scholarly journals Molecular Pathogenesis of Shigella spp.: Controlling Host Cell Signaling, Invasion, and Death by Type III Secretion

2008 ◽  
Vol 21 (1) ◽  
pp. 134-156 ◽  
Author(s):  
Gunnar N. Schroeder ◽  
Hubert Hilbi
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Molecular Mechanisms ◽  
Inflammatory Responses ◽  
Shigella Flexneri ◽  
Effector Proteins ◽  
Target Cells ◽  
Type Iii ◽  
Shigella Spp

SUMMARY Shigella spp. are gram-negative pathogenic bacteria that evolved from harmless enterobacterial relatives and may cause devastating diarrhea upon ingestion. Research performed over the last 25 years revealed that a type III secretion system (T3SS) encoded on a large plasmid is a key virulence factor of Shigella flexneri. The T3SS determines the interactions of S. flexneri with intestinal cells by consecutively translocating two sets of effector proteins into the target cells. Thus, S. flexneri controls invasion into EC, intra- and intercellular spread, macrophage cell death, as well as host inflammatory responses. Some of the translocated effector proteins show novel biochemical activities by which they intercept host cell signal transduction pathways. An understanding of the molecular mechanisms underlying Shigella pathogenesis will foster the development of a safe and efficient vaccine, which, in parallel with improved hygiene, should curb infections by this widespread pathogen.

scholarly journals Antibodies Inhibiting the Type III Secretion System of Gram-Negative Pathogenic Bacteria

Antibodies ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 35
Author(s):  
Julia A. Hotinger ◽  
Aaron E. May
Keyword(s):  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Type Iii Secretion System ◽  
The United States ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Gram Negative ◽  
Type Iii ◽  
Shigella Spp

Pathogenic bacteria are a global health threat, with over 2 million infections caused by Gram-negative bacteria every year in the United States. This problem is exacerbated by the increase in resistance to common antibiotics that are routinely used to treat these infections, creating an urgent need for innovative ways to treat and prevent virulence caused by these pathogens. Many Gram-negative pathogenic bacteria use a type III secretion system (T3SS) to inject toxins and other effector proteins directly into host cells. The T3SS has become a popular anti-virulence target because it is required for pathogenesis and knockouts have attenuated virulence. It is also not required for survival, which should result in less selective pressure for resistance formation against T3SS inhibitors. In this review, we will highlight selected examples of direct antibody immunizations and the use of antibodies in immunotherapy treatments that target the bacterial T3SS. These examples include antibodies targeting the T3SS of Pseudomonas aeruginosa, Yersinia pestis, Escherichia coli, Salmonella enterica, Shigella spp., and Chlamydia trachomatis.

scholarly journals Identification of Novel Host Interactors of Effectors Secreted by Salmonella and Citrobacter

mSystems ◽  
2016 ◽  
Vol 1 (4) ◽  
Author(s):  
Ryan L. Sontag ◽  
Ernesto S. Nakayasu ◽  
Roslyn N. Brown ◽  
George S. Niemann ◽  
Michael A. Sydor ◽  
...  
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Defense Mechanisms ◽  
Pathogenic Bacteria ◽  
Effector Proteins ◽  
Host Cells ◽  
Secretion Systems ◽  
Type Iii ◽  
Cell Cytosol ◽  
Host Cell Cytosol

ABSTRACT During infection, pathogenic bacteria face an adverse environment of factors driven by both cellular and humoral defense mechanisms. To help evade the immune response and ultimately proliferate inside the host, many bacteria evolved specialized secretion systems to deliver effector proteins directly into host cells. Translocated effector proteins function to subvert host defense mechanisms. Numerous pathogenic bacteria use a specialized secretion system called type III secretion to deliver effectors into the host cell cytosol. Here, we identified 75 new host targets of Salmonella and Citrobacter effectors, which will help elucidate their mechanisms of action. Many pathogenic bacteria of the family Enterobacteriaceae use type III secretion systems to inject virulence proteins, termed “effectors,” into the host cell cytosol. Although host-cellular activities of several effectors have been demonstrated, the function and host-targeted pathways of most of the effectors identified to date are largely undetermined. To gain insight into host proteins targeted by bacterial effectors, we performed coaffinity purification of host proteins from cell lysates using recombinant effectors from the Enterobacteriaceae intracellular pathogens Salmonella enterica serovar Typhimurium and Citrobacter rodentium. We identified 54 high-confidence host interactors for the Salmonella effectors GogA, GtgA, GtgE, SpvC, SrfH, SseL, SspH1, and SssB collectively and 21 interactors for the Citrobacter effectors EspT, NleA, NleG1, and NleK. We biochemically validated the interaction between the SrfH Salmonella protein and the extracellular signal-regulated kinase 2 (ERK2) host protein kinase, which revealed a role for this effector in regulating phosphorylation levels of this enzyme, which plays a central role in signal transduction. IMPORTANCE During infection, pathogenic bacteria face an adverse environment of factors driven by both cellular and humoral defense mechanisms. To help evade the immune response and ultimately proliferate inside the host, many bacteria evolved specialized secretion systems to deliver effector proteins directly into host cells. Translocated effector proteins function to subvert host defense mechanisms. Numerous pathogenic bacteria use a specialized secretion system called type III secretion to deliver effectors into the host cell cytosol. Here, we identified 75 new host targets of Salmonella and Citrobacter effectors, which will help elucidate their mechanisms of action.

