scholarly journals Identification of New Secreted Effectors in Salmonella enterica Serovar Typhimurium

2005 ◽  
Vol 73 (10) ◽  
pp. 6260-6271 ◽  
Author(s):  
Kaoru Geddes ◽  
Micah Worley ◽  
George Niemann ◽  
Fred Heffron
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Bacterial Species ◽  
Effector Proteins ◽  
Host Cells ◽  
Secretion Systems ◽  
Cell Cytoplasm ◽  
Type Iii ◽  
Serovar Typhimurium ◽  
Infected Cells

ABSTRACTA common theme in bacterial pathogenesis is the secretion of bacterial products that modify cellular functions to overcome host defenses. Gram-negative bacterial pathogens use type III secretion systems (TTSSs) to inject effector proteins into host cells. The genes encoding the structural components of the type III secretion apparatus are conserved among bacterial species and can be identified by sequence homology. In contrast, the sequences of secreted effector proteins are less conserved and are therefore difficult to identify. A strategy was developed to identify virulence factors secreted bySalmonella entericaserovar Typhimurium into the host cell cytoplasm. We constructed a transposon, which we refer to as mini-Tn5-cycler, to generate translational fusions betweenSalmonellachromosomal genes and a fragment of the calmodulin-dependent adenylate cyclase gene derived fromBordetella pertussis(cyaA′). In-frame fusions to bacterial proteins that are secreted into the eukaryotic cell cytoplasm were identified by high levels of cyclic AMP in infected cells. The assay was sufficiently sensitive that a single secreted fusion could be identified among several hundred that were not secreted. This approach identified three new effectors as well as seven that have been previously characterized. A deletion of one of the new effectors,steA(Salmonellatranslocated effector A), attenuated virulence. In addition, SteA localizes to thetrans-Golgi network in both transfected and infected cells. This approach has identified new secreted effector proteins inSalmonellaand will likely be useful for other organisms, even those in which genetic manipulation is more difficult.

scholarly journals Cytosporone B, an Inhibitor of the Type III Secretion System of Salmonella enterica Serovar Typhimurium

2013 ◽  
Vol 57 (5) ◽  
pp. 2191-2198 ◽  
Author(s):  
Jianfang Li ◽  
Chao Lv ◽  
Weiyang Sun ◽  
Zhenyu Li ◽  
Xiaowei Han ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Bacterial Resistance ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Content Type ◽  
Type Iii ◽  
Serovar Typhimurium

ABSTRACTBacterial virulence factors have been increasingly regarded as attractive targets for development of novel antibacterial agents. Virulence inhibitors are less likely to generate bacterial resistance, which makes them superior to traditional antibiotics that target bacterial viability.Salmonella entericaserovar Typhimurium, an important food-borne human pathogen, has type III secretion system (T3SS) as its major virulence factor. T3SS secretes effector proteins to facilitate invasion into host cells. In this study, we identified several analogs of cytosporone B (Csn-B) that strongly block the secretion ofSalmonellapathogenicity island 1 (SPI-1)-associated effector proteins, without affecting the secretion of flagellar protein FliCin vitro. Csn-B and two other derivatives exhibited a strong inhibitory effect on SPI-1-mediated invasion to HeLa cells, while no significant toxicity to bacteria was observed. Nucleoid proteins Hha and H-NS bind to the promoters of SPI-1 regulator geneshilD,hilC, andrtsAto repress their expression and consequently regulate the expression of SPI-1 apparatus and effector genes. We found that Csn-B upregulated the transcription ofhhaandhns, implying that Csn-B probably affected the secretion of effectors through the Hha–H-NS regulatory pathway. In summary, this study presented an effective SPI-1 inhibitor, Csn-B, which may have potential in drug development against antibiotic-resistantSalmonella.

scholarly journals The First 45 Amino Acids of SopA Are Necessary for InvB Binding and SPI-1 Secretion

