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Functional recruitment of human complement inhibitor C4B-binding protein to outer membrane protein Rck of Salmonella.

ABSTRACT: Resistance to complement mediated killing, or serum resistance, is a common trait of pathogenic bacteria. Rck is a 17 kDa outer membrane protein encoded on the virulence plasmid of Salmonella enterica serovars Typhimurium and Enteritidis. When expressed in either E. coli or S. enterica Typhimurium, Rck confers LPS-independent serum resistance as well as the ability to bind to and invade mammalian cells. Having recently shown that Rck binds the inhibitor of the alternative pathway of complement, factor H (fH), we hypothesized that Rck can also bind the inhibitor of the classical and lectin pathways, C4b-binding protein (C4BP). Using flow cytometry and direct binding assays, we demonstrate that E. coli expressing Rck binds C4BP from heat-inactivated serum and by using the purified protein. No binding was detected in the absence of Rck expression. C4BP bound to Rck is functional, as we observed factor I-mediated cleavage of C4b in cofactor assays. In competition assays, binding of radiolabeled C4BP to Rck was reduced by increasing concentrations of unlabeled protein. No effect was observed by increasing heparin or salt concentrations, suggesting mainly non-ionic interactions. Reduced binding of C4BP mutants lacking complement control protein domains (CCPs) 7 or 8 was observed compared to wt C4BP, suggesting that these CCPs are involved in Rck binding. While these findings are restricted to Rck expression in E. coli, these data suggest that C4BP binding may be an additional mechanism of Rck-mediated complement resistance.

SUBMITTER: Ho DK

PROVIDER: S-EPMC3213152 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Functional recruitment of human complement inhibitor C4B-binding protein to outer membrane protein Rck of Salmonella.

Ho Derek K DK Tissari Jorma J Järvinen Hanna M HM Blom Anna M AM Meri Seppo S Jarva Hanna H

PloS one 20111110 11

Resistance to complement mediated killing, or serum resistance, is a common trait of pathogenic bacteria. Rck is a 17 kDa outer membrane protein encoded on the virulence plasmid of Salmonella enterica serovars Typhimurium and Enteritidis. When expressed in either E. coli or S. enterica Typhimurium, Rck confers LPS-independent serum resistance as well as the ability to bind to and invade mammalian cells. Having recently shown that Rck binds the inhibitor of the alternative pathway of complement, ...[more]

PMID: 22102907

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Project description:The rck open reading frame (ORF) on the pefI-srgC operon encodes an outer membrane protein responsible for invasion of nonphagocytic cell lines and resistance to complement-mediated killing. Until now, the rck ORF was only detected on the virulence plasmids of three serovars of Salmonella subsp. enterica (i.e., Bovismorbificans, Enteritidis, and Typhimurium). The increasing number of Salmonella genome sequences allowed us to use a combination of reference sequences and whole-genome multilocus sequence typing (wgMLST) data analysis to probe the presence of the operon and of rck in a wide array of isolates belonging to all Salmonella species and subspecies. We established the presence of partial or complete operons in 61 subsp. enterica serovars as well as in 4 other subspecies with various syntenies and frequencies. The rck ORF itself was retrieved in 36 subsp. enterica serovars and in two subspecies with either chromosomal or plasmid-borne localization. It displays high conservation of its sequence within the genus, and we demonstrated that most of the allelic variations identified did not alter the virulence properties of the protein. However, we demonstrated the importance of the residue at position 38 (at the level of the first extracellular loop of the protein) in the invasin function of Rck. Altogether, our results highlight that rck is not restricted to the three formerly identified serovars and could therefore have a more important role in virulence than previously expected. Moreover, this work raises questions about the mechanisms involved in rck acquisition and about virulence plasmid distribution and evolution. IMPORTANCE The foodborne pathogen Salmonella is responsible for a wide variety of pathologies depending on the infected host, the infecting serovars, and its set of virulence factors. However, the implication of each of these virulence factors and their role in the specific host-pathogen interplay are not fully understood. The significance of our research is in determining the distribution of one of these factors, the virulence plasmid-encoded invasin and resistance to complement killing protein Rck. In addition to providing elements of reflection concerning the mechanisms of acquisition of specific virulence genes in certain serotypes, this work will help to understand the role of Rck in the pathogenesis of Salmonella.

Dataset Information

Functional recruitment of human complement inhibitor C4B-binding protein to outer membrane protein Rck of Salmonella.

Publications

Functional recruitment of human complement inhibitor C4B-binding protein to outer membrane protein Rck of Salmonella.

Similar Datasets

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