scholarly journals CesL Regulates Type III Secretion Substrate Specificity of the Enteropathogenic E. coli Injectisome

2021 ◽  
Vol 9 (5) ◽  
pp. 1047
Author(s):  
Miguel Díaz-Guerrero ◽  
Meztlli O. Gaytán ◽  
Eduardo Soto ◽  
Norma Espinosa ◽  
Elizabeth García-Gómez ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Effector Proteins ◽  
Host Cells ◽  
Target Cells ◽  
Remarkable Feature ◽  
Type Iii ◽  
Pairwise Interactions ◽  
Effector Secretion

The type III secretion system (T3SS) is a complex molecular device used by several pathogenic bacteria to translocate effector proteins directly into eukaryotic host cells. One remarkable feature of the T3SS is its ability to secrete different categories of proteins in a hierarchical manner, to ensure proper assembly and timely delivery of effectors into target cells. In enteropathogenic Escherichia coli, the substrate specificity switch from translocator to effector secretion is regulated by a gatekeeper complex composed of SepL, SepD, and CesL proteins. Here, we report a characterization of the CesL protein using biochemical and genetic approaches. We investigated discrepancies in the phenotype among different cesL deletion mutants and showed that CesL is indeed essential for translocator secretion and to prevent premature effector secretion. We also demonstrated that CesL engages in pairwise interactions with both SepL and SepD. Furthermore, while association of SepL to the membrane does not depended on CesL, the absence of any of the proteins forming the heterotrimeric complex compromised the intracellular stability of each component. In addition, we found that CesL interacts with the cytoplasmic domains of the export gate components EscU and EscV. We propose a mechanism for substrate secretion regulation governed by the SepL/SepD/CesL complex.

scholarly journals The Shigella Type III Secretion System: An Overview from Top to Bottom

2021 ◽  
Vol 9 (2) ◽  
pp. 451
Author(s):  
Meenakumari Muthuramalingam ◽  
Sean K. Whittier ◽  
Wendy L. Picking ◽  
William D. Picking
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Shigella Flexneri ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Effector Functions ◽  
Type Iii ◽  
Host Cytoplasm ◽  
Host Cell Membranes

Shigella comprises four species of human-restricted pathogens causing bacillary dysentery. While Shigella possesses multiple genetic loci contributing to virulence, a type III secretion system (T3SS) is its primary virulence factor. The Shigella T3SS nanomachine consists of four major assemblies: the cytoplasmic sorting platform; the envelope-spanning core/basal body; an exposed needle; and a needle-associated tip complex with associated translocon that is inserted into host cell membranes. The initial subversion of host cell activities is carried out by the effector functions of the invasion plasmid antigen (Ipa) translocator proteins, with the cell ultimately being controlled by dedicated effector proteins that are injected into the host cytoplasm though the translocon. Much of the information now available on the T3SS injectisome has been accumulated through collective studies on the T3SS from three systems, those of Shigella flexneri, Salmonella typhimurium and Yersinia enterocolitica/Yersinia pestis. In this review, we will touch upon the important features of the T3SS injectisome that have come to light because of research in the Shigella and closely related systems. We will also briefly highlight some of the strategies being considered to target the Shigella T3SS for disease prevention.

scholarly journals Role of Autocleavage in the Function of a Type III Secretion Specificity Switch Protein in Salmonella enterica Serovar Typhimurium

