title: The pore-forming toxin listeriolysin O mediates a novel entry pathway of L. monocytogenes into human hepatocytes.
abstract: Intracellular pathogens have evolved diverse strategies to invade and survive within host cells. Among the most studied facultative intracellular pathogens, Listeria monocytogenes is known to express two invasins-InlA and InlB-that induce bacterial internalization into nonphagocytic cells. The pore-forming toxin listeriolysin O (LLO) facilitates bacterial escape from the internalization vesicle into the cytoplasm, where bacteria divide and undergo cell-to-cell spreading via actin-based motility. In the present study we demonstrate that in addition to InlA and InlB, LLO is required for efficient internalization of L. monocytogenes into human hepatocytes (HepG2). Surprisingly, LLO is an invasion factor sufficient to induce the internalization of noninvasive Listeria innocua or polystyrene beads into host cells in a dose-dependent fashion and at the concentrations produced by L. monocytogenes. To elucidate the mechanisms underlying LLO-induced bacterial entry, we constructed novel LLO derivatives locked at different stages of the toxin assembly on host membranes. We found that LLO-induced bacterial or bead entry only occurs upon LLO pore formation. Scanning electron and fluorescence microscopy studies show that LLO-coated beads stimulate the formation of membrane extensions that ingest the beads into an early endosomal compartment. This LLO-induced internalization pathway is dynamin-and F-actin-dependent, and clathrin-independent. Interestingly, further linking pore formation to bacteria/bead uptake, LLO induces F-actin polymerization in a tyrosine kinase-and pore-dependent fashion. In conclusion, we demonstrate for the first time that a bacterial pathogen perforates the host cell plasma membrane as a strategy to activate the endocytic machinery and gain entry into the host cell.
authors.affiliation: Departments of Microbiology and Internal Medicine, Center for Microbial Interface Biology, Ohio State University, Columbus, Ohio, United States of America.
authors.forename: Stephen
authors.initials: S
authors.lastname: Vadia
authors.forename: Eusondia
authors.initials: E
authors.lastname: Arnett
authors.forename: Anne-Cécile
authors.initials: AC
authors.lastname: Haghighat
authors.forename: Elisabeth M
authors.initials: EM
authors.lastname: Wilson-Kubalek
authors.forename: Rodney K
authors.initials: RK
authors.lastname: Tweten
authors.forename: Stephanie
authors.initials: S
authors.lastname: Seveau
mesh_terms: D000199:Actins / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism / Q000382:microbiology; D002966:Clathrin / Q000378:metabolism; D003603:Cytotoxins / Q000378:metabolism; D034281:Dynamins / Q000378:metabolism; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000378:metabolism*; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000382:microbiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D008089:Listeria monocytogenes / Q000472:pathogenicity* / Q000502:physiology; D008863:Microspheres; D011137:Polystyrenes; D052899:Pore Forming Cytotoxic Proteins / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism
chemical_list: D000199:Actins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D002966:Clathrin; D003603:Cytotoxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; D011137:Polystyrenes; D052899:Pore Forming Cytotoxic Proteins; D011505:Protein-Tyrosine Kinases; D034281:Dynamins; C038705:hlyA protein, Listeria monocytogenes
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