title: Rickettsial outer-membrane protein B (rOmpB) mediates bacterial invasion through Ku70 in an actin, c-Cbl, clathrin and caveolin 2-dependent manner.
abstract: Rickettsia conorii, an obligate intracellular tick-borne pathogen and the causative agent of Mediterranean spotted fever, binds to and invades non-phagocytic mammalian cells. Previous work identified Ku70 as a mammalian receptor involved in the invasion process and identified the rickettsial autotransporter protein, rOmpB, as a ligand; however, little is known about the role of Ku70-rOmpB interactions in the bacterial invasion process. Using an Escherichia coli heterologous expression system, we show here that rOmpB mediates attachment to mammalian cells and entry in a Ku70-dependent process. A purified recombinant peptide corresponding to the rOmpB passenger domain interacts with Ku70 and serves as a competitive inhibitor of adherence. We observe that rOmpB-mediated infection culminates in actin recruitment at the bacterial foci, and that this entry process relies in part on actin polymerization likely imparted through protein tyrosine kinase and phosphoinositide 3-kinase-dependent activities and microtubule stability. Small-interfering RNA studies targeting components of the endocytic pathway reveal that entry by rOmpB is dependent on c-Cbl, clathrin and caveolin-2. Together, these results illustrate that rOmpB is sufficient to mediate Ku70-dependent invasion of mammalian cells and that clathrin- and caveolin-dependent endocytic events likely contribute to the internalization process.
authors.affiliation: Department of Microbiology, University of Chicago, 920 East 58th Street, Cummings Life Sciences Center 707A, Chicago, IL 60637, USA.
authors.forename: Yvonne G Y
authors.initials: YG
authors.lastname: Chan
authors.forename: Marissa M
authors.initials: MM
authors.lastname: Cardwell
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Hermanas
authors.forename: Tsuneo
authors.initials: T
authors.lastname: Uchiyama
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Martinez
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D034961:Antigens, Nuclear / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D051243:Caveolin 2 / Q000378:metabolism; D002522:Chlorocebus aethiops; D002966:Clathrin / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004847:Epithelial Cells / Q000382:microbiology*; D006367:HeLa Cells / Q000382:microbiology; D054884:Host-Pathogen Interactions; D006801:Humans; D000072200:Ku Autoantigen; D050721:Proto-Oncogene Proteins c-cbl / Q000378:metabolism; D020114:Rickettsia conorii / Q000378:metabolism / Q000472:pathogenicity* / Q000502:physiology; D014709:Vero Cells / Q000382:microbiology
chemical_list: D000199:Actins; D034961:Antigens, Nuclear; D001425:Bacterial Outer Membrane Proteins; D051243:Caveolin 2; D002966:Clathrin; D004268:DNA-Binding Proteins; D050721:Proto-Oncogene Proteins c-cbl; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen; C067419:CBL protein, human