Comparative Analysis of EspF from Enteropathogenic and Enterohemorrhagic Escherichia coli in Alteration of Epithelial Barrier Function

Abstract

Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) are related intestinal pathogens that harbor highly similar pathogenicity islands known as the locus of enterocyte effacement (LEE). Despite their genetic similarity, these two pathogens disrupt epithelial tight junction barrier function with distinct kinetics. EHEC-induced reduction in transepithelial electrical resistance (TER), a measure of barrier function disruption, is significantly slower and more modest in comparison to that induced by EPEC. The variation in bacterial adherence only partially accounted for these differences. The LEE-encoded effector protein EspF has been shown to be critical for EPEC-induced alterations in TER. EspF from both EPEC and EHEC is expressed and secreted upon growth in tissue culture medium. The mutation of EHEC cesF suggested that the optimal expression and secretion of EHEC EspF required its chaperone CesF, as has been shown for EPEC. In contrast to EPEC espF and cesF, mutation of the corresponding EHEC homologs did not dramatically alter the decrease in TER. These differences could possibly be explained by the presence of additional espF-like sequences (designated U-and M-espF, where the letter designations refer to the specific cryptic prophage sequences on the EHEC chromosome closest to the respective genes) in EHEC. Reverse transcription-PCR analyses revealed coordinate regulation of EHEC U-espF and the LEE-encoded espF, with enhanced expression in bacteria grown in Dulbecco-Vogt modified Eagle's medium compared to bacteria grown in Luria broth. Both EHEC espF and U-espF complemented an EPEC espF deletion strain for barrier function alteration. The overexpression of U-espF, but not espF, in wild-type EHEC potentiated the TER response. These studies reveal further similarities and differences in the pathogenesis of EPEC and EHEC.

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

“…Unfortunately, we did not screen for Abcf2 and N-WASP, but screened for the same ATP-binding cassette family of ABCA3. First, the espF gene sequences of EPEC and EHEC were 87% identical, although they had high similarity, their pathogenicity appeared to be different, for example, in reducing the function of epithelial resistance (TER) [69]. Second, the N-terminal of EspF protein in EPEC and EHEC includes the bacterial secretion signal site and the two organellar targeting domains.…”

Analysis of the expression, regulation and export of NleA–E in Escherichia coli O157 : H7

“…Using tandem MS, we observed many of the established proteins that have been shown to be secreted by EHEC O157. These included the Esp family of translocon proteins (EspA, B, D and F) (Abe et al, 1998;Viswanathan et al, 2004), the translocated intimin receptor, Tir (DeVinney et al, 1999), the mitochondrion-associated protein, Map (Kenny & Jepson, 2000), secreted proteases EspP (Brunder et al, 1997) and StcE (Lathem et al, 2002) (both encoded on pO157), and enterohaemolysin (Schmidt et al, 1995) (also on pO157). Using this wild-type strain, only NleA of the Nle A subpopulation of bacteria expressed nleA : : gfp, and only these bacteria were analysed for fluorescence output.…”

Mechanism underlying inhibition of intestinal apical Cl–/OH– exchange following infection with enteropathogenic E. coli

“…Therefore, it could be argued that the observed inhibition of apical Cl -/OH -exchange activity in response to EPEC infection could also be contributed by the DIDS-sensitive basolateral anion transporters. In this regard, our previous studies have suggested that infection of human intestinal epithelial cells with EPEC does not cause any transepithelial electrical resistance (TER) changes until 3 hours (5,39). Thus, in the absence of permeability changes, apical 36 Cl -uptake would be primarily facilitated by DRA and PAT-1.…”