title: EspC promotes epithelial cell detachment by enteropathogenic Escherichia coli via sequential cleavages of a cytoskeletal protein and then focal adhesion proteins.
abstract: EspC is a non-locus of enterocyte effacement (LEE)-encoded autotransporter produced by enteropathogenic Escherichia coli (EPEC) that is secreted to the extracellular milieu by a type V secretion system and then translocated into epithelial cells by the type III secretion system. Here, we show that this efficient EspC delivery into the cell leads to a cytopathic effect characterized by cell rounding and cell detachment. Thus, EspC is the main protein involved in epithelial cell cytotoxicity detected during EPEC adhesion and pedestal formation assays. The cell detachment phenotype is triggered by cytoskeletal and focal adhesion disruption. EspC-producing EPEC is able to cleave fodrin, paxillin, and focal adhesion kinase (FAK), but these effects are not observed when cells are infected with an espC isogenic mutant. Recovery of these phenotypes by complementing the mutant with the espC gene but not with the espC gene mutated in the serine protease motif highlights the role of the protease activity of EspC in the cell detachment phenotype. In vitro assays using purified proteins showed that EspC, but not EspC with an S256I substitution [EspCS256I], directly cleaves these cytoskeletal and focal adhesion proteins. Kinetics of protein degradation indicated that EspC-producing EPEC first cleaves fodrin (within the 11th and 9th repetitive units at the Q1219 and D938 residues, respectively), and this event sequentially triggers paxillin degradation, FAK dephosphorylation, and FAK degradation. Thus, cytoskeletal and focal adhesion protein cleavage leads to the cell rounding and cell detachment promoted by EspC-producing EPEC.
authors.affiliation: Department of Cell Biology, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Mexico City, Mexico.
authors.forename: Fernando
authors.initials: F
authors.lastname: Navarro-Garcia
authors.forename: Antonio
authors.initials: A
authors.lastname: Serapio-Palacios
authors.forename: Jorge E
authors.initials: JE
authors.lastname: Vidal
authors.forename: M Isabel
authors.initials: MI
authors.lastname: Salazar
authors.forename: Gabriela
authors.initials: G
authors.lastname: Tapia-Pastrana
mesh_terms: D001422:Bacterial Adhesion / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism*; D002448:Cell Adhesion; D002460:Cell Line; D054308:Enteropathogenic Escherichia coli / Q000472:pathogenicity*; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology; D004927:Escherichia coli Infections / Q000382:microbiology; D029968:Escherichia coli Proteins / Q000502:physiology*; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism*; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism*; D051419:Paxillin / Q000378:metabolism*; D013049:Spectrin
chemical_list: D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D051416:Focal Adhesion Protein-Tyrosine Kinases; D008840:Microfilament Proteins; D051419:Paxillin; D013049:Spectrin; C102541:EspC protein, E coli