2006 ◽  
Vol 188 (7) ◽  
pp. 2411-2420 ◽  
Author(s):  
Wendy Higashide ◽  
Daoguo Zhou
Keyword(s):  
Type Iii Secretion ◽  
Catalytic Domain ◽  
Effector Proteins ◽  
Host Cells ◽  
Amino Acid Residues ◽  
Secretion Systems ◽  
Reporter System ◽  
Type Iii ◽  
Serovar Typhimurium ◽  
Type Iii Protein Secretion

ABSTRACT Salmonella enterica serovar Typhimurium encodes two type III secretion systems (TTSSs) within pathogenicity island 1 (SPI-1) and island 2 (SPI-2). These type III protein secretion and translocation systems transport a panel of bacterial effector proteins across both the bacterial and the host cell membranes to promote bacterial entry and subsequent survival inside host cells. Effector proteins contain secretion and translocation signals that are often located at their N termini. We have developed a ruffling-based translocation reporter system that uses the secretion- and translocation-deficient catalytic domain of SopE, SopE78-240, as a reporter. Using this assay, we determined that the N-terminal 45 amino acid residues of Salmonella SopA are necessary and sufficient for directing its secretion and translocation through the SPI-1 TTSS. SopA1-45, but not SopA1-44, is also able to bind to its chaperone, InvB, indicating that SPI-1 type III secretion and translocation of SopA require its chaperone.

scholarly journals Optimization of the Delivery of Heterologous Proteins by the Salmonella enterica Serovar Typhimurium Type III Secretion System for Vaccine Development

2006 ◽  
Vol 74 (10) ◽  
pp. 5826-5833 ◽  
Author(s):  
Li-Mei Chen ◽  
Gabriel Briones ◽  
Ruben O. Donis ◽  
Jorge E. Galán
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Vaccine Development ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Host Cells ◽  
Antigen Delivery ◽  
Secretion Systems ◽  
Type Iii ◽  
Serovar Typhimurium

ABSTRACT Type III protein secretion systems, which are organelles with the capacity to deliver bacterial proteins into host cells, have been adapted to deliver heterologous antigens for vaccine development. A limitation of these antigen delivery systems is that some proteins are not amenable to secretion through this pathway. We show here that proteins from the simian and human immunodeficiency viruses that are not permissive for secretion through a Salmonella enterica serovar Typhimurium type III secretion system can be modified to travel this secretion pathway by introduction of discrete mutations. Proteins optimized for secretion were presented more efficiently via the major histocompatibility complex class I pathway and were able to induce a better immune response.

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 Salmonella type III effector, PipA, works in a different manner than the PipA family effectors GogA and GtgA

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0248975
Author(s):  
Momo Takemura ◽  
Takeshi Haneda ◽  
Hikari Idei ◽  
Tsuyoshi Miki ◽  
Nobuhiko Okada
Keyword(s):  
Hela Cells ◽  
Critical Role ◽  
Effector Proteins ◽  
Host Cells ◽  
Microbial Pathogens ◽  
Secretion Systems ◽  
Type Iii ◽  
Zinc Metalloprotease ◽  
Type Iii Secretion Systems ◽  
Serovar Typhimurium

Nuclear factor-kappa B (NF-κB) plays a critical role in the host defense against microbial pathogens. Many pathogens modulate NF-κB signaling to establish infection in their host. Salmonella enterica serovar Typhimurium (S. Typhimurium) possesses two type III secretion systems (T3SS-1 and T3SS-2) and directly injects many effector proteins into host cells. It has been reported that some effectors block NF-κB signaling, but the molecular mechanism of the inactivation of NF-κB signaling in S. Typhimurium is poorly understood. Here, we identified seven type III effectors—GogA, GtgA, PipA, SseK1, SseK2, SseK3, and SteE—that inhibited NF-κB activation in HeLa cells stimulated with TNF-α. We also determined that only GogA and GtgA are involved in regulation of the activation of NF-κB in HeLa cells infected with S. Typhimurium. GogA, GtgA, and PipA are highly homologous to one another and have the consensus zinc metalloprotease HEXXH motif. Our experiments demonstrated that GogA, GtgA, and PipA each directly cleaved NF-κB p65, whereas GogA and GtgA, but not PipA, inhibited the NF-κB activation in HeLa cells infected with S. Typhimurium. Further, expressions of the gogA or gtgA gene were induced under the SPI-1-and SPI-2-inducing conditions, but expression of the pipA gene was induced only under the SPI-2-inducing condition. We also showed that PipA was secreted into RAW264.7 cells through T3SS-2. Finally, we indicated that PipA elicits bacterial dissemination in the systemic stage of infection of S. Typhimurium via a T3SS-1-independent mechanism. Collectively, our results suggest that PipA, GogA and GtgA contribute to S. Typhimurium pathogenesis in different ways.

