scholarly journals The Role of the Membrane-Associated Domain of the Export Apparatus Protein, EscV (SctV), in the Activity of the Type III Secretion System

2021 ◽  
Vol 12 ◽  
Author(s):  
Boško Mitrović ◽  
Shir Lezerovich ◽  
Neta Sal-Man
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Host Cells ◽  
Health Concern ◽  
Loss Of Function ◽  
Reporter System ◽  
Host Cell Membrane ◽  
Type Iii

Diarrheal diseases remain a major public health concern worldwide. Many of the causative bacterial pathogens that cause these diseases have a specialized protein complex, the type III secretion system (T3SS), which delivers effector proteins directly into host cells. These effectors manipulate host cell processes for the benefit of the infecting bacteria. The T3SS structure resembles a syringe anchored within the bacterial membrane, projecting toward the host cell membrane. The entry port of the T3SS substrates, called the export apparatus, is formed by five integral membrane proteins. Among the export apparatus proteins, EscV is the largest, and as it forms a nonamer, it constitutes the largest portion of the export apparatus complex. While there are considerable data on the soluble cytoplasmic domain of EscV, our knowledge of its membrane-associated section and its transmembrane domains (TMDs) is still very limited. In this study, using an isolated genetic reporter system, we found that TMD5 and TMD6 of EscV mediate strong self-oligomerization. Substituting these TMDs within the full-length protein with a random hydrophobic sequence resulted in a complete loss of function of the T3SS, further suggesting that the EscV TMD5 and TMD6 sequences have a functional role in addition to their structural role as membrane anchors. As we observed only mild reduction in the ability of the TMD-exchanged variants to integrate into the full or intermediate T3SS complexes, we concluded that EscV TMD5 and TMD6 are not crucial for the global assembly or stability of the T3SS complex but are rather involved in promoting the necessary TMD–TMD interactions within the complex and the overall TMD orientation to allow channel opening for the entry of T3SS substrates.

scholarly journals Identification of Two Translocon Proteins of Vibrio parahaemolyticus Type III Secretion System 2

2008 ◽  
Vol 76 (9) ◽  
pp. 4282-4289 ◽  
Author(s):  
Toshio Kodama ◽  
Hirotaka Hiyoshi ◽  
Kazuyoshi Gotoh ◽  
Yukihiro Akeda ◽  
Shigeaki Matsuda ◽  
...  
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Vibrio Parahaemolyticus ◽  
Critical Role ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Host Cell Membrane ◽  
Type Iii

ABSTRACT The type III secretion system (T3SS) translocon complex is composed of several associated proteins, which form a translocation channel through the host cell plasma membrane. These proteins are key molecules that are involved in the pathogenicity of many T3SS-positive bacteria, because they are necessary to deliver effector proteins into host cells. A T3SS designated T3SS2 of Vibrio parahaemolyticus is thought to be related to the enterotoxicity of this bacterium in humans, but the effector translocation mechanism of T3SS2 is unclear because there is only one gene (the VPA1362 gene) in the T3SS2 region that is homologous to other translocon protein genes. It is also not known whether the VPA1362 protein is functional in the translocon of T3SS2 or whether it is sufficient to form the translocation channel of T3SS2. In this study, we identified both VPA1362 (designated VopB2) and VPA1361 (designated VopD2) as T3SS2-dependent secretion proteins. Functional analysis of these proteins showed that they are essential for T3SS2-dependent cytotoxicity, for the translocation of one of the T3SS2 effector proteins (VopT), and for the contact-dependent activity of pore formation in infected cells in vitro. Their targeting to the host cell membrane depends on T3SS2, and furthermore, they are necessary for T3SS2-dependent enterotoxicity in vivo. These results indicate that VopB2 and VopD2 act as translocon proteins of V. parahaemolyticus T3SS2 and hence have a critical role in the T3SS2-dependent enterotoxicity of this bacterium.

