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Role of the Group B antigen of Streptococcus agalactiae: a peptidoglycan-anchored polysaccharide involved in cell wall biogenesis.

Caliot É, Dramsi S, Chapot-Chartier MP, Courtin P, Kulakauskas S, Péchoux C, Trieu-Cuot P, Mistou MY - PLoS Pathog. (2012)

Bottom Line: Furthermore, vancomycin labeling and peptidoglycan structure analysis demonstrated that, in the absence of GBC, cells failed to initiate normal PG synthesis and cannot complete polymerization of the murein sacculus.Collectively, these findings show that GBC is an essential component of the cell wall of S. agalactiae whose function is reminiscent of that of conventional wall teichoic acids found in Staphylococcus aureus or Bacillus subtilis.Furthermore, our findings raise the possibility that GBC-like molecules play a major role in the growth of most if not all beta-hemolytic streptococci.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité des Bactéries Pathogènes à Gram positif, Paris, France.

ABSTRACT
Streptococcus agalactiae (Group B streptococcus, GBS) is a leading cause of infections in neonates and an emerging pathogen in adults. The Lancefield Group B carbohydrate (GBC) is a peptidoglycan-anchored antigen that defines this species as a Group B Streptococcus. Despite earlier immunological and biochemical characterizations, the function of this abundant glycopolymer has never been addressed experimentally. Here, we inactivated the gene gbcO encoding a putative UDP-N-acetylglucosamine-1-phosphate:lipid phosphate transferase thought to catalyze the first step of GBC synthesis. Indeed, the gbcO mutant was unable to synthesize the GBC polymer, and displayed an important growth defect in vitro. Electron microscopy study of the GBC-depleted strain of S. agalactiae revealed a series of growth-related abnormalities: random placement of septa, defective cell division and separation processes, and aberrant cell morphology. Furthermore, vancomycin labeling and peptidoglycan structure analysis demonstrated that, in the absence of GBC, cells failed to initiate normal PG synthesis and cannot complete polymerization of the murein sacculus. Finally, the subcellular localization of the PG hydrolase PcsB, which has a critical role in cell division of streptococci, was altered in the gbcO mutant. Collectively, these findings show that GBC is an essential component of the cell wall of S. agalactiae whose function is reminiscent of that of conventional wall teichoic acids found in Staphylococcus aureus or Bacillus subtilis. Furthermore, our findings raise the possibility that GBC-like molecules play a major role in the growth of most if not all beta-hemolytic streptococci.

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Fluorescent immunolocalization of the putative peptidoglycan hydrolase PcsB.Exponentially growing NEM316 WT, ΔgbcO mutant and ΔgbcOpTCVΩgbcO complemented strains were harvested, transferred to glass slide, and fixed. IFM with anti-PcsB serum and DAPI staining were performed as described in Materials and Methods.
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ppat-1002756-g008: Fluorescent immunolocalization of the putative peptidoglycan hydrolase PcsB.Exponentially growing NEM316 WT, ΔgbcO mutant and ΔgbcOpTCVΩgbcO complemented strains were harvested, transferred to glass slide, and fixed. IFM with anti-PcsB serum and DAPI staining were performed as described in Materials and Methods.

Mentions: The consequences of the inactivation of the gbcO gene on cell morphology and division were reminiscent to those observed for a PcsB- mutant of S. agalactiae strain 6313 [32], [33]. PcsB (protein required for cell wall separation) is a cell surface located putative PG hydrolase that has orthologs in all species of the Streptococcaceae family. This protein possesses a cysteine-histidine-dependent-amidohydrolase-peptidase (CHAP) domain and is required for proper cell wall synthesis and efficient cell separation in S. agalactiae and other streptococci [31], [33]–[35], suggesting that it is involved in PG remodeling. We therefore hypothesized that GBC deprivation could impact the localization of PcsB and the associated PG hydrolase activity. To test this hypothesis, IFM experiments were performed to localize PcsB on the surface of bacterial cells harvested in the exponential phase of growth (Figure 8). A PcsB-specific signal located in the equatorial zone of dividing cells corresponding to the site of active PG synthesis was observed in NEM316 WT and complemented strains. This localization was recently observed for the orthologous PcsB protein in Streptococcus pneumonie[36]. By contrast, no regular labeling pattern can be distinguished in ΔgbcO mutant and PcsB-associated signals were unevenly distributed on the cell surface and in some instances accumulated in foci. This result indicates that GBC is involved in the proper localization of PcsB, a cell wall protein involved in bacterial division and PG biosynthesis.


