Disruption of cell polarity by enteropathogenic Escherichia coli enables basolateral membrane proteins to migrate apically and to potentiate physiological consequences.
Enteropathogenic Escherichia coli (EPEC) disrupts the structure and barrier function of host intestinal epithelial tight junctions (TJs). The impact of EPEC on TJ "fence function," i.e., maintenance of cell polarity, has not been investigated. In polarized cells, proteins such as beta(1)-integrin and Na(+)/K(+) ATPase are restricted to basolateral (BL) membranes. The outer membrane EPEC protein intimin possesses binding sites for the EPEC translocated intimin receptor (Tir) and beta(1)-integrin. Restriction of beta(1)-integrin to BL domains, however, precludes opportunity for interaction. We hypothesize that EPEC perturbs TJ fence function and frees BL proteins such as beta(1)-integrin to migrate to apical (AP) membranes of host cells, thus allowing interactions with bacterial adhesins such as intimin. The aim of this study was to determine whether EPEC alters the polar distribution of BL proteins, in particular beta(1)-integrin, and if such redistribution contributes to pathogenesis. Human intestinal epithelial T84 cells and EPEC strain E2348/69 were used. Selective biotinylation of AP or BL membrane proteins and confocal microscopy showed the presence of beta(1)-integrin and Na(+)/K(+) ATPase on the AP membrane following infection. beta(1)-Integrin antibody afforded no protection against the initial EPEC-induced decrease in transepithelial electrical resistance (TER) but halted the progressive decrease at later time points. While the effects of EPEC on TJ barrier and fence function were Tir dependent, disruption of cell polarity by calcium chelation allowed a tir mutant to be nearly as effective as wild-type EPEC. In contrast, deletion of espD, which renders the type III secretory system ineffective, had no effect on TER even after calcium chelation, suggesting that the putative beta(1)-integrin-intimin interaction serves to provide intimate contact, like that of Tir and intimin, making translocation of effector molecules more efficient. We conclude that the initial alterations of TJ barrier and fence function by EPEC are Tir dependent but that later disruption of cell polarity and accessibility of EPEC to BL membrane proteins, such as beta(1)-integrin, potentiates the physiological perturbations.
Muza-Moons
Michelle M
MM
Section of Digestive Diseases and Nutrition, Department of Medicine, University of Illinois at Chicago, 60612, USA.
Koutsouris
Athanasia
A
Hecht
Gail
G
D002462:Cell Membrane / Q000473:pathology* / Q000502:physiology; D016764:Cell Polarity / Q000502:physiology; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000382:microbiology / Q000473:pathology; D004926:Escherichia coli / Q000472:pathogenicity*; D029968:Escherichia coli Proteins; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D007422:Intestines / Q000166:cytology; D008565:Membrane Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D011956:Receptors, Cell Surface; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D019108:Tight Junctions / Q000473:pathology*
D029968:Escherichia coli Proteins; D019012:Integrin beta1; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; C109287:Tir protein, E coli; D000254:Sodium-Potassium-Exchanging ATPase