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Julia V. Monjarás Feria ◽  
Matthew D. Lefebre ◽  
York-Dieter Stierhof ◽  
Jorge E. Galán ◽  
Samuel Wagner
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Effector Proteins ◽  
Switching Function ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Gram Negative ◽  
Type Iii ◽  
Autocatalytic Cleavage ◽  
Serovar Typhimurium

ABSTRACTType III secretion systems (T3SSs) are multiprotein machines employed by many Gram-negative bacteria to inject bacterial effector proteins into eukaryotic host cells to promote bacterial survival and colonization. The core unit of T3SSs is the needle complex, a supramolecular structure that mediates the passage of the secreted proteins through the bacterial envelope. A distinct feature of the T3SS is that protein export occurs in a strictly hierarchical manner in which proteins destined to form the needle complex filament and associated structures are secreted first, followed by the secretion of effectors and the proteins that will facilitate their translocation through the target host cell membrane. The secretion hierarchy is established by complex mechanisms that involve several T3SS-associated components, including the “switch protein,” a highly conserved, inner membrane protease that undergoes autocatalytic cleavage. It has been proposed that the autocleavage of the switch protein is the trigger for substrate switching. We show here that autocleavage of theSalmonella entericaserovar Typhimurium switch protein SpaS is an unregulated process that occurs after its folding and before its incorporation into the needle complex. Needle complexes assembled with a precleaved form of SpaS function in a manner indistinguishable from that of the wild-type form. Furthermore, an engineered mutant of SpaS that is processed by an external protease also displays wild-type function. These results demonstrate that the cleavage eventper sedoes not provide a signal for substrate switching but support the hypothesis that cleavage allows the proper conformation of SpaS to render it competent for its switching function.IMPORTANCEBacterial interaction with eukaryotic hosts often involves complex molecular machines for targeted delivery of bacterial effector proteins. One such machine, the type III secretion system of some Gram-negative bacteria, serves to inject a multitude of structurally diverse bacterial proteins into the host cell. Critical to the function of these systems is their ability to secrete proteins in a strict hierarchical order, but it is unclear how the mechanism of switching works. Central to the switching mechanism is a highly conserved inner membrane protease that undergoes autocatalytic cleavage. Although it has been suggested previously that the autocleavage event is the trigger for substrate switching, we show here that this is not the case. Rather, our results show that cleavage allows the proper conformation of the protein to render it competent for its switching function. These findings may help develop inhibitors of type III secretion machines that offer novel therapeutic avenues to treat various infectious diseases.

scholarly journals Small-Molecule Type III Secretion System Inhibitors Block Assembly of the Shigella Type III Secreton

2008 ◽  
Vol 191 (2) ◽  
pp. 563-570 ◽  
Author(s):  
Andreas K. J. Veenendaal ◽  
Charlotta Sundin ◽  
Ariel J. Blocker
Keyword(s):  
Type Iii Secretion ◽  
Bacterial Infections ◽  
Shigella Flexneri ◽  
Effector Proteins ◽  
Host Cells ◽  
Incomplete Penetrance ◽  
Secretion Systems ◽  
Bacterial Surface ◽  
Type Iii ◽  
Type Iii Secretion Systems

ABSTRACT Type III secretion systems (T3SSs) are essential virulence devices for many gram-negative bacteria that are pathogenic for plants, animals, and humans. They serve to translocate virulence effector proteins directly into eukaryotic host cells. T3SSs are composed of a large cytoplasmic bulb and a transmembrane region into which a needle is embedded, protruding above the bacterial surface. The emerging antibiotic resistance of bacterial pathogens urges the development of novel strategies to fight bacterial infections. Therapeutics that rather than kill bacteria only attenuate their virulence may reduce the frequency or progress of resistance emergence. Recently, a group of salicylidene acylhydrazides were identified as inhibitors of T3SSs in Yersinia, Chlamydia, and Salmonella species. Here we show that these are also effective on the T3SS of Shigella flexneri, where they block all related forms of protein secretion so far known, as well as the epithelial cell invasion and induction of macrophage apoptosis usually demonstrated by this bacterium. Furthermore, we show the first evidence for the detrimental effect of these compounds on T3SS needle assembly, as demonstrated by increased numbers of T3S apparatuses without needles or with shorter needles. Therefore, the compounds generate a phenocopy of T3SS export apparatus mutants but with incomplete penetrance. We discuss why this would be sufficient to almost completely block the later secretion of effector proteins and how this begins to narrow the search for the molecular target of these compounds.