scholarly journals The Ruler Protein EscP of the Enteropathogenic Escherichia coli Type III Secretion System Is Involved in Calcium Sensing and Secretion Hierarchy Regulation by Interacting with the Gatekeeper Protein SepL

mBio ◽  
2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Lihi Shaulov ◽  
Jenia Gershberg ◽  
Wanyin Deng ◽  
B. Brett Finlay ◽  
Neta Sal-Man
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Bacterial Pathogens ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Bacterial Virulence ◽  
Effector Proteins ◽  
Host Cells ◽  
Gram Negative ◽  
Type Iii

ABSTRACT The type III secretion system (T3SS) is a multiprotein complex that plays a central role in the virulence of many Gram-negative bacterial pathogens. To ensure that effector proteins are efficiently translocated into the host cell, bacteria must be able to sense their contact with the host cell. In this study, we found that EscP, which was previously shown to function as the ruler protein of the enteropathogenic Escherichia coli T3SS, is also involved in the switch from the secretion of translocator proteins to the secretion of effector proteins. In addition, we demonstrated that EscP can interact with the gatekeeper protein SepL and that the EscP-SepL complex dissociates upon a calcium concentration drop. We suggest a model in which bacterial contact with the host cell is accompanied by a drop in the calcium concentration that causes SepL-EscP complex dissociation and triggers the secretion of effector proteins. IMPORTANCE The emergence of multidrug-resistant bacterial strains, especially those of pathogenic bacteria, has serious medical and clinical implications. At the same time, the development and approval of new antibiotics have been limited for years. Recently, antivirulence drugs have received considerable attention as a novel antibiotic strategy that specifically targets bacterial virulence rather than growth, an approach that applies milder evolutionary pressure on the bacteria to develop resistance. A highly attractive target for the development of antivirulence compounds is the type III secretion system, a specialized secretory system possessed by many Gram-negative bacterial pathogens for injecting virulence factors (effectors) into host cells. In this study, we shed light on the molecular mechanism that allows bacteria to sense their contact with the host cell and to respond with the timed secretion of effector proteins. Understanding this critical step for bacterial virulence may provide a new therapeutic strategy. IMPORTANCE The emergence of multidrug-resistant bacterial strains, especially those of pathogenic bacteria, has serious medical and clinical implications. At the same time, the development and approval of new antibiotics have been limited for years. Recently, antivirulence drugs have received considerable attention as a novel antibiotic strategy that specifically targets bacterial virulence rather than growth, an approach that applies milder evolutionary pressure on the bacteria to develop resistance. A highly attractive target for the development of antivirulence compounds is the type III secretion system, a specialized secretory system possessed by many Gram-negative bacterial pathogens for injecting virulence factors (effectors) into host cells. In this study, we shed light on the molecular mechanism that allows bacteria to sense their contact with the host cell and to respond with the timed secretion of effector proteins. Understanding this critical step for bacterial virulence may provide a new therapeutic strategy.

scholarly journals Transcriptional Induction of the Pseudomonas aeruginosa Type III Secretion System by Low Ca2+ and Host Cell Contact Proceeds through Two Distinct Signaling Pathways

2006 ◽  
Vol 74 (6) ◽  
pp. 3334-3341 ◽  
Author(s):  
Nandini Dasgupta ◽  
Alix Ashare ◽  
Gary W. Hunninghake ◽  
Timothy L. Yahr
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Mammalian Cells ◽  
Type Iii Secretion System ◽  
Growth Conditions ◽  
Secretion System ◽  
Host Cells ◽  
Dependent Manner ◽  
Type Iii

