Proteomic Analysis of the Intestinal Epithelial Cell Response to Enteropathogenic Escherichia coli

We present the first large scale proteomic analysis of a human cellular response to a pathogen. Enteropathogenic Escherichia coli (EPEC) is an enteric human pathogen responsible for much childhood morbidity and mortality worldwide. EPEC uses a type III secretion system (TTSS) to inject bacterial pro...

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Bibliographic Details
Published in:The Journal of biological chemistry 2004-05, Vol.279 (19), p.20127-20136
Main Authors: Hardwidge, Philip R, Rodriguez-Escudero, Isabel, Goode, David, Donohoe, Sam, Eng, Jimmy, Goodlett, David R, Aebersold, Reudi, Finlay, B Brett
Format: Article
Language:English
Subjects:
Biological Transport
Blotting, Western
Caco-2 Cells
Cytoskeleton - metabolism
Cytosol - metabolism
Down-Regulation
Epithelial Cells - microbiology
Escherichia coli
Escherichia coli - metabolism
Gene Expression Regulation
HeLa Cells
Humans
Immunoblotting
Intestines - microbiology
Ions
Microscopy, Fluorescence
Proteome
Spectrometry, Mass, Electrospray Ionization
Up-Regulation
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Summary:We present the first large scale proteomic analysis of a human cellular response to a pathogen. Enteropathogenic Escherichia coli (EPEC) is an enteric human pathogen responsible for much childhood morbidity and mortality worldwide. EPEC uses a type III secretion system (TTSS) to inject bacterial proteins into the cytosol of intestinal epithelial cells, resulting in diarrhea. We analyzed the host response to TTSS-delivered EPEC effector proteins by infecting polarized intestinal epithelial monolayers with either wild-type or TTSS-deficient EPEC. Host proteins were isolated and subjected to quantitative profiling using isotope-coded affinity tagging (ICAT) combined with electrospray ionization tandem mass spectrometry. We identified over 2000 unique proteins from infected Caco-2 monolayers, of which ∼13% are expressed differentially in the presence of TTSS-delivered EPEC effector proteins. We validated these data in silico and through immunoblotting and immunofluorescence microscopy. The identified changes extend cytoskeletal observations made in less relevant cell types and generate testable hypotheses with regard to host proteins potentially involved in EPEC-induced diarrhea. These data provide a framework for future biochemical analyses of host-pathogen interactions.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M401228200