title: Vimentin-mediated signalling is required for IbeA+ E. coli K1 invasion of human brain microvascular endothelial cells.
abstract: IbeA in meningitic Escherichia coli K1 strains has been described previously for its role in invasion of BMECs (brain microvascular endothelial cells). Vimentin was identified as an IbeA-binding protein on the surface of HBMECs (human BMECs). In the present study, we demonstrated that vimentin is a primary receptor required for IbeA+ E. coli K1-induced signalling and invasion of HBMECs, on the basis of the following observations. First, E44 (IbeA+ E. coli K1 strain) invasion was blocked by vimentin inhibitors (withaferin A and acrylamide), a recombinant protein containing the vimentin head domain and an antibody against the head domain respectively. Secondly, overexpression of GFP (green fluorescent protein)-vimentin and GFP-VDM (vimentin head domain deletion mutant) significantly increased and decreased bacterial invasion respectively. Thirdly, bacterial invasion was positively correlated with phosphorylation of vimentin at Ser82 by CaMKII (Ca2+/calmodulin-dependent protein kinase II) and IbeA+ E. coli-induced phosphorylation of ERK (extracellular-signal-regulated kinase). Blockage of CaMKII by KN93 and inhibition of ERK1/2 phosphorylation by PD098059 resulted in reduced IbeA+ E. coli invasion. Fourthly, IbeA+ E. coli and IbeA-coated beads induced the clustering of vimentin that was correlated with increased entry of bacteria and beads. Lastly, IbeA+ E. coli K1 invasion was inhibited by lipid-raft-disrupting agents (filipin and nystatin) and caveolin-1 siRNA (small interfering RNA), suggesting that caveolae/lipid rafts are signalling platforms for inducing IbeA-vimentin-mediated E. coli invasion of HBMECs. Taken together, the present studies suggest that a dynamic and function-related interaction between IbeA and its primary receptor vimentin at HBMEC membrane rafts leads to vimentin phosphorylation and ERK-mediated signalling, which modulate meningitic E. coli K1 invasion.
authors.affiliation: Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, USA.
authors.forename: Feng
authors.initials: F
authors.lastname: Chi
authors.forename: Timothy D
authors.initials: TD
authors.lastname: Jong
authors.forename: Lin
authors.initials: L
authors.lastname: Wang
authors.forename: Yannan
authors.initials: Y
authors.lastname: Ouyang
authors.forename: Chunhua
authors.initials: C
authors.lastname: Wu
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Sheng-He
authors.initials: SH
authors.lastname: Huang
mesh_terms: D020106:Acrylamide / Q000494:pharmacology; D015153:Blotting, Western; D001921:Brain / Q000098:blood supply*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000235:genetics / Q000378:metabolism; D051242:Caveolin 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D004730:Endothelium, Vascular / Q000382:microbiology*; D004875:Ergosterol / Q000031:analogs & derivatives / Q000494:pharmacology; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D014746:Vimentin / Q000037:antagonists & inhibitors / Q000502:physiology*; D054358:Withanolides
chemical_list: D051242:Caveolin 1; C096350:CusC protein, E coli; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014746:Vimentin; D054358:Withanolides; D020106:Acrylamide; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D048049:Extracellular Signal-Regulated MAP Kinases; C009684:withaferin A; D004875:Ergosterol