title: The type III protein translocation system of enteropathogenic Escherichia coli involves EspA-EspB protein interactions.
abstract: Enteropathogenic Escherichia coli (EPEC), like many bacterial pathogens, use a type III secretion system to deliver effector proteins across the bacterial cell wall. In EPEC, four proteins, EspA, EspB, EspD and Tir are known to be exported by a type III secretion system and to be essential for 'attaching and effacing' (A/E) lesion formation, the hallmark of EPEC pathogenicity. EspA was recently shown to be a structural protein and a major component of a large, transiently expressed, filamentous surface organelle which forms a direct link between the bacterium and the host cell. In contrast, EspB is translocated into the host cell where it is localized to both membrane and cytosolic cell fractions. EspA and EspB are required for translocation of Tir to the host cell membrane suggesting that they may both be components of the translocation apparatus. In this study, we show that EspB co-immunoprecipitates with the EspA filaments and that, during EPEC infection of HEp-2 cells, EspB localizes closely with EspA. Using a number of binding assays, we also show that EspB can bind and be copurified with EspA. Nevertheless, binding of EspA filaments to the host cell membranes occurred even in the absence of EspB. These results suggest that following initial attachment of the EspA filaments to the target cells, EspB is delivered into the host cell membrane and that the interaction between EspA and EspB may be important for protein translocation.
authors.affiliation: Department of Biochemistry, Imperial College of Science, Technology and Medicine, London SW7 2AZ, UK.
authors.forename: E L
authors.initials: EL
authors.lastname: Hartland
authors.forename: S J
authors.initials: SJ
authors.lastname: Daniell
authors.forename: R M
authors.initials: RM
authors.lastname: Delahay
authors.forename: B C
authors.initials: BC
authors.lastname: Neves
authors.forename: T
authors.initials: T
authors.lastname: Wallis
authors.forename: R K
authors.initials: RK
authors.lastname: Shaw
authors.forename: C
authors.initials: C
authors.lastname: Hale
authors.forename: S
authors.initials: S
authors.lastname: Knutton
authors.forename: G
authors.initials: G
authors.lastname: Frankel
mesh_terms: D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D001692:Biological Transport; D002460:Cell Line / Q000382:microbiology; D004847:Epithelial Cells / Q000382:microbiology; D004926:Escherichia coli / Q000378:metabolism* / Q000472:pathogenicity*; D029968:Escherichia coli Proteins; D005982:Glutathione Transferase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation; D011233:Precipitin Tests / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; C087863:EaeB protein, E coli; D029968:Escherichia coli Proteins; C102051:EspA protein, E coli; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase