Cytokeratin 18 interacts with the enteropathogenic Escherichia coli secreted protein F (EspF) and is redistributed after infection.
Enteropathogenic Escherichia coli (EPEC) pathogenesis requires the delivery of effector proteins into host cytosol by a type III secretion system. The effector protein EspF, while critical for disruption of epithelial barrier function through alteration of tight junctions, is not required for bacterial viability or attachment. Yeast two-hybrid analyses revealed host intermediate filament (IF) protein cytokeratin 18 (CK18) as an interacting partner of EspF. This was confirmed by co-immunoprecipitation of extracts from EPEC-infected epithelial cells. EPEC infection increased detergent-soluble CK18 amounts without significantly altering CK18 expression. The adaptor protein 14-3-3 binds to CK18 and modulates its solubility. EPEC infection promoted CK18/14-3-3 interactions, corresponding to the increase of CK18 in the soluble fractions. 14-3-3 also co-immunoprecipitated with EspF, suggesting that CK18, 14-3-3 and EspF may form a complex in infected cells. The 14-3-3zeta isoform was co-immunoprecipitated with CK18, suggesting the involvement of specific signalling pathways. Immunofluorescence studies revealed a dramatic alteration in the architecture of the IF network in EPEC-infected epithelial cells. IF fragmentation, evident at 2 h post infection, progressed to a collapse of this network at later time points. The secretion mutant (DeltaescN) failed to alter CK18 solubility and IF morphology, while deletion of espF partially impaired the ability of EPEC to induce CK18 modifications. These results suggest that modifications in 14-3-3 interactions and IF network, modulated by type III secreted proteins, may be an important step in EPEC pathogenesis.
Viswanathan
V K
VK
Department of Medicine, M/C 716, Section of Digestive Diseases and Nutrition, University of Illinois at Chicago, Room 718, Clinical Sciences Building, 840S. Wood Street, Chicago, IL 60612-7323, USA.
Lukic
Sandra
S
Koutsouris
Athanasia
A
Miao
Richard
R
Muza
Michelle M
MM
Hecht
Gail
G
D048948:14-3-3 Proteins / Q000378:metabolism; D002460:Cell Line; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D004926:Escherichia coli / Q000472:pathogenicity*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007382:Intermediate Filaments / Q000378:metabolism; D007633:Keratins / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D048948:14-3-3 Proteins; D029968:Escherichia coli Proteins; D007633:Keratins