scholarly journals SseBCD Proteins Are Secreted by the Type III Secretion System of Salmonella Pathogenicity Island 2 and Function as a Translocon

2001 ◽  
Vol 183 (20) ◽  
pp. 6036-6045 ◽  
Author(s):  
Thomas Nikolaus ◽  
Jörg Deiwick ◽  
Catherine Rappl ◽  
Jeremy A. Freeman ◽  
Werner Schröder ◽  
...  
Keyword(s):  
Cell Surface ◽  
Type Iii Secretion ◽  
Bacterial Cell ◽  
Type Iii Secretion System ◽  
Pathogenicity Island ◽  
Secretion System ◽  
Effector Proteins ◽  
Type Iii ◽  
Bacterial Cell Surface ◽  
Substrate Proteins

ABSTRACT The type III secretion system encoded bySalmonella pathogenicity island 2 (SPI2) is required for systemic infections and intracellular accumulation of Salmonella enterica. This system is induced by intracellularSalmonella and subsequently transfers effector proteins into the host cell. Growth conditions either inducing expression of the type III secretion system or the secretion of substrate proteins were defined. Here we report the identification of a set of substrate proteins consisting of SseB, SseC, and SseD that are secreted by the SPI2 system in vitro. Secretion was observed if bacterial cells were exposed to acidic pH after growth in minimal medium with limitation of Mg2+ or phosphate. SseB, -C, and -D were isolated in a fraction detached from the bacterial cell surface by mechanical shearing, indicating that these proteins are predominantly assembled into complexes on the bacterial cell surface. The three proteins were required for the translocation of SPI2 effector proteins SspH1 and SspH2 into infected host cells. Thus, SseB, SseC, and SseD function as the translocon for effector proteins by intracellular Salmonella.

scholarly journals YscP and YscU Regulate Substrate Specificity of the Yersinia Type III Secretion System

2003 ◽  
Vol 185 (7) ◽  
pp. 2259-2266 ◽  
Author(s):  
Petra J. Edqvist ◽  
Jan Olsson ◽  
Moa Lavander ◽  
Lena Sundberg ◽  
Åke Forsberg ◽  
...  
Keyword(s):  
Cell Surface ◽  
Substrate Specificity ◽  
Type Iii Secretion ◽  
Bacterial Cell ◽  
Type Iii Secretion System ◽  
Eukaryotic Cell ◽  
Secretion System ◽  
Cell Contact ◽  
Type Iii ◽  
Bacterial Cell Surface

ABSTRACT Pathogenic Yersinia species use a type III secretion system to inhibit phagocytosis by eukaryotic cells. At 37°C, the secretion system is assembled, forming a needle-like structure on the bacterial cell surface. Upon eukaryotic cell contact, six effector proteins, called Yops, are translocated into the eukaryotic cell cytosol. Here, we show that a yscP mutant exports an increased amount of the needle component YscF to the bacterial cell surface but is unable to efficiently secrete effector Yops. Mutations in the cytoplasmic domain of the inner membrane protein YscU suppress the yscP phenotype by reducing the level of YscF secretion and increasing the level of Yop secretion. These results suggest that YscP and YscU coordinately regulate the substrate specificity of the Yersinia type III secretion system. Furthermore, we show that YscP and YscU act upstream of the cell contact sensor YopN as well as the inner gatekeeper LcrG in the pathway of substrate export regulation. These results further strengthen the strong evolutionary link between flagellar biosynthesis and type III synthesis.

SseA is a chaperone for the SseB and SseD translocon components of the Salmonella pathogenicity-island-2-encoded type III secretion system

Microbiology ◽  
2003 ◽  
Vol 149 (5) ◽  
pp. 1103-1111 ◽  
Author(s):  
Javier Ruiz-Albert ◽  
Rosanna Mundy ◽  
Xiu-Jun Yu ◽  
Carmen R. Beuzón ◽  
David W. Holden
Keyword(s):  
Type Iii Secretion ◽  
Sequence Similarity ◽  
Coiled Coil ◽  
Type Iii Secretion System ◽  
Pathogenicity Island ◽  
Secretion System ◽  
Effector Proteins ◽  
Type Iii ◽  
Virulence Attenuation

