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RrgA is a pilus-associated adhesin in Streptococcus pneumoniae.

Nelson AL, Ries J, Bagnoli F, Dahlberg S, Fälker S, Rounioja S, Tschöp J, Morfeldt E, Ferlenghi I, Hilleringmann M, Holden DW, Rappuoli R, Normark S, Barocchi MA, Henriques-Normark B - Mol. Microbiol. (2007)

Bottom Line: Non-adherent RrgA-negative, RrgB- and RrgC-positive organisms produce pili, suggesting that pilus-mediated adherence is due to expression of RrgA, rather than the pilus backbone itself.In contrast, RrgA-positive strains with disrupted rrgB and rrgC genes exhibit wild-type adherence despite failure to produce pili by Western blot or immunoelectron microscopy.These data suggest that RrgA is central in pilus-mediated adherence and disease, even in the absence of polymeric pilus production.

View Article: PubMed Central - PubMed

Affiliation: Swedish Institute for Infectious Disease Control and Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
Adherence to host cells is important in microbial colonization of a mucosal surface, and Streptococcus pneumoniae adherence was significantly enhanced by expression of an extracellular pilus composed of three subunits, RrgA, RrgB and RrgC. We sought to determine which subunit(s) confers adherence. Bacteria deficient in RrgA are significantly less adherent than wild-type organisms, while overexpression of RrgA enhances adherence. Recombinant monomeric RrgA binds to respiratory cells, as does RrgC with less affinity, and pre-incubation of epithelial cells with RrgA reduces adherence of wild-type piliated pneumococci. Non-adherent RrgA-negative, RrgB- and RrgC-positive organisms produce pili, suggesting that pilus-mediated adherence is due to expression of RrgA, rather than the pilus backbone itself. In contrast, RrgA-positive strains with disrupted rrgB and rrgC genes exhibit wild-type adherence despite failure to produce pili by Western blot or immunoelectron microscopy. The density of bacteria colonizing the upper respiratory tract of mice inoculated with piliated RrgA-negative pneumococci was significantly less compared with wild-type; in contrast, non-piliated pneumococci expressing non-polymeric RrgA had similar numbers of bacteria in the nasopharynx as piliated wild-type bacteria. These data suggest that RrgA is central in pilus-mediated adherence and disease, even in the absence of polymeric pilus production.

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Expression of rrgA confers pilus-mediated adherence to human respiratory epithelial cells. A. rrgA, but not rrgB and rrgC, is necessary for pilus-mediated adherence to epithelial cells. Adherence of wild-type piliated TIGR4 (‘T4’) to A549 human respiratory epithelial cells was significantly greater than both non-piliated T4Δ(rrgA-srtD) compared with T4 deficient in the rrgA gene (‘T4ΔrrgA’). Adherence of T4 deficient in both rrgB and rrgC (‘T4ΔrrgBC’) was not notably different from wild-type organisms, but was significantly greater compared with T4ΔrrgA. Repeated-measure anova of data collected from three independent determinations indicates statistically significant differences within experimental conditions. Post hoc Bonferroni analyses identify specific significant differences: **P< 0.01. B. rrgA expression determines adherence to epithelial cells. T4 deficient in rrgA are significantly deficient in adherence (‘ΔrrgA’), compared with wild-type (T4), while trans-complementation restores wild-type adherence [‘ΔrrgA∇(lacE::rrgA’)]. Introduction of a second copy of rrgA inserted in trans in the lacE locus [‘∇(lacE::rrgA’] results in significant enhancement of adherence over wild-type levels. Statistical analyses were performed with repeated-measure anova of data collected from three independent determinations. Post hoc Bonferroni analyses identify specific significant differences: *P< 0.01 and **P< 0.001.
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fig01: Expression of rrgA confers pilus-mediated adherence to human respiratory epithelial cells. A. rrgA, but not rrgB and rrgC, is necessary for pilus-mediated adherence to epithelial cells. Adherence of wild-type piliated TIGR4 (‘T4’) to A549 human respiratory epithelial cells was significantly greater than both non-piliated T4Δ(rrgA-srtD) compared with T4 deficient in the rrgA gene (‘T4ΔrrgA’). Adherence of T4 deficient in both rrgB and rrgC (‘T4ΔrrgBC’) was not notably different from wild-type organisms, but was significantly greater compared with T4ΔrrgA. Repeated-measure anova of data collected from three independent determinations indicates statistically significant differences within experimental conditions. Post hoc Bonferroni analyses identify specific significant differences: **P< 0.01. B. rrgA expression determines adherence to epithelial cells. T4 deficient in rrgA are significantly deficient in adherence (‘ΔrrgA’), compared with wild-type (T4), while trans-complementation restores wild-type adherence [‘ΔrrgA∇(lacE::rrgA’)]. Introduction of a second copy of rrgA inserted in trans in the lacE locus [‘∇(lacE::rrgA’] results in significant enhancement of adherence over wild-type levels. Statistical analyses were performed with repeated-measure anova of data collected from three independent determinations. Post hoc Bonferroni analyses identify specific significant differences: *P< 0.01 and **P< 0.001.

