Identification of a domain in Rck, a product of the Salmonella typhimurium virulence plasmid, required for both serum resistance and cell invasion

Abstract

Rck is encoded on the Salmonella typhimurium virulence plasmid and is a member of a family of related 17to 19-kDa outer membrane proteins of Enterobacteriaceae, including Ail (Yersinia enterocolitica) and PagC (S. typhimurium). Structural models for these proteins predict eight membrane-spanning domains alternating with hydrophilic inner and outer loops. When expressed in Escherichia coli, Rck and Ail, but not PagC, confer highlevel resistance to the bactericidal activity of complement as well as the ability to adhere to and invade mammalian cell lines. To identify functional domains of Rck, we made and screened random mutations in Rck for decreased bioactivity. We found that a single amino acid substitution (glycine to aspartic acid) in the putative third outer loop greatly reduced Rck-mediated serum resistance and eukaryotic cell invasion. We then constructed two chimeric proteins between Rck and PagC. Substitution of the C-terminal half of Rck with the corresponding PagC fragment containing both the third and the fourth outer loops abolishes the Rck-mediated serum resistance and invasion phenotypes. Substitution of Rck with a smaller C-terminal portion of PagC containing the fourth outer loop did not affect the invasive phenotype or serum resistance. These data reveal that the third putative outer membrane loop region is important for the virulence-associated properties of the Rck protein and suggest a similarity between the mechanism of serum resistance and epithelial cell invasion involving the same domain of Rck.

An AIL family protein promotes type three secretion system-1-independent invasion and pathogenesis ofSalmonella entericaserovar Typhi

“…Although the domains of laminin and fibronectin that bind Salmonella AIL are not known, 46 amino acids of Rck (113-159) are responsible for host cell adhesion and internalization, of which peptide 140-150 may be involved in adhesion and peptide 150-159 for invasion (Rosselin et al, 2010). In addition, G118D mutation greatly reduces Rck-mediated adhesion and invasion (Cirillo et al, 1996). We show that four loop-like structures of T2942 (amino acids 37-48, 70-89, 116-131 and 155-164) are important for adhesion, while only the 20-mer peptide (70-89) is directly involved in the invasion of non-phagocytic cells.…”

“…Rck and PagC belong to a family of Yersinia AIL-like proteins and are required for laminin and fibronectin binding (Crago and Koronakis, 1999). Rck, but not PagC also promotes epithelial invasion (Cirillo et al, 1996). MisL and ShdA were reported to bind fibronectin and help in intestinal colonization of S. Typhimurium (Kingsley et al, 2002;Dorsey et al, 2005), while giant, non-fimbrial adhesin SiiE binds to N-acetyl glucosamine and sialic acid of the apical membrane of the intestinal epithelial cells (IECs; Wagner et al, 2014).…”

Salmonellastress management and its relevance to behaviour during intestinal colonisation and infection

“…sdiA in Salmonella was first described by Ahmer et al [167] when this group suggested a link between sdiA and positive regulation of ten genes on the virulence plasmid including the previously characterised rck gene responsible for increased resistance to complement killing and adhesion to epithelial cells [168]. Although sdiA in S. typhimurium seems to be suppressed by conditioned medium to the extent observed in E. coli [169], no AI-1 like metabolite was ever detected also in S. typhimurium conditioned media.…”

Ail provides multiple mechanisms of serum resistance to Yersinia pestis

“…Extensive studies on the involvement of Y. enterocolitica Ail residues in adhesion and serum resistance revealed the contribution of D90 and V91 (numbered according to the unprocessed form) to serum resistance (Miller et al, 2001). Expressing double mutants, of Y. pestis Ail residues corresponding to homologous residues that no longer confer full serum resistance in Yersinia enterocolitica Ail (Miller et al, 2001) and Salmonella enterica Rck (Cirillo et al, 1996). B.…”

“…Each has the ability to recruit functional C4BP and factor H to the surface of the bacteria, providing potential mechanisms of serum resistance (Biedzka-Sarek et al, 2008a;2008b;Ho et al, 2011;Ho et al, 2010;Ho et al, 2012a;2012b). Specific amino acids in the extracellular loops of Y. enterocolitica Ail and S. enterica Rck are required for serum resistance in these bacteria (Cirillo et al, 1996;Miller et al, 2001).…”