The Serine Protease Motif of EspC from Enteropathogenic Escherichia coli Produces Epithelial Damage by a Mechanism Different from That of Pet Toxin from Enteroaggregative E. coli

Abstract

EspC (Escherichia coli secreted protein C) of enteropathogenic E. coli (EPEC) shows the three classical domains of the autotransporter proteins and has a conserved serine protease motif belonging to the SPATE (serine protease autotransporters of Enterobacteriaceae) subfamily. EspC and its homolog Pet in enteroaggregative E. coli (EAEC) bear the same sequence within the serine protease motif, and both proteins produce enterotoxic effects, suggesting that like Pet, EspC could be internalized to reach and cleave the calmodulinbinding domain of fodrin, causing actin cytoskeleton disruption. Even though both proteins cause cytoskeleton damage by virtue of their serine protease motifs, the following evidence supports the hypothesis that the mechanisms are different. All these data suggest that the protein conformational structure is very important for the activity of the catalytic site, influencing its interaction with the target protein and its internalization. The differences between these proteins may explain the reduced ability of EspC to cause cytopathic effects. However, these differences may confer a specialized role on EspC in the pathogenesis of EPEC, which is different from that of Pet in EAEC pathogenesis.Enteropathogenic Escherichia coli (EPEC), a leading cause of infantile diarrhea in developing countries, secretes effector molecules which allow bacteria to interact with their hosts and cause disease (7,11). EPEC secretes at least six proteins (Esp [E. coli secreted proteins]), four of which, EspA (11), EspD (14), EspB (26), and EspF (16), are secreted by a type III secretion system. Esp molecules as well as the secretion apparatus are encoded within a 35.6-kb pathogenicity island termed the locus of enterocyte effacement (LEE) (4, 15). EspA, EspD, and EspB proteins form a translocon for delivering effector molecules into the host cytosol (12,22). Tir (translocated intimin receptor) is translocated through this apparatus (10) and serves as a receptor for the EPEC adhesin intimin (9). Intimin is required for intimate bacterial attachment to host cells (8) and to focus the polymerization of cytoskeletal rearrangements leading to the disruption of intestinal microvilli, a phenotype known as the attaching and effacing (A/E) lesion (18).In addition to the secretion of the proteins via the type III secretion system mentioned above, EPEC also secretes a protein via the type V secretion system, the autotransporter pathway (25). This protein, EspC, shows the three classical domains (signal peptide, passenger domain, and ␤-domain) of the autotransporter proteins, which were first described for the immunoglobulin A (IgA) protease of Neisseria gonorrhoeae (23,25). EspC also has a conserved serine protease motif similar to the IgA protease but does not cleave IgA, as do several other members of the autotransporter family of proteins. In fact, EspC belongs to the subfamily of serine protease autotransporters of Enterobacteriaceae (SPATE) including Tsh, SepA, Pic, EspP, Sat, and Pet; none of these cleave IgA. Moreover, ...

Molecular characterization of virulence factors in diarrhoeagenic Escherichia coli isolates from children in Nairobi, Kenya.

“…This result concurs with observations by Navarro-García et al [15] who determined that it mainly confers virulence properties to EPEC. The presence of EspC PAI shows significance of acquisition of other virulent PAIs in aiding pathogenesis of diarrhoeal illness.…”

“…A multiplex PCR cycle was used as previously described [10] in amplification of DEC pathogenic genes. PCR conditions for the detection of PAIs were used as earlier described for she PAI (pic gene) [11], HPI PAI (irp2 and fyu A gene) [12], LEE PAI (eae gene) [13], TIA PAI (tia gene) [14], SHI-2 PAI (iutA gene) [13], EspC PAI (espC gene) [15] and O-island PAI (efa/lifA gene and pagC gene) [16]. The amplified PCR products were separated by electrophoresis in 2% agarose gels stained with ethidium bromide in Tris Borate (TBE) buffer at 100 V for one and half hours.…”

Bacterial Macroscopic Rope-like Fibers with Cytopathic and Adhesive Properties

“…The EspC protein of EPEC shows considerable homology with the IgA protease of Neisseria gonorrhoeae and Haemophilus influenza, although it does not display this activity (2,11). Enterotoxin and cytotoxin activities on rat jejunal segments and cultured epithelial cells have been demonstrated for EspC (12,13). Cytotoxicity is more efficient when EspC is translocated during contact of EPEC with epithelial cells (14), a process that apparently also requires the participation of the T3SS (15).…”

Efficient Translocation of EspC into Epithelial Cells Depends on Enteropathogenic Escherichia coli and Host Cell Contact

“…Our group has also found that EspC causes cytotoxic effects, including cytoskeletal damage that depends on EspC internalization and on its functional serine protease motif (30). These activities are similar to those induced by the Pet toxin, a homolog autotransporter to EspC (70% similarity) that is secreted by enteroaggregative E. coli.…”

“…Recently, we showed that the purified EspC has enterotoxic and cytotoxic activities on rat jejunum preparations mounted in Ussing chambers and on cultured epithelial cells, respectively (24,30). Our group has also found that EspC causes cytotoxic effects, including cytoskeletal damage that depends on EspC internalization and on its functional serine protease motif (30).…”