Mentions: T4ΔrrgA was eightfold less adherent than wild-type T4, and therefore demonstrated adherence properties comparable to the non-piliated negative control strains, T4Δ(rrgA-srtD), which was fivefold less adherent than the wild-type parent (Fig. 1). T4ΔrrgBC, which expresses only the rrgA gene product, was found to be as adherent as the wild-type T4 strain, and was eightfold more adherent than T4ΔrrgA, and fivefold more adherent than T4Δ(rrgA-srtD) (Fig. 1). These data demonstrate that expression of RrgA, but not RrgB or RrgC, is required for pilus-mediated adherence to human respiratory epithelial cells. Further, expression of RrgA alone is sufficient to enhance adherence over non-piliated levels. In support of this model, it is notable that ΔrrgA organisms are out-competed by wild-type S. pneumoniae in a mouse model of pneumococcal pneumonia (Hava and Camilli, 2002).


RrgA is a pilus-associated adhesin in Streptococcus pneumoniae.

Nelson AL, Ries J, Bagnoli F, Dahlberg S, Fälker S, Rounioja S, Tschöp J, Morfeldt E, Ferlenghi I, Hilleringmann M, Holden DW, Rappuoli R, Normark S, Barocchi MA, Henriques-Normark B - Mol. Microbiol. (2007)

Expression of rrgA confers pilus-mediated adherence to human respiratory epithelial cells. A. rrgA, but not rrgB and rrgC, is necessary for pilus-mediated adherence to epithelial cells. Adherence of wild-type piliated TIGR4 (‘T4’) to A549 human respiratory epithelial cells was significantly greater than both non-piliated T4Δ(rrgA-srtD) compared with T4 deficient in the rrgA gene (‘T4ΔrrgA’). Adherence of T4 deficient in both rrgB and rrgC (‘T4ΔrrgBC’) was not notably different from wild-type organisms, but was significantly greater compared with T4ΔrrgA. Repeated-measure anova of data collected from three independent determinations indicates statistically significant differences within experimental conditions. Post hoc Bonferroni analyses identify specific significant differences: **P< 0.01. B. rrgA expression determines adherence to epithelial cells. T4 deficient in rrgA are significantly deficient in adherence (‘ΔrrgA’), compared with wild-type (T4), while trans-complementation restores wild-type adherence [‘ΔrrgA∇(lacE::rrgA’)]. Introduction of a second copy of rrgA inserted in trans in the lacE locus [‘∇(lacE::rrgA’] results in significant enhancement of adherence over wild-type levels. Statistical analyses were performed with repeated-measure anova of data collected from three independent determinations. Post hoc Bonferroni analyses identify specific significant differences: *P< 0.01 and **P< 0.001.
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Related In: Results  -  Collection

