental specimens for each phenotype outlined above. The set included pandemic isolates of serogroup O1, as well as several serogroup O139 and non-O1/non-O139 strains. We found that while chitin utilization was common, only 22.6% of the isolates tested were proficient at chitin-induced natural transformation, suggesting that transformation is expendable. Constitutive contact-dependent killing ofEscherichia coliprey, which is indicative of a functional T6SS, was rare among clinical isolates (only 4 of 29) but common among environmental isolates (22 of 24). These results bolster the pathoadaptive model in which tight regulation of T6SS-mediated bacterial killing is beneficial in a human host, whereas constitutive killing by environmental isolates may give a competitive advantage in natural settings. Future sequence analysis of this set of diverse isolates may identify previously unknown regulators and structural components for both natural transformation and T6SS.

scholarly journals Transcriptional Silencing by TsrA in the Evolution of Pathogenic Vibrio cholerae Biotypes

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Florence Caro ◽  
José A. Caro ◽  
Nicole M. Place ◽  
John J. Mekalanos
Keyword(s):  
Gene Expression ◽  
Vibrio Cholerae ◽  
Virulence Factors ◽  
Virulence Genes ◽  
Secretion System ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Genes Encoding ◽  
El Tor

ABSTRACT Vibrio cholerae is a globally important pathogen responsible for the severe epidemic diarrheal disease called cholera. The current and ongoing seventh pandemic of cholera is caused by El Tor strains, which have completely replaced the sixth-pandemic classical strains of V. cholerae. To successfully establish infection and disseminate to new victims, V. cholerae relies on key virulence factors encoded on horizontally acquired genetic elements. The expression of these factors relies on the regulatory architecture that coordinates the timely expression of virulence determinants during host infection. Here, we apply transcriptomics and structural modeling to understand how type VI secretion system regulator A (TsrA) affects gene expression in both the classical and El Tor biotypes of V. cholerae. We find that TsrA acts as a negative regulator of V. cholerae virulence genes encoded on horizontally acquired genetic elements. The TsrA regulon comprises genes encoding cholera toxin (CT), the toxin-coregulated pilus (TCP), and the type VI secretion system (T6SS), as well as genes involved in biofilm formation. The majority of the TsrA regulon is carried on horizontally acquired AT-rich genetic islands whose loss or acquisition could be directly ascribed to the differences between the classical and El Tor strains studied. Our modeling predicts that the TsrA protein is a structural homolog of the histone-like nucleoid structuring protein (H-NS) oligomerization domain and is likely capable of forming higher-order superhelical structures, potentially with DNA. These findings describe how TsrA can integrate into the intricate V. cholerae virulence gene expression program, controlling gene expression through transcriptional silencing. IMPORTANCE Pathogenic Vibrio cholerae strains express multiple virulence factors that are encoded by bacteriophage and chromosomal islands. These include cholera toxin and the intestinal colonization pilus called the toxin-coregulated pilus, which are essential for causing severe disease in humans. However, it is presently unclear how the expression of these horizontally acquired accessory virulence genes can be efficiently integrated with preexisting transcriptional programs that are presumably fine-tuned for optimal expression in V. cholerae before its conversion to a human pathogen. Here, we report the role of a transcriptional regulator (TsrA) in silencing horizontally acquired genes encoding important virulence factors. We propose that this factor could be critical to the efficient acquisition of accessory virulence genes by silencing their expression until other signals trigger their transcriptional activation within the host.

scholarly journals Pandemic Vibrio cholerae Acquired Competitive Traits from an Environmental Vibrio Species

2021 ◽  
Author(s):  
Francis J Santoriello ◽  
Paul Kirchberger ◽  
Yann Boucher ◽  
Stefan Pukatzki
Keyword(s):  
Gene Transfer ◽  
Vibrio Cholerae ◽  
Secretion System ◽  
Human Pathogen ◽  
Environmental Niche ◽  
Fish Pathogen ◽  
Type Vi Secretion System ◽  
Type Vi Secretion ◽  
Potential Reservoir ◽  
Environmental Reservoir

