Inhibition of NF-κB signaling in human dendritic cells by the enteropathogenic Escherichia coli effector protein NleE.
Intestinal dendritic cells (DCs) send processes between epithelial cells into the gut lumen to sample pathogens. Noninvasive enteropathogenic Escherichia coli (EPEC) colonize the gut using a type three secretion system (T3SS) to inject effector proteins into epithelial cells. We hypothesized that EPEC might also inject proteins into DC processes to dampen immune recognition. Using a T3SS-linked fluorescence resonance energy transfer-based system we show that EPEC injects effectors into in vitro grown human myeloid DCs. Injected cells emit a blue signal due to cleavage of the green fluorescence resonance energy transfer-based substrate CCF2/AM by β-lactamase. When cultured with a mutant EPEC unable to translocate effector proteins, myeloid DCs show rapid activation of NF-κB, secrete large amounts of proinflammatory cytokines and increase expression of CD80, CD83, and CD86, whereas wild-type EPEC barely elicits cytokine production and shuts off nuclear translocation of NF-κB p65. By deleting effector protein genes, we identified NleE as being critical for this effect. Expression of NleE in HeLa cells completely prevented nuclear p65 accumulation in response to IL1-β, and luciferase production in an NF-κB reporter cell line. DCs cocultured with wild-type EPEC or NleE-complemented strains were less potent at inducing MLR. EPEC was also able to inject effectors into DCs sending processes through model gut epithelium in a transwell system and into Peyer's patch myeloid DCs. Thus, EPEC translocate effectors into human DCs to dampen the inflammatory response elicited by its own pathogen-associated molecular patterns.
Vossenkämper
Anna
A
Institute of Cell and Molecular Science, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, Whitechapel, London. a.vossenkaemper@qmul.ac.uk
Marchès
Olivier
O
Fairclough
Peter D
PD
Warnes
Gary
G
Stagg
Andrew J
AJ
Lindsay
James O
JO
Evans
Paul C
PC
Luong
Le A
le A
Croft
Nicholas M
NM
Naik
Sandhia
S
Frankel
Gad
G
MacDonald
Thomas T
TT
D015153:Blotting, Western; D002469:Cell Separation; D018920:Coculture Techniques; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism / Q000382:microbiology*; D054308:Enteropathogenic Escherichia coli / Q000276:immunology / Q000378:metabolism / Q000472:pathogenicity*; D004797:Enzyme-Linked Immunosorbent Assay; D004927:Escherichia coli Infections / Q000276:immunology / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000276:immunology / Q000378:metabolism*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D007959:Lymphocyte Culture Test, Mixed; D018613:Microscopy, Confocal; D016328:NF-kappa B / Q000276:immunology / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology*; D037521:Virulence Factors / Q000276:immunology / Q000378:metabolism*
D029968:Escherichia coli Proteins; D016328:NF-kappa B; C526043:NleE protein, E coli; D037521:Virulence Factors