The type III secretion system (TTSS) encoded by the Salmonella pathogenicity island 2 (SPI-2) is required for bacterial replication inside macrophages and for systemic infection in mice. Many TTSS secreted proteins, including effectors and components of the translocon, require chaperones which promote their stability, prevent their premature interactions or facilitate their secretion. In this study, the function of the first gene (sseA) of one of the SPI-2 operons (sseA–G) was investigated. This operon includes genes that encode translocon components (SseB, SseC and SseD), translocated proteins (SseF and SseG) and putative chaperones (SscA and SscB). sseA encodes a 12·5 kDa protein with a C-terminal region with the potential to form a coiled-coil structure, but no sequence similarity to other proteins. Mutation of sseA results in severe virulence attenuation and an intracellular replication defect. It is shown here that SseA is not a secreted protein, but is required for SPI-2-dependent translocation of two effector proteins (SifA and PipB). Furthermore, the translocon components SseB and SseD were not detected in an sseA mutant strain. By using a yeast two-hybrid assay and column binding experiments, it is demonstrated that SseA interacts directly with SseB and SseD. These results indicate that SseA is a chaperone for SseB and SseD. The inability of an sseA mutant to assemble the SPI-2 TTSS translocon accounts for its high level of virulence attenuation in vivo. To the authors' knowledge, this is the first chaperone described for the SPI-2 TTSS.

scholarly journals Essential Role of the Type III Secretion System Effector NleB in Colonization of Mice by Citrobacter rodentium

2006 ◽  
Vol 74 (4) ◽  
pp. 2328-2337 ◽  
Author(s):  
Michelle Kelly ◽  
Emily Hart ◽  
Rosanna Mundy ◽  
Olivier Marchès ◽  
Siouxsie Wiles ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Pathogenicity Island ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Virulence Determinants ◽  
Citrobacter Rodentium ◽  
Gene Encoding ◽  
Type Iii

ABSTRACT Attaching and effacing (A/E) pathogens are a significant cause of gastrointestinal illness in humans and animals. All A/E pathogens carry a large pathogenicity island, termed the locus for enterocyte effacement (LEE), which encodes a type III secretion system that translocates several effector proteins into host cells. To identify novel virulence determinants in A/E pathogens, we performed a signature-tagged mutagenesis screen in C57BL/6 mice by using the mouse A/E pathogen Citrobacter rodentium. Five hundred seventy-six derivatives of C. rodentium were tested in pools of 12 mutants. One attenuated mutant carried a transposon insertion in nleB, which encodes a putative effector of the LEE-encoded type III secretion system (T3SS). nleB is present in a genomic pathogenicity island that also encodes another putative effector, NleE, immediately downstream. Using translational fusions with β-lactamase (TEM-1), we showed that both NleB and NleE were translocated into host cells by the LEE-encoded T3SS of enteropathogenic Escherichia coli. In addition, deletion of the gene encoding NleB in C. rodentium resulted in reduced colonization of mice in single infections and reduced colonic hyperplasia. In contrast, the deletion of other non-LEE-encoded effector genes in C. rodentium, nleC, nleD, or nleE, had no effect on host colonization or disease. These results suggest that nleB encodes an important virulence determinant of A/E pathogens.

scholarly journals RtsA Coordinately Regulates DsbA and the Salmonella Pathogenicity Island 1 Type III Secretion System

2004 ◽  
Vol 186 (1) ◽  
pp. 68-79 ◽  
Author(s):  
Craig D. Ellermeier ◽  
James M. Slauch
Keyword(s):  
Disulfide Bond ◽  
Type Iii Secretion ◽  
Feedback Inhibition ◽  
Regulatory Protein ◽  
Type Iii Secretion System ◽  
Pathogenicity Island ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Type Iii

ABSTRACT Salmonella serovars cause a wide variety of diseases ranging from mild gastroenteritis to life-threatening systemic infections. An important step in Salmonella enterica serovar Typhimurium infection is the invasion of nonphagocytic epithelial cells, mediated by a type III secretion system (TTSS) encoded on Salmonella pathogenicity island 1 (SPI1). The SPI1 TTSS forms a needle complex through which effector proteins are injected into the cytosol of host cells, where they promote actin rearrangement and engulfment of the bacteria. We previously identified the Salmonella-specific regulatory protein RtsA, which induces expression of hilA and, thus, the SPI1 genes. Here we show that the hilA regulators RtsA, HilD, and HilC can each induce transcription of dsbA, which encodes a periplasmic disulfide bond isomerase. RtsA induces expression of dsbA independent of either the SPI1 TTSS or the only known regulator of dsbA, the CpxRA two-component system. We show that DsbA is required for both the SPI1 and SPI2 TTSS to translocate effector proteins into the cytosol of host cells. DsbA is also required for survival during the systemic stages of infection. We also present evidence that production of SPI1 effector proteins is coupled to assembly of the TTSS. This feedback regulation is mediated at either the transcriptional or posttranscriptional level, depending on the particular effector. Loss of DsbA leads to feedback inhibition, which is consistent with the hypothesis that disulfide bond formation plays a role in TTSS assembly or function.

