Initial binding of Shiga toxin‐producing <i>Escherichia coli</i> to host cells and subsequent induction of actin rearrangements depend on filamentous EspA‐containing surface appendages

Ebel, Frank; Podzadel, Till; Rohde, Manfred; Kresse, Andreas U.; Krämer, Sarah; Deibel, Christina; Guzmán, Carlos A.; Chakraborty, Trinad

doi:10.1046/j.1365-2958.1998.01046.x

Cited by 157 publications

(166 citation statements)

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“…The enteropathogenic E. coli espA mutant is defective in the transport of effector proteins into the host cell but is normal in the secretion of effector proteins across the bacterial cell wall (13,14). In this regard, class IB mutants pheno-copy the espA mutant.…”

Section: Toward Understanding the Molecular Defects Of Nonfunctional mentioning

confidence: 99%

“…In enteropathogenic Escherichia coli, the needle is connected with another extracellular filament called the EspA filament (11,12). The EspA filament is 12 nm in diameter and can be several micrometers long (13,14). The TTSS of plant pathogenic bacteria assembles an extracellular appendage called the Hrp pilus, which is 6 -8 nm wide and several micrometers long (15)(16)(17)(18)(19)(20).…”

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confidence: 99%

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Use of Dominant-negative HrpA Mutants to Dissect Hrp Pilus Assembly and Type III Secretion in Pseudomonas syringae pv. tomato

Lee

Kolade²,

Nomura³

et al. 2005

Journal of Biological Chemistry

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The Hrp pilus plays an essential role in the long-distance type III translocation of effector proteins from bacteria into plant cells. HrpA is the structural subunit of the Hrp pilus in Pseudomonas syringae pv. tomato (Pst) DC3000. Little is known about the molecular features in the HrpA protein for pilus assembly or for transporting effector proteins. From previous collections of nonfunctional HrpA derivatives that carry random pentapeptide insertions or single amino acid mutations, we identified several dominant-negative mutants that blocked the ability of wild-type Pst DC3000 to elicit host responses. The dominant-negative phenotype was correlated with the disappearance of the Hrp pilus in culture and inhibition of wild-type HrpA protein self-assembly in vitro. Dominant-negative HrpA mutants can be grouped into two functional classes: one class exerted a strong dominant-negative effect on the secretion of effector proteins AvrPto and HopPtoM in culture, and the other did not. The two classes of mutant HrpA proteins carry pentapeptide insertions in discrete regions, which are interrupted by insertions without a dominant-negative effect. These results enable prediction of possible subunit-subunit interaction sites in the assembly of the Hrp pilus and suggest the usefulness of dominant-negative mutants in dissection of the role of the wild-type HrpA protein in various stages of type III translocation: protein exit across the bacterial cell wall, the assembly and/or stabilization of the Hrp pilus in the extracellular space, and Hrp pilus-mediated long-distance transport beyond the bacterial cell wall. The bacterial type III secretion system (TTSS)1 is a longdistance protein transport system, moving bacterial effector proteins from the bacterial cytoplasm into a eukaryotic cell (1-4). During this long-distance transport, several physical barriers must be traversed: the bacterial cell wall, the host extracellular matrix layer (e.g. plant cell wall or animal mucous layer/glycocalyx), and the host plasma membrane. The TTSS has adapted to such long-distance transport by assembling extracellular needle/pilus-like appendages of various lengths (3, 4). For example, the TTSS of mammalian pathogenic bacteria assembles a needle complex, which consists of a base structure embedded in the bacterial cell wall and a primarily extracellular hollow needle of 6 -8 nm in diameter and 50 -80 nm in length (5-10). In enteropathogenic Escherichia coli, the needle is connected with another extracellular filament called the EspA filament (11,12). The EspA filament is 12 nm in diameter and can be several micrometers long (13,14). The TTSS of plant pathogenic bacteria assembles an extracellular appendage called the Hrp pilus, which is 6 -8 nm wide and several micrometers long (15-20). The needle, EspA filament, and Hrp pilus are believed to be tunnels linking the type III "secreton" embedded in the bacterial cell wall and a type III "translocon" in the host plasma membrane. The secreton allows the exit of effector proteins across the bacte...