scholarly journals A NanoLuc luciferase-based assay enabling the real-time analysis of protein secretion and injection by bacterial type III secretion systems

2019 ◽  
Author(s):  
Sibel Westerhausen ◽  
Melanie Nowak ◽  
Claudia Torres-Vargas ◽  
Ursula Bilitewski ◽  
Erwin Bohn ◽  
...  
Keyword(s):  
Protein Secretion ◽  
High Throughput ◽  
Type Iii Secretion ◽  
Molecular Mechanisms ◽  
Host Cells ◽  
Target Cells ◽  
Secretion Systems ◽  
Type Iii ◽  
Nanoluc Luciferase ◽  
Type Iii Secretion Systems

AbstractThe elucidation of the molecular mechanisms of secretion through bacterial protein secretion systems is impeded by a lack of assays to quantitatively assess secretion kinetics. Also the analysis of the biological role of these secretion systems as well as the identification of inhibitors targeting these systems would greatly benefit from the availability of a simple, quick and quantitative assay to monitor principle secretion and injection into host cells. Here we present a versatile solution to this need, utilizing the small and very bright NanoLuc luciferase to assess secretion and injection through the type III secretion system encoded by Salmonella pathogenicity island 1. The NanoLuc-based secretion assay features a very high signal-to-noise ratio and sensitivity down to the nanoliter scale. The assay enables monitoring of secretion kinetics and is adaptable to a high throughput screening format in 384-well microplates. We further developed NanoLuc and split-NanoLuc-based assays that enable the monitoring of type III secretion-dependent injection of effector proteins into host cells.ImportanceThe ability to secrete proteins to the bacterial cell surface, to the extracellular environment, or even into target cells is one of the foundations of interbacterial as well as pathogen-host interaction. While great progress has been made in elucidating assembly and structure of secretion systems, our understanding of their secretion mechanism often lags behind, not last because of the challenge to quantitatively assess secretion function. Here, we developed a luciferase-based assay to enable the simple, quick, quantitative, and high throughput-compatible assessment of secretion and injection through virulence-associated type III secretion systems. The assay allows detection of minute amounts of secreted substrate proteins either in the supernatant of the bacterial culture or within eukaryotic host cells. It thus provides an enabling technology to elucidate the mechanisms of secretion and injection of type III secretion systems and is likely adaptable to assay secretion through other bacterial secretion systems.

scholarly journals Identification of Novel Type III Secretion Effectors in Xanthomonas oryzae pv. oryzae

2009 ◽  
Vol 22 (1) ◽  
pp. 96-106 ◽  
Author(s):  
Ayako Furutani ◽  
Minako Takaoka ◽  
Harumi Sanada ◽  
Yukari Noguchi ◽  
Takashi Oku ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Regulatory Protein ◽  
Effector Proteins ◽  
Xanthomonas Oryzae ◽  
Dependent Manner ◽  
Plant Pathogenic Bacteria ◽  
Type Iii ◽  
Genes Encoding

Many gram-negative bacteria secrete so-called effector proteins via a type III secretion (T3S) system. Through genome screening for genes encoding potential T3S effectors, 60 candidates were selected from rice pathogen Xanthomonas oryzae pv. oryzae MAFF311018 using these criteria: i) homologs of known T3S effectors in plant-pathogenic bacteria, ii) genes with expression regulated by hrp regulatory protein HrpX, or iii) proteins with N-terminal amino acid patterns associated with T3S substrates of Pseudomonas syringae. Of effector candidates tested with the Bordetella pertussis calmodulin-dependent adenylate cyclase reporter for translocation into plant cells, 16 proteins were translocated in a T3S system-dependent manner. Of these 16 proteins, nine were homologs of known effectors in other plant-pathogenic bacteria and seven were not. Most of the effectors were widely conserved in Xanthomonas spp.; however, some were specific to X. oryzae. Interestingly, all these effectors were expressed in an HrpX-dependent manner, suggesting coregulation of effectors and the T3S system. In X. campestris pv. vesicatoria, HpaB and HpaC (HpaP in X. oryzae pv. oryzae) have a central role in recruiting T3S substrates to the secretion apparatus. Secretion of all but one effector was reduced in both HpaB– and HpaP– mutant strains, indicating that HpaB and HpaP are widely involved in efficient secretion of the effectors.

