title: Escherichia coli interaction with human brain microvascular endothelial cells induces signal transducer and activator of transcription 3 association with the C-terminal domain of Ec-gp96, the outer membrane protein A receptor for invasion.
abstract: Our inability to develop new therapeutic strategies to prevent meningitis due to Escherichia coli K1 is attributed to our incomplete understanding of the pathophysiology of the disease. Previously, we demonstrated that outer membrane protein A of E. coli interacts with a gp96 homologue, Ec-gp96, on human brain microvascular endothelial cells (HBMEC) for invasion. However, signalling events mediated by Ec-gp96 that allow internalization of E. coli are incompletely understood. Here, we demonstrate that signal transducer and activator of transcription 3 (Stat3) activation and its interaction with Ec-gp96 were critical for E. coli invasion. The activated Stat3 was colocalized with Ec-gp96 at the actin condensation sites, and overexpressing a dominant negative (DN) form of Stat3 in HBMEC significantly abrogated the invasion. Furthermore, overexpression of Ec-gp96Delta200, the C-terminal 214-amino-acid truncated Ec-gp96, prevented the invasion of E. coli in HBMEC. In contrast, lack of ATP binding by gp96 did not affect the invasion. Overexpression of DN forms of either phosphatidyl inositol-3 kinase (PI3-kinase) subunit p85 or protein kinase C-alpha (PKC-alpha) had no effect on the activation of Stat3 and its association with Ec-gp96, whereas overexpression of DN-Stat3 abolished the activation of both PI3-kinase and PKC-alpha. Together, our findings identified a novel interaction of Stat3 with Ec-gp96, upstream of PI3-kinase and PKC-alpha activation that is required for the invasion of E. coli into HBMEC.
authors.affiliation: Division of Infectious Diseases, The Saban Research Institute, Childrens Hospital Los Angeles, CA 90027, USA.
authors.forename: Ravi
authors.initials: R
authors.lastname: Maruvada
authors.forename: Yair
authors.initials: Y
authors.lastname: Argon
authors.forename: Nemani V
authors.initials: NV
authors.lastname: Prasadarao
mesh_terms: D000199:Actins / Q000378:metabolism; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001921:Brain / Q000098:blood supply / Q000378:metabolism; D051242:Caveolin 1 / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008833:Microcirculation; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D001425:Bacterial Outer Membrane Proteins; D051242:Caveolin 1; D029968:Escherichia coli Proteins; D008562:Membrane Glycoproteins; D019869:Phosphatidylinositol 3-Kinases; D051571:Protein Kinase C-alpha; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C053387:endoplasmin; C078167:OMPA outer membrane proteins