title: The Bacillus cereus Hbl and Nhe tripartite enterotoxin components assemble sequentially on the surface of target cells and are not interchangeable.
abstract: Bacillus cereus is a spore-forming, Gram-positive bacterium commonly associated with outbreaks of food poisoning. It is also known as an opportunistic pathogen causing clinical infections such as bacteremia, meningitis, pneumonia, and gas gangrene-like cutaneous infections, mostly in immunocompromised patients. B. cereus secretes a plethora of toxins of which four are associated with the symptoms of food poisoning. Two of these, the non-hemolytic enterotoxin Nhe and the hemolysin BL (Hbl) toxin, are predicted to be structurally similar and are unique in that they require the combined action of three toxin proteins to induce cell lysis. Despite their dominant role in disease, the molecular mechanism of their toxic function is still poorly understood. We report here that B. cereus strain ATCC 10876 harbors not only genes encoding Nhe, but also two copies of the hbl genes. We identified Hbl as the major secreted toxin responsible for inducing rapid cell lysis both in cultured cells and in an intraperitoneal mouse toxicity model. Antibody neutralization and deletion of Hbl-encoding genes resulted in significant reductions of cytotoxic activity. Microscopy studies with Chinese Hamster Ovary cells furthermore showed that pore formation by both Hbl and Nhe occurs through a stepwise, sequential binding of toxin components to the cell surface and to each other. This begins with binding of Hbl-B or NheC to the eukaryotic membrane, and is followed by the recruitment of Hbl-L1 or NheB, respectively, followed by the corresponding third protein. Lastly, toxin component complementation studies indicate that although Hbl and Nhe can be expressed simultaneously and are predicted to be structurally similar, they are incompatible and cannot complement each other.
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.
authors.forename: Inka
authors.initials: I
authors.lastname: Sastalla
authors.forename: Rasem
authors.initials: R
authors.lastname: Fattah
authors.forename: Nicole
authors.initials: N
authors.lastname: Coppage
authors.forename: Poulomi
authors.initials: P
authors.lastname: Nandy
authors.forename: Devorah
authors.initials: D
authors.lastname: Crown
authors.forename: Andrei P
authors.initials: AP
authors.lastname: Pomerantsev
authors.forename: Stephen H
authors.initials: SH
authors.lastname: Leppla
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001409:Bacillus cereus / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D016466:CHO Cells; D002460:Cell Line; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004768:Enterotoxins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D018628:Gene Dosage; D023061:Gene Order; D005816:Genetic Complementation Test; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism / Q000382:microbiology; D051379:Mice; D008807:Mice, Inbred BALB C; D008811:Mice, Inbred DBA; D018613:Microscopy, Confocal; D009154:Mutation; D011485:Protein Binding
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D017077:Culture Media, Conditioned; D004768:Enterotoxins; D006460:Hemolysin Proteins; C086257:hemolysin B