scholarly journals Posttranscriptional Control of Microbe-Induced Rearrangement of Host Cell Actin

mBio ◽  
2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Charley C. Gruber ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Cell Biology ◽  
Molecular Mechanisms ◽  
Bacterial Attachment ◽  
Posttranscriptional Control ◽  
Content Type ◽  
Type Iii ◽  
Pedestal Formation

ABSTRACTRemodeling of the host cytoskeleton is a common strategy employed by bacterial pathogens. Although there is vigorous investigation of the cell biology underlying these bacterially mediated cytoskeleton modifications, knowledge of the plasticity and dynamics of the bacterial signaling networks that regulate the expression of genes necessary for these phenotypes is lacking. EnterohemorrhagicEscherichia coliattaches to enterocytes, forming pedestal-like structures. Pedestal formation requires the expression of the locus-of-enterocyte-effacement (LEE) andespFugenes. The LEE encodes a molecular syringe, a type III secretion system (T3SS) used by pathogens to translocate effectors such as EspFu into the host cell. By using a combination of genetic, biochemical, and cell biology approaches, we show that pedestal formation relies on posttranscriptional regulation by two small RNAs (sRNAs), GlmY and GlmZ. The GlmY and GlmZ sRNAs are unique; they have extensive secondary structures and work in concert. Although these sRNAs may offer unique insights into RNA and posttranscriptional biology, thus far, only one target and one mechanism of action (exposure of the ribosome binding site from theglmSgene to promote its translation) has been described. Here we uncovered new targets and two different molecular mechanisms of action of these sRNAs. In the case of EspFu expression, they promote translation by cleavage of the transcript, while in regard to the LEE, they promote destabilization of the mRNA. Our findings reveal that two unique sRNAs act in concert through different molecular mechanisms to coordinate bacterial attachment to mammalian cells.IMPORTANCEPathogens evolve by horizontal acquisition of pathogenicity islands. We describe here how two sRNAs, GlmY and GlmZ, involved in cellular metabolism and cellular architecture, through the posttranscriptional control of GlmS (the previously only known target of GlmY and GlmZ), which controls amino sugar synthesis, have been coopted to modulate the expression of virulence. These sRNAs quickly allow for plasticity in gene expression in order for enterohemorrhagicEscherichia colito fine-tune the expression of its complex type III secretion machinery and its effectors to promote bacterial attachment and subsequent actin rearrangement on host cells. Pedestal formation is a very dynamic process. Many of the genes necessary for pedestal formation are located within the same operon to evolutionarily guarantee that they are inherited together. However, it is worth noting that within these operons, several genes need to yield more proteins than others and that these differences cannot be efficiently regulated at the transcriptional level.

scholarly journals Identification of a Specific Chaperone for SptP, a Substrate of the Centisome 63 Type III Secretion System ofSalmonella typhimurium

1998 ◽  
Vol 180 (13) ◽  
pp. 3393-3399 ◽  
Author(s):  
Yixin Fu ◽  
Jorge E. Galán
Keyword(s):  
Actin Cytoskeleton ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Tyrosine Phosphatase ◽  
Secretion System ◽  
Effector Proteins ◽  
Loss Of Function ◽  
Secretion Systems ◽  
Type Iii ◽  
Type Iii Protein Secretion

ABSTRACT Salmonella typhimurium uses of a type III protein secretion system encoded at centisome 63 of its chromosome to deliver effector molecules into the host cell. These proteins stimulate host cell responses such as reorganization of the actin cytoskeleton and activation of transcription factors. One of these effector proteins is SptP, a tyrosine phosphatase that causes disruption of the host cell actin cytoskeleton. A characteristic feature of many substrates of type III secretion systems is their association with specific cytoplasmic chaperones which appears to be required for secretion and/or translocation of these proteins into the host cell. We report here the identification of SicP, a 13-kDa acidic polypeptide that is encoded immediately upstream of sptP. A loss-of-function mutation in sicP resulted in drastically reduced levels of SptP but did not affect sptP expression, indicating that SicP exerts its effect posttranscriptionally. Pulse-chase experiments demonstrated that the loss of SicP leads to increased degradation of SptP. In addition, we show that SicP binds to SptP directly and that the binding site is located between residues 15 and 100 of the tyrosine phosphatase. Taken together, these results indicate that SicP acts as a specific chaperone for SptP.

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