Background: Vibrio cholerae, the causative agent of cholera, is a human pathogen that thrives in estuarine environments. V. cholerae competes with neighboring microbes by the contact-dependent translocation of toxic effectors with the type VI secretion system (T6SS). Effector types are highly variable across V. cholerae strains, but all pandemic isolates encode the same set of distinct effectors. It is possible that acquisition of these effectors via horizontal gene transfer played a role in the development of pandemic V. cholerae. Results: We assessed the distribution of V. cholerae T6SS loci across multiple Vibrio species. We showed that the fish-pathogen V. anguillarum encodes all three V. cholerae core loci as well as two of the four additional auxiliary clusters. We further demonstrated that V. anguillarum shares T6SS effectors with V. cholerae, including every pandemic-associated V. cholerae effector. We identified a novel T6SS cluster (Accessory Aux1) that is widespread in V. anguillarum and encodes the pandemic V. cholerae effector TseL. We highlighted potential gene transfer events of Accessory Aux1 from V. anguillarum to V. cholerae. Finally, we showed that TseL from V. cholerae can be neutralized by the V. anguillarum Accessory Aux1 immunity protein and vice versa, indicating V. anguillarum as the donor of tseL to the V. cholerae species. Conclusions: V. anguillarum constitutes an environmental reservoir of pandemic-associated V. cholerae T6SS effectors. V. anguillarum and V. cholerae likely share an environmental niche, compete, and exchange T6SS effectors. Further, our findings highlight the fish as a potential reservoir of pandemic V. cholerae.

scholarly journals Secretome Analysis of Vibrio cholerae Type VI Secretion System Reveals a New Effector-Immunity Pair

mBio ◽  
2015 ◽  
Vol 6 (2) ◽  
Author(s):  
Emrah Altindis ◽  
Tao Dong ◽  
Christy Catalano ◽  
John Mekalanos
Keyword(s):  
Vibrio Cholerae ◽  
Bioinformatics Analysis ◽  
Secretion System ◽  
Comparative Proteomics ◽  
Gram Negative Bacteria ◽  
Type Vi Secretion System ◽  
Immunity Protein ◽  
Gram Negative ◽  
Content Type ◽  
Type Vi Secretion

ABSTRACT The type VI secretion system (T6SS) is a dynamic macromolecular organelle that many Gram-negative bacteria use to inhibit or kill other prokaryotic or eukaryotic cells. The toxic effectors of T6SS are delivered to the prey cells in a contact-dependent manner. In Vibrio cholerae, the etiologic agent of cholera, T6SS is active during intestinal infection. Here, we describe the use of comparative proteomics coupled with bioinformatics to identify a new T6SS effector-immunity pair. This analysis was able to identify all previously identified secreted substrates of T6SS except PAAR (proline, alanine, alanine, arginine) motif-containing proteins. Additionally, this approach led to the identification of a new secreted protein encoded by VCA0285 (TseH) that carries a predicted hydrolase domain. We confirmed that TseH is toxic when expressed in the periplasm of Escherichia coli and V. cholerae cells. The toxicity observed in V. cholerae was suppressed by coexpression of the protein encoded by VCA0286 (TsiH), indicating that this protein is the cognate immunity protein of TseH. Furthermore, exogenous addition of purified recombinant TseH to permeabilized E. coli cells caused cell lysis. Bioinformatics analysis of the TseH protein sequence suggest that it is a member of a new family of cell wall-degrading enzymes that include proteins belonging to the YD repeat and Rhs superfamilies and that orthologs of TseH are likely expressed by species belonging to phyla as diverse as Bacteroidetes and Proteobacteria. IMPORTANCE The Gram-negative bacterium Vibrio cholerae causes cholera, a severe and often lethal diarrheal disease. The 2010-2012 epidemic in Haiti and new explosive epidemics in Africa show that cholera remains a significant global public health problem. The type VI secretion system (T6SS) is a dynamic organelle expressed by many Gram-negative bacteria, which use it to inject toxic effector proteins into eukaryotic and bacterial prey cells. In this study, we applied a comparative proteomics approach to the V. cholerae T6SS secretome to identify new substrates of this secretion apparatus. We show that the product of the gene VCA0285 is likely a new peptidoglycan hydrolase that is secreted by T6SS and that its cognate immunity protein is encoded by the gene that is immediately downstream (VCA0286). Bioinformatics analysis shows that VCA0285 carries four conserved motifs that likely define a large family of hydrolases with antibacterial activity. The identification of new antibacterial T6SS effectors provides useful information for the development of novel antibiotics and therapeutic agents.

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