title: Bacterial surface association of heat-labile enterotoxin through lipopolysaccharide after secretion via the general secretory pathway.
abstract: Heat-labile enterotoxin (LT) is an important virulence factor expressed by enterotoxigenic Escherichia coli. The route of LT secretion through the outer membrane and the cellular and extracellular localization of secreted LT were examined. Using a fluorescently labeled receptor, LT was found to be specifically secreted onto the surface of wild type enterotoxigenic Escherichia coli. The main terminal branch of the general secretory pathway (GSP) was necessary and sufficient to localize LT to the bacterial surface in a K-12 strain. LT is a heteromeric toxin, and we determined that its cell surface localization was mediated by the its B subunit independent of an intact G(M1) ganglioside binding site and that LT binds lipopolysaccharide and G(M1) concurrently. The majority of LT secreted into the culture supernatant by the GSP in E. coli associated with vesicles. Only a mutation in hns, not overexpression of the GSP or LT, caused an increase in vesicle yield, supporting a specific vesicle formation machinery regulated by the nucleoid-associated protein HNS. We propose a model in which LT is secreted by the GSP across the outer membrane, secreted LT binds lipopolysaccharide via a G(M1)-independent binding region on its B subunit, and LT on the surface of released outer membrane vesicles interacts with host cell receptors, leading to intoxication. These data explain a novel mechanism of vesicle-mediated receptor-dependent delivery of a bacterial toxin into a host cell.
authors.affiliation: Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Amanda L
authors.initials: AL
authors.lastname: Horstman
authors.forename: Meta J
authors.initials: MJ
authors.lastname: Kuehn
mesh_terms: D001427:Bacterial Toxins / Q000378:metabolism*; D001896:Boron Compounds / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D004768:Enterotoxins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes / Q000494:pharmacology; D005677:G(M1) Ganglioside / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism*; D008954:Models, Biological; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001427:Bacterial Toxins; D001896:Boron Compounds; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes; D008070:Lipopolysaccharides; D005677:G(M1) Ganglioside; C027538:heat-labile enterotoxin, E coli