scholarly journals Pseudomonas syringae Strains Naturally Lacking the Classical P. syringae hrp/hrc Locus Are Common Leaf Colonizers Equipped with an Atypical Type III Secretion System

2010 ◽  
Vol 23 (2) ◽  
pp. 198-210 ◽  
Author(s):  
Christopher R. Clarke ◽  
Rongman Cai ◽  
David J. Studholme ◽  
David S. Guttman ◽  
Boris A. Vinatzer
Keyword(s):  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Plant Cells ◽  
Type Iii Secretion System ◽  
Ice Nucleation ◽  
Secretion System ◽  
Effector Proteins ◽  
Population Densities ◽  
Type Iii ◽  
Effector Genes

Pseudomonas syringae is best known as a plant pathogen that causes disease by translocating immune-suppressing effector proteins into plant cells through a type III secretion system (T3SS). However, P. syringae strains belonging to a newly described phylogenetic subgroup (group 2c) are missing the canonical P. syringae hrp/hrc cluster coding for a T3SS, flanking effector loci, and any close orthologue of known P. syringae effectors. Nonetheless, P. syringae group 2c strains are common leaf colonizers and grow on some tested plant species to population densities higher than those obtained by other P. syringae strains on nonhost species. Moreover, group 2c strains have genes necessary for the production of phytotoxins, have an ice nucleation gene, and, most interestingly, contain a novel hrp/hrc cluster, which is only distantly related to the canonical P. syringae hrp/hrc cluster. This hrp/hrc cluster appears to encode a functional T3SS although the genes hrpK and hrpS, present in the classical P. syringae hrp/hrc cluster, are missing. The genome sequence of a representative group 2c strain also revealed distant orthologues of the P. syringae effector genes avrE1 and hopM1 and the P. aeruginosa effector genes exoU and exoY. A putative life cycle for group 2c P. syringae is discussed.

scholarly journals Use of the Galleria mellonella Caterpillar as a Model Host To Study the Role of the Type III Secretion System in Pseudomonas aeruginosa Pathogenesis

2003 ◽  
Vol 71 (5) ◽  
pp. 2404-2413 ◽  
Author(s):  
Sachiko Miyata ◽  
Monika Casey ◽  
Dara W. Frank ◽  
Frederick M. Ausubel ◽  
Eliana Drenkard
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Galleria Mellonella ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Triple Mutant ◽  
Type Iii ◽  
Wax Moth

ABSTRACT Nonvertebrate model hosts represent valuable tools for the study of host-pathogen interactions because they facilitate the identification of bacterial virulence factors and allow the discovery of novel components involved in host innate immune responses. In this report, we determined that the greater wax moth caterpillar Galleria mellonella is a convenient nonmammalian model host for study of the role of the type III secretion system (TTSS) in Pseudomonas aeruginosa pathogenesis. Based on the observation that a mutation in the TTSS pscD gene of P. aeruginosa strain PA14 resulted in a highly attenuated virulence phenotype in G. mellonella, we examined the roles of the four known effector proteins of P. aeruginosa (ExoS, ExoT, ExoU, and ExoY) in wax moth killing. We determined that in P. aeruginosa strain PA14, only ExoT and ExoU play a significant role in G. mellonella killing. Strain PA14 lacks the coding sequence for the ExoS effector protein and does not seem to express ExoY. Moreover, using ΔexoU ΔexoY, ΔexoT ΔexoY, and ΔexoT ΔexoU double mutants, we determined that individual translocation of either ExoT or ExoU is sufficient to obtain nearly wild-type levels of G. mellonella killing. On the other hand, data obtained with a ΔexoT ΔexoU ΔexoY triple mutant and a ΔpscD mutant suggested that additional, as-yet-unidentified P. aeruginosa components of type III secretion are involved in virulence in G. mellonella. A high level of correlation between the results obtained in the G. mellonella model and the results of cytopathology assays performed with a mammalian tissue culture system validated the use of G. mellonella for the study of the P. aeruginosa TTSS.

scholarly journals RNase E Promotes Expression of Type III Secretion System Genes in Pseudomonas aeruginosa