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Section: Toward Understanding the Molecular Defects Of Nonfunctional mentioning

confidence: 99%

mentioning

confidence: 99%

Use of Dominant-negative HrpA Mutants to Dissect Hrp Pilus Assembly and Type III Secretion in Pseudomonas syringae pv. tomato

Lee

Kolade²,

Nomura³

et al. 2005

Journal of Biological Chemistry

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“…EspA is secreted by the type III secretion system across the bacterial membranes and is found in ®lamentous appendages on the bacterial surface . The same organelle is also present in Shiga toxin-producing E. coli (STEC) (Ebel et al, 1998a), which also possesses a LEE. This structure could form a transport channel, connecting the type III secretion apparatus in the bacterial envelope and the host cell membrane ( Fig.…”

Section: Bacterial Factors Involved In Epec Attachment To Host Cellsmentioning

confidence: 99%

Enteropathogenic Escherichia coli (EPEC) attachment to epithelial cells: exploiting the host cell cytoskeleton from the outside

2000

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SummaryEnteropathogenic Escherichia coli (EPEC), a leading cause of human infantile diarrhoea, is the prototype for a family of intestinal bacterial pathogens that induce attaching and effacing (A/E) lesions on host cells. A/E lesions are characterized by localized effacement of the brush border of enterocytes, intimate bacterial attachment and pedestal formation beneath the adherent bacteria. As a result of some recent breakthrough discoveries, EPEC has now emerged as a fascinating paradigm for the study of host±pathogen interactions and cytoskeletal rearrangements that occur at the host cell membrane. EPEC uses a type III secretion machinery to attach to epithelial cells, translocating its own receptor for intimate attachment, Tir, into the host cell, which then binds to intimin on the bacterial surface. Studies of EPECinduced cytoskeletal rearrangements have begun to provide clues as to the mechanisms used by this pathogen to subvert the host cell cytoskeleton and signalling pathways. These ®ndings have unravelled new ways by which pathogenic bacteria exploit host processes from the cell surface and have shed new light on how EPEC might cause diarrhoea. EPEC, a model extracellular pathogenAmong the many themes in bacterial pathogenesis, determining how bacteria interact with host cells, including subversion of the cellular cytoskeleton and signal transduction pathways, remains of major interest. Most of the pathogens for which these aspects have been extensively studied, such as Salmonella, Shigella and Listeria species, illustrate that pathogens have evolved strategies to hijack the host cell machinery for invasive purposes. They use the intestinal barrier as an entry site, exploiting the target host cell cytoskeletal machinery to trigger their uptake by non-phagocytic cells, thereby gaining access to the host. Enteropathogenic Escherichia coli (EPEC), a major cause of diarrhoea in the developing world, colonizes the intestinal epithelial surface and exerts its pathologic effects by intimately attaching to the surface of enterocytes, causing histopathological alterations called the attaching/effacing (A/E) lesion. A/E lesions result from degeneration of the enterocyte brush border through the reorganization of the underlying cytoskeleton. This is triggered by bacteria attached to the surface and leads to the localized loss of microvilli and the formation of actin-rich pedestals upon which EPEC reside (Frankel et al., 1998a). Several other pathogens, such as enterohaemorrhagic E. coli (EHEC), which is associated with haemolytic±uraemic syndrome, Citrobacter rodentium and Hafnia alvei, also induce A/E lesion formation upon infection, but EPEC has been the most well studied of these pathogens and thus serves as the paradigm.Like several Gram-negative animal and plant pathogens, EPEC possesses a type III secretion machinery, which is designed to secrete key virulence factors and directly inject some of them into the host cell cytosol by functioning as a`molecular syringe ' (Hueck, 1998). Recently, some new and s...

