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J. Biol. Chem., Vol. 275, Issue 47, 36769-36774, November 24, 2000
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From the § Division of Infectious Diseases, Childrens
Hospital Los Angeles and the Departments of ¶ Pediatrics and
Invasion of brain microvascular endothelial cells
(BMEC) is a prerequisite for successful crossing of the blood-brain
barrier by Escherichia coli K1. We have previously
demonstrated the requirement of cytoskeletal rearrangements and
activation of focal adhesion kinase (FAK) in E. coli K1
invasion of human BMEC (HBMEC). The current study investigated the role
of phosphatidylinositol 3-kinase (PI3K) activation and PI3K interaction
with FAK in E. coli invasion of HBMEC. PI3K inhibitor
LY294002 blocked E. coli K1 invasion of HBMEC in a
dose-dependent manner, whereas an inactive analogue LY303511 had no such effect. In HBMEC, E. coli K1 increased
phosphorylation of Akt, a downstream effector of PI3K, which was
completely blocked by LY294002. In contrast, non-invasive E. coli failed to activate PI3K. Overexpression of PI3K mutants
Phosphatidylinositol 3-Kinase Activation and Interaction with
Focal Adhesion Kinase in Escherichia coli K1 Invasion of
Human Brain Microvascular Endothelial Cells*
§,
**
Molecular Microbiology and Immunology, University of Southern
California School of Medicine, Los Angeles, California 90027
p85 and catalytically inactive p110 in HBMEC significantly inhibited
both PI3K/Akt activation and E. coli K1 invasion of HBMEC.
Stimulation of HBMEC with E. coli K1 increased PI3K
association with FAK. Furthermore, PI3K/Akt activation was blocked in
HBMEC-overexpressing FAK dominant-negative mutants (FRNK and
Phe397FAK). These results demonstrated the involvement of PI3K
signaling in E. coli K1 invasion of HBMEC and identified a
novel role for PI3K interaction with FAK in the pathogenesis of
E. coli meningitis.
*
This work was supported by National Institutes of Health
Grants R01-NS-26310, AI-47225, and HL-61951 (to K. S. K.) and
R29-AI-40567 (to N. V. P.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Current address: Division of Diabetes, Endocrinology and
Metabolism, Beckman Research Inst. City of Hope National Medical Center, Duarte, CA 91010.
**
To whom correspondence should be addressed: Pediatric Infectious
Diseases Division, Johns Hopkins University School of Medicine, 600 North Wolfe St., Park 256, Baltimore, MD 21287. Tel.:
410-614-3917; Fax: 410-614-1491; E-mail: kwangkim@jhmi.edu.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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