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.
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Research Article|
March 15 2010
Vimentin-mediated signalling is required for IbeA+ E. coli K1 invasion of human brain microvascular endothelial cells
Feng Chi;
Feng Chi
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
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Timothy D. Jong;
Timothy D. Jong
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
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Lin Wang;
Lin Wang
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
†Department of Histology and Embryology, School of Basic Medical Science, Wuhan University, Wuhan 430071, China
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Yannan Ouyang;
Yannan Ouyang
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
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Chunhua Wu;
Chunhua Wu
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
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Wei Li;
Wei Li
‡Department of Medicine, Norris Cancer Center, University of Southern California, Los Angeles, CA 90027, U.S.A.
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Sheng-He Huang
Sheng-He Huang
1
*Saban Research Institute of Childrens Hospital Los Angeles, Department of Pediatrics, University of Southern California, Los Angeles, CA 90027, U.S.A.
1To whom correspondence should be addressed (email shhuang@hsc.usc.edu).
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Publisher: Portland Press Ltd
Received:
July 20 2009
Revision Received:
January 05 2010
Accepted:
January 20 2010
Accepted Manuscript online:
January 20 2010
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2010 Biochemical Society
2010
Biochem J (2010) 427 (1): 79–90.
Article history
Received:
July 20 2009
Revision Received:
January 05 2010
Accepted:
January 20 2010
Accepted Manuscript online:
January 20 2010
Citation
Feng Chi, Timothy D. Jong, Lin Wang, Yannan Ouyang, Chunhua Wu, Wei Li, Sheng-He Huang; Vimentin-mediated signalling is required for IbeA+ E. coli K1 invasion of human brain microvascular endothelial cells. Biochem J 1 April 2010; 427 (1): 79–90. doi: https://doi.org/10.1042/BJ20091097
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