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fig01: Expression of rrgA confers pilus-mediated adherence to human respiratory epithelial cells. A. rrgA, but not rrgB and rrgC, is necessary for pilus-mediated adherence to epithelial cells. Adherence of wild-type piliated TIGR4 (‘T4’) to A549 human respiratory epithelial cells was significantly greater than both non-piliated T4Δ(rrgA-srtD) compared with T4 deficient in the rrgA gene (‘T4ΔrrgA’). Adherence of T4 deficient in both rrgB and rrgC (‘T4ΔrrgBC’) was not notably different from wild-type organisms, but was significantly greater compared with T4ΔrrgA. Repeated-measure anova of data collected from three independent determinations indicates statistically significant differences within experimental conditions. Post hoc Bonferroni analyses identify specific significant differences: **P< 0.01. B. rrgA expression determines adherence to epithelial cells. T4 deficient in rrgA are significantly deficient in adherence (‘ΔrrgA’), compared with wild-type (T4), while trans-complementation restores wild-type adherence [‘ΔrrgA∇(lacE::rrgA’)]. Introduction of a second copy of rrgA inserted in trans in the lacE locus [‘∇(lacE::rrgA’] results in significant enhancement of adherence over wild-type levels. Statistical analyses were performed with repeated-measure anova of data collected from three independent determinations. Post hoc Bonferroni analyses identify specific significant differences: *P< 0.01 and **P< 0.001.
Mentions: T4ΔrrgA was eightfold less adherent than wild-type T4, and therefore demonstrated adherence properties comparable to the non-piliated negative control strains, T4Δ(rrgA-srtD), which was fivefold less adherent than the wild-type parent (Fig. 1). T4ΔrrgBC, which expresses only the rrgA gene product, was found to be as adherent as the wild-type T4 strain, and was eightfold more adherent than T4ΔrrgA, and fivefold more adherent than T4Δ(rrgA-srtD) (Fig. 1). These data demonstrate that expression of RrgA, but not RrgB or RrgC, is required for pilus-mediated adherence to human respiratory epithelial cells. Further, expression of RrgA alone is sufficient to enhance adherence over non-piliated levels. In support of this model, it is notable that ΔrrgA organisms are out-competed by wild-type S. pneumoniae in a mouse model of pneumococcal pneumonia (Hava and Camilli, 2002).

Bottom Line: Non-adherent RrgA-negative, RrgB- and RrgC-positive organisms produce pili, suggesting that pilus-mediated adherence is due to expression of RrgA, rather than the pilus backbone itself.In contrast, RrgA-positive strains with disrupted rrgB and rrgC genes exhibit wild-type adherence despite failure to produce pili by Western blot or immunoelectron microscopy.These data suggest that RrgA is central in pilus-mediated adherence and disease, even in the absence of polymeric pilus production.

View Article: PubMed Central - PubMed

Affiliation: Swedish Institute for Infectious Disease Control and Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
Adherence to host cells is important in microbial colonization of a mucosal surface, and Streptococcus pneumoniae adherence was significantly enhanced by expression of an extracellular pilus composed of three subunits, RrgA, RrgB and RrgC. We sought to determine which subunit(s) confers adherence. Bacteria deficient in RrgA are significantly less adherent than wild-type organisms, while overexpression of RrgA enhances adherence. Recombinant monomeric RrgA binds to respiratory cells, as does RrgC with less affinity, and pre-incubation of epithelial cells with RrgA reduces adherence of wild-type piliated pneumococci. Non-adherent RrgA-negative, RrgB- and RrgC-positive organisms produce pili, suggesting that pilus-mediated adherence is due to expression of RrgA, rather than the pilus backbone itself. In contrast, RrgA-positive strains with disrupted rrgB and rrgC genes exhibit wild-type adherence despite failure to produce pili by Western blot or immunoelectron microscopy. The density of bacteria colonizing the upper respiratory tract of mice inoculated with piliated RrgA-negative pneumococci was significantly less compared with wild-type; in contrast, non-piliated pneumococci expressing non-polymeric RrgA had similar numbers of bacteria in the nasopharynx as piliated wild-type bacteria. These data suggest that RrgA is central in pilus-mediated adherence and disease, even in the absence of polymeric pilus production.

Show MeSH
Related in: MedlinePlus