Enteropathogenic Escherichia coli infection leads to appearance of aberrant tight junctions strands in the lateral membrane of intestinal epithelial cells

Abstract

SummaryInfection of intestinal epithelial cells with enteropathogenic Escherichia coli (EPEC) disrupts tight junction (TJ) architecture and barrier function. The aim of this study was to determine the impact of EPEC on TJ protein interactions and localization. Human intestinal epithelial cells (T84) were infected for 1, 3 or 6 h with EPEC. To probe the TJ protein-protein interactions, co-immunoprecipitations were performed. The associations between ZO-1, occludin and claudin-1 progressively decreased after infection. Corresponding morphological changes were analysed by immunofluorescence confocal microscopy. Tight junction proteins progressively lost their apically restricted localization. Freeze-fracture electron microscopy revealed the appearance of aberrant strands throughout the lateral membrane that contained claudin-1 and occludin as determined by immunogold labelling. These structural alterations were accompanied by a loss of barrier function. Mutation of the gene encoding EspF, important in the disruption of TJs by EPEC, prevented the disruption of TJs. Tight junction structure normalized following eradication of EPEC with gentamicin and overnight recovery. This is the first demonstration that a microbial pathogen can cause aberrant TJ strands in the lateral membrane of host cells. We speculate that the disruption of integral and cytoplasmic TJ protein interactions following EPEC infection allows TJ strands to form or diffuse into the lateral plasma membrane.

Enteropathogenic Escherichia coli , Shigella flexneri , and Listeria monocytogenes Recruit a Junctional Protein, Zonula Occludens-1, to Actin Tails and Pedestals

“…Upon infection by EPEC, the peripheral lining of ZO-1 began to disperse at 3 h postinfection and became discontinuous at 5 h or later. These changes are basically consistent with the observations made with T84 monolayers (10,37,43,48). In addition to these changes, we noticed the recruitment of ZO-1 to the sites of bacterial attachment (Fig.…”

“…Amieva and colleagues have reported that Helicobacter pylori causes a similar recruitment of ZO-1 to the bacterial attachment sites in Madin-Darby canine kidney (MDCK) cell monolayers and proposed that the recruitment is linked to a dysfunctional paracellular seal (1). Because EPEC has been reported to affect the integrity of the epithelial barrier (11,32,34,36,37,43,54), we examined if the EPEC-induced recruitment of ZO-1 is also linked with the disruption of the barrier. A polarized Caco/B7 monolayer was infected with wild-type EPEC and the mutants, and the TER across the monolayer, which is a hallmark of the integrity of the epithelial barrier, was measured.…”

Screening for Host Proteins Interacting with Escherichia Coli O157:H7 EspF using Bimolecular Fluorescence Complementation

“…Moreover, VK Viswanathan et al have shown that EspF can interact with the host intermediate filament protein cytokeratin 18 (CK18) in a complex with adaptor protein 14-3-3, which can alter the architecture of the intermediate filament network in EPEC-infected cells [19]. It cannot only modulate the structure and function of TJs but can also remodel the brush border [20,21]. In addition, EspF can inhibit EPEC internalization by J774.A1 macrophages through a PI-3 kinase-dependent pathway [22].…”

The role of epithelial malfunction in the pathogenesis of enteropathogenic E. coli-induced diarrhea

“…Indeed, EPEC infection induces redistribution of occludin, which results in a significant decrease in barrier function both in vitro and in vivo, in an EspF-dependent manner. [40][41][42][43] Redistribution of ZO-1, cytoplasmic clearing of claudin-1, and a gradual decrease in protein-protein interaction within the tight junctions have also been reported during EPEC infection. 41 Finally, EspF and Map have been shown to induce cytosolic translocation of claudin-1, -3, and -5 in C. rodentium-infected mice.…”

“…[40][41][42][43] Redistribution of ZO-1, cytoplasmic clearing of claudin-1, and a gradual decrease in protein-protein interaction within the tight junctions have also been reported during EPEC infection. 41 Finally, EspF and Map have been shown to induce cytosolic translocation of claudin-1, -3, and -5 in C. rodentium-infected mice. 44,45 This mechanism requires the locus of enterocyte effacementencoded EspF chaperone, CesF, for the translocation of EspF into the host cytosol and the subsequent reduction of TER.…”