ABSTRACT The opportunistic pathogen Pseudomonas aeruginosa utilizes a type III secretion system (T3SS) to intoxicate eukaryotic host cells. Transcription of the T3SS is induced under calcium-limited growth conditions or following intimate contact of P. aeruginosa with host cells. In the present study, we demonstrate that expression of the T3SS is controlled by two distinct regulatory mechanisms and that these mechanisms are differentially activated in a host cell-dependent manner. The first mechanism is dependent upon ExsC, a regulatory protein that couples transcription of the T3SS to the activity of the type III secretion machinery. ExsC is essential for induction of the T3SS under low-calcium-growth conditions and for T3SS-dependent cytotoxicity towards social amoebae, insect cells, and erythrocytes. The second regulatory mechanism functions independently of ExsC and is sufficient to elicit T3SS-dependent cytotoxicity towards certain types of mammalian cells. Although this second pathway (ExsC independent) is sufficient, an exsC mutant demonstrates a lag in the induction of cytotoxicity towards Chinese hamster ovary cells and is attenuated for virulence in a mouse pneumonia model. We propose that the ExsC-dependent pathway is required for full cytotoxicity towards all host cell types tested whereas the ExsC-independent pathway may represent an adaptation that allows P. aeruginosa to increase expression of the T3SS in response to specific types of mammalian cells.

scholarly journals IpaD Localizes to the Tip of the Type III Secretion System Needle of Shigella flexneri

2006 ◽  
Vol 74 (8) ◽  
pp. 4391-4400 ◽  
Author(s):  
Marianela Espina ◽  
Andrew J. Olive ◽  
Roma Kenjale ◽  
David S. Moore ◽  
S. Fernando Ausar ◽  
...  
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Actin Polymerization ◽  
Shigella Flexneri ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Erythrocyte Membranes ◽  
Host Cell Membrane ◽  
Type Iii ◽  
Surface Localization

ABSTRACT Shigella flexneri, the causative agent of shigellosis, is a gram-negative bacterial pathogen that initiates infection by invading cells within the colonic epithelium. Contact with host cell surfaces induces a rapid burst of protein secretion via the Shigella type III secretion system (TTSS). The first proteins secreted are IpaD, IpaB, and IpaC, with IpaB and IpaC being inserted into the host cell membrane to form a pore for translocating late effectors into the target cell cytoplasm. The resulting pathogen-host cross talk results in localized actin polymerization, membrane ruffling, and, ultimately, pathogen entry. IpaD is essential for host cell invasion, but its role in this process is just now coming to light. IpaD is a multifunctional protein that controls the secretion and presentation of IpaB and IpaC at the pathogen-host interface. We show here that antibodies recognizing the surface-exposed N terminus of IpaD neutralize Shigella's ability to promote pore formation in erythrocyte membranes. We further show that MxiH and IpaD colocalize on the bacterial surface. When TTSS needles were sheared from the Shigella surface, IpaD was found at only the needle tips. Consistent with this, IpaD localized to the exposed tips of needles that were still attached to the bacterium. Molecular analyses then showed that the IpaD C terminus is required for this surface localization and function. Furthermore, mutations that prevent IpaD surface localization also eliminate all IpaD-related functions. Thus, this study demonstrates that IpaD localizes to the TTSA needle tip, where it functions to control the secretion and proper insertion of translocators into host cell membranes.

scholarly journals The Type III System-Secreted Effector EspZ Localizes to Host Mitochondria and Interacts with the Translocase of Inner Mitochondrial Membrane 17b

2011 ◽  
Vol 79 (12) ◽  
pp. 4784-4790 ◽  
Author(s):  
Stephanie R. Shames ◽  
Matthew A. Croxen ◽  
Wanyin Deng ◽  
B. Brett Finlay
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Mitochondrial Membrane ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Infected Host ◽  
Inner Mitochondrial Membrane ◽  
Type Iii

