title: Identification of cell surface-exposed proteins involved in the fimbria-mediated adherence of enteroaggregative Escherichia coli to intestinal cells.
abstract: Fimbria-mediated adherence to the intestinal epithelia is a key step in enteroaggregative Escherichia coli (EAEC) pathogenesis. To date, four fimbriae have been described for EAEC; aggregative adherence fimbria II (AAF/II) is the most important adherence factor for EAEC prototype strain 042. Previously, we described results showing that extracellular matrix (ECM) components might be involved in the recognition of AAF/II fimbriae by intestinal cells. In this study, we sought to identify novel potential receptors on intestinal epithelial cells recognized by the AAF/II fimbriae. Purified AafA-dsc protein, the major subunit of AAF/II fimbriae, was incubated with a monolayer of T84 cells, cross-linked to the surface-exposed T84 cell proteins, and immunoprecipitated by using anti-AafA antibodies. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of cellular proteins bound to AafA-dsc protein identified laminin (previously recognized as a potential receptor for AAF/II) and cytokeratin 8 (CK8). Involvement of the major subunit of AAF/II fimbriae (AafA protein) in the binding to recombinant CK8 was confirmed by adherence assays with purified AAF/II fimbriae, AafA-dsc protein, and strain 042. Moreover, HEp-2 cells transfected with CK8 small interfering RNA (siRNA) showed reduced 042 adherence compared with cells transfected with scrambled siRNA as a control. Adherence of 042 to HEp-2 cells preincubated with antibodies against ECM proteins or CK8 was substantially reduced. Altogether, our results supported the idea of a role of CK8 as a potential receptor for EAEC.
authors: Centro de Estudios Moleculares, Departamento de Pediatría, Hospital Dr. Luis Calvo Mackenna, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors: Mariana
authors: M
authors: Izquierdo
authors: Fernando
authors: F
authors: Navarro-Garcia
authors: Raul
authors: R
authors: Nava-Acosta
authors: James P
authors: JP
authors: Nataro
authors: Fernando
authors: F
authors: Ruiz-Perez
authors: Mauricio J
authors: MJ
authors: Farfan
mesh_terms: D018830:Adhesins, Escherichia coli; D001422:Bacterial Adhesion / Q000502:physiology*; D002460:Cell Line; D004847:Epithelial Cells / Q000382:microbiology* / Q000502:physiology; D004926:Escherichia coli / Q000502:physiology*; D005353:Fibronectins / Q000276:immunology; D010861:Fimbriae, Bacterial / Q000502:physiology*; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology; D053533:Keratin-8 / Q000378:metabolism / Q000502:physiology*; D007797:Laminin / Q000276:immunology / Q000502:physiology*; D008565:Membrane Proteins
chemical_list: C108511:AafA protein, E coli; D018830:Adhesins, Escherichia coli; D005353:Fibronectins; D053533:Keratin-8; D007797:Laminin; D008565:Membrane Proteins