Identification of regions of Ail required for the invasion and serum resistance phenotypes

Abstract

Yersinia enterocolitica is an enteric pathogen that has served as a model system for the study of microbial pathogenesis. Numerous virulence gene have been identified both on the virulence plasmid and on the chromosome. One of the chromosomal genes that is highly correlated with virulence is ail, a gene identified along with inv in a screen for Y. enterocolitica genes that could confer an invasive phenotype to Escherichia coli. Ail also promotes serum resistance in both E. coli and Y. enterocolitica. Several virulence factors homologous to Ail have been identified in other pathogens, yet very little is known about what constitutes the functional domain(s) of these proteins. Proteins in this family are predicted to consist of eight transmembrane β‐sheets and four cell surface‐exposed loops. We constructed and characterized a number of insertion, deletion and point mutations in the regions of ail predicted to encode the cell surface loops. The results from the analysis of these mutants indicate that cell surface loops one and four do not directly promote invasion or serum resistance, whereas mutations in loop three appear to modulate both phenotypes. Analysis of mutations in loop 2 suggests that this surface‐exposed loop contains sequences required for serum resistance and invasion. In addition, a peptide derived from the sequence of loop 2 was able specifically to inhibit Ail‐mediated invasion in a dose‐dependent manner. These results suggest that Ail directly promotes invasion and that loop 2 contains an active site, perhaps a receptor‐binding domain. Analyses of the mutations also suggest that the serum resistance and invasion phenotypes may be separable, because there are numerous mutations that affect one phenotype but not the other.

The Different Strategies Used by Salmonella to Invade Host Cells

“…These proteins do not exhibit homologous regions that could be related to these two identical roles. In Ail, the cell invasion property is associated with loop2 whereas loop3 and more precisely the region from the amino acids G113 to V159 was shown to be the minimal region of Rck required and sufficient for cell adhesion and invasion (Miller et al, 2001;Rosselin et al, 2010).…”

Biochemical characterization of the enterotoxigenic Escherichia coli LeoA protein

“…Kuehn and co-workers noted in their pioneering studies of LT secretion via vesicles that OmpX is present in ETEC outer membrane and vesicle preparations, that the abundance of OmpX differs as a function of the growth medium, and that this protein is not detected in preparations from non-pathogenic strains (Horstman & Kuehn, 2000). These authors also noted that OmpX is homologous to the Yersinia Ail protein, which has been suggested to promote bacterial invasion (Miller et al, 2001), and the OmpX adhesin of Enterobacter cloacae, which promotes invasiveness in rabbit ileal enterocytes (de Kort et al, 1994). It remains to be determined why deletion of LeoA alters OmpX abundance in the outer membrane.…”

Phenotypic characterization of OmpX, an Ail homologue of Yersinia pestis KIM

“…Although Y. enterocolitica Ail has been extensively studied (Miller et al, 1990(Miller et al, , 2001, similar studies have not been reported for its Y. pestis homologues. Two previous studies showed that loss of an unidentified protein in the Y. pestis outer membrane affects autoaggregation (Podladchikova & Rykova, 2006) and adhesion to phagocytes (Kukleva et al, 2000).…”