ABSTRACTEnteropathogenic and enterohemorrhagicEscherichia coli(EPEC and EHEC, respectively) are attaching and effacing (A/E) bacterial pathogens that cause severe diarrheal disease worldwide. To cause disease, A/E pathogens require a type III secretion system, which facilitates transport of bacterial effector proteins directly into infected host cells. One of these effector proteins translocated by the type III secretion system, EspZ, is essential for A/E pathogen infection and functions to prevent rapid death of EPEC-infected cells. We further investigated the mechanism of EspZ-mediated protection of infected host cells and found that a severe decrease in host mitochondrial membrane potential (Δψm) occurs concurrently with host cell lysis during infection with EPEC lacking EspZ (ΔespZ). It was also demonstrated that EspZ localizes to host cell mitochondria and interacts with the translocase of inner mitochondrial membrane 17b (TIM17b). In addition, host cell cytotoxicity was exacerbated in the absence of TIM17b during wild-type (WT) EPEC infection. The findings of this study together provide the first evidence that EspZ localizes to host mitochondria and that TIM17b contributes to protection against rapid cell death during EPEC infection.

scholarly journals Vibrio parahaemolyticusSenses Intracellular K+To Translocate Type III Secretion System 2 Effectors Effectively

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Sarunporn Tandhavanant ◽  
Shigeaki Matsuda ◽  
Hirotaka Hiyoshi ◽  
Tetsuya Iida ◽  
Toshio Kodama
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Cell Attachment ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Host Cells ◽  
Content Type ◽  
Type Iii ◽  
Key Factor ◽  
Effective Delivery

ABSTRACTMany Gram-negative bacterial symbionts and pathogens employ a type III secretion system (T3SS) to live in contact with eukaryotic cells. Because T3SSs inject bacterial proteins (effectors) directly into host cells, the switching of secretory substrates between translocators and effectors in response to host cell attachment is a crucial step for the effective delivery of effectors. Here, we show that the protein secretion switch ofVibrio parahaemolyticusT3SS2, which is a main contributor to the enteropathogenicity of a food poisoning bacterium, is regulated by two gatekeeper proteins, VgpA and VgpB. In the absence of these gatekeepers, effector secretion was activated, but translocator secretion was abolished, causing the loss of virulence. We found that the K+concentration, which is high inside the host cell but low outside, is a key factor for VgpA- and VgpB-mediated secretion switching. Exposure of wild-type bacteria to K+ions provoked both gatekeeper and effector secretions but reduced the level of secretion of translocators. The secretion protein profile of wild-type bacteria cultured with 0.1 M KCl was similar to that of gatekeeper mutants. Furthermore, depletion of K+ions in host cells diminished the efficiency of T3SS2 effector translocation. Thus, T3SS2 senses the high intracellular concentration of K+of the host cell so that T3SS2 effectors can be effectively injected.IMPORTANCEThe pathogenesis of many Gram-negative bacterial pathogens arises from a type III secretion system (T3SS), whereby bacterial proteins (effectors) are directly injected into host cells. The injected effectors then modify host cell functions. For effective delivery of effector proteins, bacteria need to both recognize host cell attachment and switch the type of secreted proteins. Here, we identified gatekeeper proteins that play important roles in a T3SS2 secretion switch ofVibrio parahaemolyticus, a causative agent of food-borne gastroenteritis. We also found that K+, which is present in high concentrations inside the host cell but in low concentrations outside, is a key factor for the secretion switch. Thus,V. parahaemolyticussenses the high intracellular K+concentration, triggering the effective injection of effectors.

scholarly journals Chlamydial Lytic Exit from Host Cells Is Plasmid Regulated

mBio ◽  
2015 ◽  
Vol 6 (6) ◽  
Author(s):  
Chunfu Yang ◽  
Tregei Starr ◽  
Lihua Song ◽  
John H. Carlson ◽  
Gail L. Sturdevant ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Actin Polymerization ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Cytoskeletal Proteins ◽  
Host Cells ◽  
Genetic Mechanism ◽  
Developmental Cycle ◽  
Obligate Intracellular ◽  
Type Iii