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“…Translocation of Tir and other effector proteins into host cells requires the type III secreted EspA protein, which forms filaments connecting the bacteria to the host cell surface, as well as EspB and EspD, which are thought to form a membrane pore (reviewed by Frankel et al, 1998). EspA filaments are believed to act as a hollow conduit for the delivery of effectors; however, they also play a role in initial adherence (Cleary et al, 2004;Ebel et al, 1998), and an E. coli O157 : H7 non-polar espA deletion mutant exhibits reduced colonization in a murine model (Nagano et al, 2003). Immunization of cattle with E. coli O157 : H7 type III secreted proteins reduces faecal shedding of the bacteria following experimental infection (Potter et al, 2004); however, the contribution of the structural apparatus and individual type III secreted proteins in colonization and immunity in ruminants is unknown.…”

Section: Introductionmentioning

confidence: 99%

Identification of Escherichia coli O157 : H7 genes influencing colonization of the bovine gastrointestinal tract using signature-tagged mutagenesis

et al. 2004

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Enterohaemorrhagic Escherichia coli (EHEC) cause acute gastroenteritis in humans that may be complicated by life-threatening systemic sequelae. The predominant EHEC serotype affecting humans in the UK and North America is O157 : H7 and infections are frequently associated with contact with ruminant faeces. Strategies to reduce the carriage of EHEC in ruminants are expected to lower the incidence of human EHEC infections; however, the molecular mechanisms underlying persistence of EHEC in ruminants are poorly understood. This paper reports the first comprehensive survey for EHEC factors mediating colonization of the bovine intestines by using signature-tagged transposon mutagenesis. Seventy-nine E. coli O157 : H7 mutants impaired in their ability to colonize calves were isolated and 59 different genes required for intestinal colonization were identified by cloning and sequencing of the transposon insertion sites. Thirteen transposon insertions were clustered in the locus of enterocyte effacement (LEE), which encodes a type III protein secretion system required for the formation of attaching and effacing lesions on intestinal epithelia. A putative structural component of the apparatus (EscN) is essential for intestinal colonization; however, the type III secreted effector protein Map plays only a minor role. Other Type III secretion-associated genes were implicated in colonization of calves by E. coli O157 : H7, including z0990 (ecs0850), which encodes the non-LEE-encoded type III secreted effector NleD and the closely related z3023 (ecs2672) and z3026 (ecs2674) genes which encode homologues of Shigella IpaH proteins. We also identified a novel fimbrial locus required for intestinal colonization in calves by E. coli O157 : H7 (z2199-z2206; ecs2114-ecs2107/locus 8) and demonstrated that a mutant harbouring a deletion of the putative major fimbrial subunit gene is rapidly out-competed by the parent strain in co-infection studies. Our data provide valuable new information for the development of intervention strategies. INTRODUCTIONEnterohaemorrhagic Escherichia coli (EHEC) were first recognized as a cause of human disease in 1983 and are associated with diarrhoea and haemorrhagic colitis, which may be complicated by life-threatening renal and neurological sequelae, including haemolytic uraemic syndrome and thrombocytopaenic purpura (Karmali et al., 1983;Riley et al., 1983). EHEC, and in particular serotype O157 : H7, have since emerged as a major cause of severe diarrhoeal illness worldwide and EHEC infections are the leading antecedent to paediatric acute renal failure in many countries (Ammon, 1997; Mead et al., 1999; WHO, 1997). E. coli O157 : H7 strains are estimated to cause 73 480 cases and 60 deaths per annum in the United States, with serotypes other than O157 : H7 accounting for a further 36 740 illnesses (Mead et al., 1999). Non-O157 EHEC are an emerging problem, and indeed may be more prevalent than serogroup O157 in some countries (Bettelheim, 2000). EHEC are defined by their ability to produce one or mo...

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Initial binding of Shiga toxin‐producing Escherichia coli to host cells and subsequent induction of actin rearrangements depend on filamentous EspA‐containing surface appendages

Cited by 157 publications

References 32 publications

Use of Dominant-negative HrpA Mutants to Dissect Hrp Pilus Assembly and Type III Secretion in Pseudomonas syringae pv. tomato

Use of Dominant-negative HrpA Mutants to Dissect Hrp Pilus Assembly and Type III Secretion in Pseudomonas syringae pv. tomato

Enteropathogenic Escherichia coli (EPEC) attachment to epithelial cells: exploiting the host cell cytoskeleton from the outside

Identification of Escherichia coli O157 : H7 genes influencing colonization of the bovine gastrointestinal tract using signature-tagged mutagenesis

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