scholarly journals Interactions between effector proteins of the Pseudomonas aeruginosa type III secretion system do not significantly affect several measures of disease severity in mammals

Microbiology ◽  
2006 ◽  
Vol 152 (1) ◽  
pp. 143-152 ◽  
Author(s):  
Ciara M. Shaver ◽  
Alan R. Hauser
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Disease Severity ◽  
Type Iii Secretion ◽  
Disease Process ◽  
Effector Proteins ◽  
Host Cells ◽  
Secreted Proteins ◽  
Secretion Systems ◽  
Type Iii

The effector proteins of the type III secretion systems of many bacterial pathogens act in a coordinated manner to subvert host cells and facilitate the development and progression of disease. It is unclear whether interactions between the type-III-secreted proteins of Pseudomonas aeruginosa result in similar effects on the disease process. We have previously characterized the contributions to pathogenesis of the type-III-secreted proteins ExoS, ExoT and ExoU when secreted individually. In this study, we extend our prior work to determine whether these proteins have greater than expected effects on virulence when secreted in combination. In vitro cytotoxicity and anti-internalization activities were not enhanced when effector proteins were secreted in combinations rather than alone. Likewise in a mouse model of pneumonia, bacterial burden in the lungs, dissemination and mortality attributable to ExoS, ExoT and ExoU were not synergistically increased when combinations of these effector proteins were secreted. Because of the absence of an appreciable synergistic increase in virulence when multiple effector proteins were secreted in combination, we conclude that any cooperation between ExoS, ExoT and ExoU does not translate into a synergistically significant enhancement of disease severity as measured by these assays.

scholarly journals Impact of Salmonella enterica Type III Secretion System Effectors on the Eukaryotic Host Cell

2012 ◽  
Vol 2012 ◽  
pp. 1-36 ◽  
Author(s):  
Francisco Ramos-Morales
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Current Knowledge ◽  
Cellular Level ◽  
Host Cells ◽  
Secretion Systems ◽  
Type Iii ◽  
Cellular Processes ◽  
Type Iii Secretion Systems ◽  
Near Future

Type III secretion systems are molecular machines used by many Gram-negative bacterial pathogens to inject proteins, known as effectors, directly into eukaryotic host cells. These proteins manipulate host signal transduction pathways and cellular processes to the pathogen’s advantage. Salmonella enterica possesses two virulence-related type III secretion systems that deliver more than forty effectors. This paper reviews our current knowledge about the functions, biochemical activities, host targets, and impact on host cells of these effectors. First, the concerted action of effectors at the cellular level in relevant aspects of the interaction between Salmonella and its hosts is analyzed. Then, particular issues that will drive research in the field in the near future are discussed. Finally, detailed information about each individual effector is provided.

scholarly journals Dynamics of the Type III Secretion System Activity of Enteropathogenic Escherichia coli

mBio ◽  
2013 ◽  
Vol 4 (4) ◽  
Author(s):  
Erez Mills ◽  
Kobi Baruch ◽  
Gili Aviv ◽  
Mor Nitzan ◽  
Ilan Rosenshine
Keyword(s):  
Escherichia Coli ◽  
Type Iii Secretion ◽  
Public Health Problem ◽  
Effector Proteins ◽  
Host Cells ◽  
Global Public Health ◽  
Secretion Systems ◽  
Comprehensive Description ◽  
Content Type ◽  
Type Iii

