title: The Serine Protease EspC from Enteropathogenic Escherichia coli Regulates Pore Formation and Cytotoxicity Mediated by the Type III Secretion System.
abstract: Type III secretion systems (T3SSs) are specialized macromolecular machines critical for bacterial virulence, and allowing the injection of bacterial effectors into host cells. The T3SS-dependent injection process requires the prior insertion of a protein complex, the translocon, into host cell membranes consisting of two-T3SS hydrophobic proteins, associated with pore-forming activity. In all described T3SS to date, a hydrophilic protein connects one hydrophobic component to the T3SS needle, presumably insuring the continuum between the hollow needle and the translocon. In the case of Enteropathogenic Escherichia coli (EPEC), the hydrophilic component EspA polymerizes into a filament connecting the T3SS needle to the translocon composed of the EspB and EspD hydrophobic proteins. Here, we identify EspA and EspD as targets of EspC, a serine protease autotransporter of Enterobacteriaceae (SPATE). We found that in vitro, EspC preferentially targets EspA associated with EspD, but was less efficient at proteolyzing EspA alone. Consistently, we found that EspC did not regulate EspA filaments at the surface of primed bacteria that was devoid of EspD, but controlled the levels of EspD and EspA secreted in vitro or upon cell contact. While still proficient for T3SS-mediated injection of bacterial effectors and cytoskeletal reorganization, an espC mutant showed increased levels of cell-associated EspA and EspD, as well as increased pore formation activity associated with cytotoxicity. EspP from enterohaemorrhagic E. coli (EHEC) also targeted translocator components and its activity was interchangeable with that of EspC, suggesting a common and important function of these SPATEs. These findings reveal a novel regulatory mechanism of T3SS-mediated pore formation and cytotoxicity control during EPEC/EHEC infection.
authors.affiliation: Equipe Communication Intercellulaire et Infections Microbiennes, Centre de Recherche Interdisciplinaire en Biologie (CIRB), Collège de France, Paris, France; Institut National de la Santé et de la Recherche Médicale U1050, Paris, France; Centre National de la Recherche Scientifique UMR7241, Paris, France; MEMOLIFE Laboratory of Excellence and Paris Science Lettre, Paris, France.
authors.forename: Julie
authors.initials: J
authors.lastname: Guignot
authors.affiliation: Equipe Communication Intercellulaire et Infections Microbiennes, Centre de Recherche Interdisciplinaire en Biologie (CIRB), Collège de France, Paris, France; Institut National de la Santé et de la Recherche Médicale U1050, Paris, France; Centre National de la Recherche Scientifique UMR7241, Paris, France; MEMOLIFE Laboratory of Excellence and Paris Science Lettre, Paris, France.
authors.forename: Audrey
authors.initials: A
authors.lastname: Segura
authors.affiliation: Equipe Communication Intercellulaire et Infections Microbiennes, Centre de Recherche Interdisciplinaire en Biologie (CIRB), Collège de France, Paris, France; Institut National de la Santé et de la Recherche Médicale U1050, Paris, France; Centre National de la Recherche Scientifique UMR7241, Paris, France; MEMOLIFE Laboratory of Excellence and Paris Science Lettre, Paris, France.
authors.forename: Guy
authors.initials: G
authors.lastname: Tran Van Nhieu
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D054308:Enteropathogenic Escherichia coli / Q000378:metabolism / Q000472:pathogenicity*; D004927:Escherichia coli Infections / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D016297:Mutagenesis, Site-Directed; D052899:Pore Forming Cytotoxic Proteins / Q000378:metabolism*; D000069296:Type III Secretion Systems / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C102541:EspC protein, E coli; D052899:Pore Forming Cytotoxic Proteins; D000069296:Type III Secretion Systems