title: Phosphatidylinositol 3-kinase activation and interaction with focal adhesion kinase in Escherichia coli K1 invasion of human brain microvascular endothelial cells.
abstract: 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 Deltap85 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.
authors.affiliation: Division of Infectious Diseases, Childrens Hospital Los Angeles, CA, USA.
authors.forename: M A
authors.initials: MA
authors.lastname: Reddy
authors.forename: N V
authors.initials: NV
authors.lastname: Prasadarao
authors.forename: C A
authors.initials: CA
authors.lastname: Wass
authors.forename: K S
authors.initials: KS
authors.lastname: Kim
mesh_terms: D001812:Blood-Brain Barrier; D001921:Brain / Q000382:microbiology*; D002478:Cells, Cultured; D002560:Cerebrovascular Circulation; D002867:Chromones / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000201:enzymology* / Q000382:microbiology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology / Q000472:pathogenicity*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D008833:Microcirculation; D009025:Morpholines / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction; D013329:Structure-Activity Relationship
chemical_list: D002867:Chromones; D004791:Enzyme Inhibitors; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011505:Protein-Tyrosine Kinases; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C496050:PTK2 protein, human