ABSTRACT Type III secretion systems (TTSSs) are employed by pathogens to translocate host cells with effector proteins, which are crucial for virulence. The dynamics of effector translocation, behavior of the translocating bacteria, translocation temporal order, and relative amounts of each of the translocated effectors are all poorly characterized. To address these issues, we developed a microscopy-based assay that tracks effector translocation. We used this assay alongside a previously described real-time population-based translocation assay, focusing mainly on enteropathogenic Escherichia coli (EPEC) and partly comparing it to Salmonella. We found that the two pathogens exhibit different translocation behaviors: in EPEC, a subpopulation that formed microcolonies carried out most of the translocation activity, while Salmonella executed protein translocation as planktonic bacteria. We also noted variability in host cell susceptibility, with some cells highly resistant to translocation. We next extended the study to determine the translocation dynamics of twenty EPEC effectors and found that all exhibited distinct levels of translocation efficiency. Further, we mapped the global effects of key TTSS-related components on TTSS activity. Our results provide a comprehensive description of the dynamics of the TTSS activity of EPEC and new insights into the mechanisms that control the dynamics. IMPORTANCE EPEC and the closely related enterohemorrhagic Escherichia coli (EHEC) represent a global public health problem. New strategies to combat EPEC and EHEC infections are needed, and development of such strategies requires better understanding of their virulence machinery. The TTSS is a critical virulence mechanism employed by these pathogens, and by others, including Salmonella. In this study, we aimed at elucidating new aspects of TTSS function. The results obtained provide a comprehensive description of the dynamics of TTSS activity of EPEC and new insights into the mechanisms that control these changes. This knowledge sets the stage for further analysis of the system and may accelerate the development of new ways to treat EPEC and EHEC infections. Further, the newly described microscopy-based assay can be readily adapted to study the dynamics of TTSS activity in other pathogens.

scholarly journals Effect of the O-Antigen Length of Lipopolysaccharide on the Functions of Type III Secretion Systems in Salmonella enterica

2009 ◽  
Vol 77 (12) ◽  
pp. 5458-5470 ◽  
Author(s):  
Stefanie U. Hölzer ◽  
Markus C. Schlumberger ◽  
Daniela Jäckel ◽  
Michael Hensel
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Defense Mechanisms ◽  
Host Cells ◽  
Effector Protein ◽  
Secretion Systems ◽  
Type Iii ◽  
O Antigen ◽  
T3ss Effector ◽  
Type Iii Secretion Systems

ABSTRACT The virulence of Salmonella enterica critically depends on the functions of two type III secretion systems (T3SS), with the Salmonella pathogenicity island 1 (SPI1)-encoded T3SS required for host cell invasion and the SPI2-T3SS enabling Salmonella to proliferate within host cells. A further T3SS is required for the assembly of the flagella. Most serovars of Salmonella also possess a lipopolysaccharide with a complex O-antigen (OAg) structure. The number of OAg units attached to the core polysaccharide varies between 16 and more than 100 repeats, with a trimodal distribution. This work investigated the correlation of the OAg length with the functions of the SPI1-T3SS and the SPI2-T3SS. We observed that the number of repeats of OAg units had no effect on bacterial motility. The interaction of Salmonella with epithelial cells was altered if the OAg structure was changed by mutations in regulators of OAg. Strains defective in synthesis of very long or long and very long OAg species showed increased translocation of a SPI1-T3SS effector protein and increased invasion. Invasion of a strain entirely lacking OAg was increased, but this mutant strain also showed increased adhesion. In contrast, translocation of a SPI2-T3SS effector protein and intracellular replication were not affected by modification of the OAg length. Mutant strains lacking the entire OAg or long and very long OAg were highly susceptible to complement killing. These observations indicate that the architecture of the outer membrane of Salmonella is balanced to permit sufficient T3SS function but also to confer optimal protection against antimicrobial defense mechanisms.

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