ABSTRACTChlamydia trachomatisis an obligate intracellular bacterium that is a globally important human pathogen. The chlamydial plasmid is an attenuating virulence factor, but the molecular basis for attenuation is not understood. Chlamydiae replicate within a membrane-bound vacuole termed an inclusion, where they undergo a biphasic developmental growth cycle and differentiate from noninfectious into infectious organisms. Late in the developmental cycle, the fragile chlamydia-laden inclusion retains its integrity by surrounding itself with scaffolds of host cytoskeletal proteins. The ability of chlamydiae to developmentally free themselves from this cytoskeleton network is a fundamental virulence trait of the pathogen. Here, we show that plasmidless chlamydiae are incapable of disrupting their cytoskeletal entrapment and remain intracellular as stable mature inclusions that support high numbers of infectious organisms. By using deletion mutants of the eight plasmid-carried genes (Δpgp1to Δpgp8), we show that Pgp4, a transcriptional regulator of multiple chromosomal genes, is required for exit. Exit of chlamydiae is dependent on protein synthesis and is inhibited by the compound C1, an inhibitor of the type III secretion system (T3S). Exit of plasmid-free and Δpgp4organisms, which failed to lyse infected cells, was rescued by latrunculin B, an inhibitor of actin polymerization. Our findings describe a genetic mechanism of chlamydial exit from host cells that is dependent on an unknownpgp4-regulated chromosomal T3S effector gene.IMPORTANCEChlamydia's obligate intracellular life style requires both entry into and exit from host cells. Virulence factors that function in exiting are unknown. The chlamydial inclusion is stabilized late in the infection cycle by F-actin. A prerequisite of chlamydial exit is its ability to disassemble actin from the inclusion. We show that chlamydial plasmid-free organisms, and also a plasmid gene protein 4 (pgp4) null mutant, do not disassociate actin from the inclusion and fail to exit cells. We further provide evidence that Pgp4-regulated exit is dependent on the chlamydial type III secretion system. This study is the first to define a genetic mechanism that functions in chlamydial lytic exit from host cells. The findings also have practical implications for understanding why plasmid-free chlamydiae are highly attenuated and have the ability to elicit robust protective immune responses.

scholarly journals Salmonella enterica subspecies enterica serovar Enteritidis Salmonella pathogenicity island 2 type III secretion system: role in intestinal colonization of chickens and systemic spread

Microbiology ◽  
2010 ◽  
Vol 156 (9) ◽  
pp. 2770-2781 ◽  
Author(s):  
Amanda L. S. Wisner ◽  
Taseen S. Desin ◽  
Birgit Koch ◽  
Po-King S. Lam ◽  
Emil M. Berberov ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Host Cells ◽  
Wild Type ◽  
Type Iii ◽  
Systemic Spread

Salmonella enterica subspecies enterica serovar Enteritidis (S. Enteritidis) has been identified as a significant cause of salmonellosis in humans. Salmonella pathogenicity islands 1 and 2 (SPI-1 and SPI-2) each encode a specialized type III secretion system (T3SS) that enables Salmonella to manipulate host cells at various stages of the invasion/infection process. For the purposes of our studies we used a chicken isolate of S. Enteritidis (Sal18). In one study, we orally co-challenged 35-day-old specific pathogen-free (SPF) chickens with two bacterial strains per group. The control group received two versions of the wild-type strain Sal18: Sal18 attTn7 : : tet and Sal18 attTn7 : : cat, while the other two groups received the wild-type strain (Sal18 attTn7 : : tet) and one of two mutant strains. From this study, we concluded that S. Enteritidis strains deficient in the SPI-1 and SPI-2 systems were outcompeted by the wild-type strain. In a second study, groups of SPF chickens were challenged at 1 week of age with four different strains: the wild-type strain, and three other strains lacking either one or both of the SPI-1 and SPI-2 regions. On days 1 and 2 post-challenge, we observed a reduced systemic spread of the SPI-2 mutants, but by day 3, the systemic distribution levels of the mutants matched that of the wild-type strain. Based on these two studies, we conclude that the S. Enteritidis SPI-2 T3SS facilitates invasion and systemic spread in chickens, although alternative mechanisms for these processes appear to exist.

Sign in / Sign up

Export Citation Format

Share Document