2019 ◽  
Vol 201 (22) ◽  
Author(s):  
Josh S. Sharp ◽  
Arne Rietsch ◽  
Simon L. Dove
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Rnase E ◽  
Content Type ◽  
Type Iii ◽  
Small Regulatory Rnas

ABSTRACT Pseudomonas aeruginosa is an important opportunistic pathogen that employs a type III secretion system (T3SS) to inject effector proteins into host cells. Using a protein depletion system, we show that the endoribonuclease RNase E positively regulates expression of the T3SS genes. We also present evidence that RNase E antagonizes the expression of genes of the type VI secretion system and limits biofilm production in P. aeruginosa. Thus, RNase E, which is thought to be the principal endoribonuclease involved in the initiation of RNA degradation in P. aeruginosa, plays a key role in controlling the production of factors involved in both acute and chronic stages of infection. Although the posttranscriptional regulator RsmA is also known to positively regulate expression of the T3SS genes, we find that RNase E does not appreciably influence the abundance of RsmA in P. aeruginosa. Moreover, we show that RNase E still exerts its effects on T3SS gene expression in cells lacking all four of the key small regulatory RNAs that function by sequestering RsmA. IMPORTANCE The type III secretion system (T3SS) is a protein complex produced by many Gram-negative pathogens. It is capable of injecting effector proteins into host cells that can manipulate cell metabolism and have toxic effects. Understanding how the T3SS is regulated is important in understanding the pathogenesis of bacteria with such systems. Here, we show that RNase E, which is typically thought of as a global regulator of RNA stability, plays a role in regulating the T3SS in Pseudomonas aeruginosa. Depleting RNase E results in the loss of T3SS gene expression as well as a concomitant increase in biofilm formation. These observations are reminiscent of the phenotypes associated with the loss of activity of the posttranscriptional regulator RsmA. However, RNase E-mediated regulation of these systems does not involve changes in the abundance of RsmA and is independent of the known small regulatory RNAs that modulate RsmA activity.

scholarly journals Control of Type III Secretion System Effector/Chaperone Ratio Fosters Pathogen Adaptation to Host-Adherent Lifestyle

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Netanel Elbaz ◽  
Yaakov Socol ◽  
Naama Katsowich ◽  
Ilan Rosenshine
Keyword(s):  
Gene Expression ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Host Colonization ◽  
Content Type ◽  
Type Iii ◽  
Attached State

ABSTRACT The transition from a planktonic lifestyle to a host-attached state is often critical for bacterial virulence. Upon attachment to host cells, enteropathogenic Escherichia coli (EPEC) employs a type III secretion system (T3SS) to inject into the host cells ∼20 effector proteins, including Tir. CesT, which is encoded from the same operon downstream of tir, is a Tir-bound chaperone that facilitates Tir translocation. Upon Tir translocation, the liberated CesT remains in the bacterial cytoplasm and antagonizes the posttranscriptional regulator CsrA, thus eliciting global regulation in the infecting pathogen. Importantly, tight control of the Tir/CesT ratio is vital, since an uncontrolled surge in free CesT levels may repress CsrA in an untimely manner, thus abrogating colonization. We investigated how fluctuations in Tir translation affect the regulation of this ratio. By creating mutations that cause the premature termination of Tir translation, we revealed that the untranslated tir mRNA becomes highly unstable, resulting in a rapid drop in cesT mRNA levels and, thus, CesT levels. This mechanism couples Tir and CesT levels to ensure a stable Tir/CesT ratio. Our results expose an additional level of regulation that enhances the efficacy of the initial interaction of EPEC with the host cell, providing a better understanding of the bacterial switch from the planktonic to the cell-adherent lifestyle. IMPORTANCE Host colonization by extracellular pathogens often entails the transition from a planktonic lifestyle to a host-attached state. Enteropathogenic E. coli (EPEC), a Gram-negative pathogen, attaches to the intestinal epithelium of the host and employs a type III secretion system (T3SS) to inject effector proteins into the cytoplasm of infected cells. The most abundant effector protein injected is Tir, whose translocation is dependent on the Tir-bound chaperon CesT. Upon Tir injection, the liberated CesT binds to and inhibits the posttranscriptional regulator CsrA, resulting in reprogramming of gene expression in the host-attached bacteria. Thus, adaptation to the host-attached state involves dynamic remodeling of EPEC gene expression, which is mediated by the relative levels of Tir and CesT. Fluctuating from the optimal Tir/CesT ratio results in a decrease in EPEC virulence. Here we elucidate a posttranscriptional circuit that prevents sharp variations from this ratio, thus improving host colonization.

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