Role of the Group B antigen of Streptococcus agalactiae: a peptidoglycan-anchored polysaccharide involved in cell wall biogenesis.

Caliot É, Dramsi S, Chapot-Chartier MP, Courtin P, Kulakauskas S, Péchoux C, Trieu-Cuot P, Mistou MY - PLoS Pathog. (2012)

Fluorescent immunolocalization of the putative peptidoglycan hydrolase PcsB.Exponentially growing NEM316 WT, ΔgbcO mutant and ΔgbcOpTCVΩgbcO complemented strains were harvested, transferred to glass slide, and fixed. IFM with anti-PcsB serum and DAPI staining were performed as described in Materials and Methods.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375309&req=5

ppat-1002756-g008: Fluorescent immunolocalization of the putative peptidoglycan hydrolase PcsB.Exponentially growing NEM316 WT, ΔgbcO mutant and ΔgbcOpTCVΩgbcO complemented strains were harvested, transferred to glass slide, and fixed. IFM with anti-PcsB serum and DAPI staining were performed as described in Materials and Methods.
Mentions: The consequences of the inactivation of the gbcO gene on cell morphology and division were reminiscent to those observed for a PcsB- mutant of S. agalactiae strain 6313 [32], [33]. PcsB (protein required for cell wall separation) is a cell surface located putative PG hydrolase that has orthologs in all species of the Streptococcaceae family. This protein possesses a cysteine-histidine-dependent-amidohydrolase-peptidase (CHAP) domain and is required for proper cell wall synthesis and efficient cell separation in S. agalactiae and other streptococci [31], [33]–[35], suggesting that it is involved in PG remodeling. We therefore hypothesized that GBC deprivation could impact the localization of PcsB and the associated PG hydrolase activity. To test this hypothesis, IFM experiments were performed to localize PcsB on the surface of bacterial cells harvested in the exponential phase of growth (Figure 8). A PcsB-specific signal located in the equatorial zone of dividing cells corresponding to the site of active PG synthesis was observed in NEM316 WT and complemented strains. This localization was recently observed for the orthologous PcsB protein in Streptococcus pneumonie[36]. By contrast, no regular labeling pattern can be distinguished in ΔgbcO mutant and PcsB-associated signals were unevenly distributed on the cell surface and in some instances accumulated in foci. This result indicates that GBC is involved in the proper localization of PcsB, a cell wall protein involved in bacterial division and PG biosynthesis.

Bottom Line: Furthermore, vancomycin labeling and peptidoglycan structure analysis demonstrated that, in the absence of GBC, cells failed to initiate normal PG synthesis and cannot complete polymerization of the murein sacculus.Collectively, these findings show that GBC is an essential component of the cell wall of S. agalactiae whose function is reminiscent of that of conventional wall teichoic acids found in Staphylococcus aureus or Bacillus subtilis.Furthermore, our findings raise the possibility that GBC-like molecules play a major role in the growth of most if not all beta-hemolytic streptococci.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité des Bactéries Pathogènes à Gram positif, Paris, France.

ABSTRACT
Streptococcus agalactiae (Group B streptococcus, GBS) is a leading cause of infections in neonates and an emerging pathogen in adults. The Lancefield Group B carbohydrate (GBC) is a peptidoglycan-anchored antigen that defines this species as a Group B Streptococcus. Despite earlier immunological and biochemical characterizations, the function of this abundant glycopolymer has never been addressed experimentally. Here, we inactivated the gene gbcO encoding a putative UDP-N-acetylglucosamine-1-phosphate:lipid phosphate transferase thought to catalyze the first step of GBC synthesis. Indeed, the gbcO mutant was unable to synthesize the GBC polymer, and displayed an important growth defect in vitro. Electron microscopy study of the GBC-depleted strain of S. agalactiae revealed a series of growth-related abnormalities: random placement of septa, defective cell division and separation processes, and aberrant cell morphology. Furthermore, vancomycin labeling and peptidoglycan structure analysis demonstrated that, in the absence of GBC, cells failed to initiate normal PG synthesis and cannot complete polymerization of the murein sacculus. Finally, the subcellular localization of the PG hydrolase PcsB, which has a critical role in cell division of streptococci, was altered in the gbcO mutant. Collectively, these findings show that GBC is an essential component of the cell wall of S. agalactiae whose function is reminiscent of that of conventional wall teichoic acids found in Staphylococcus aureus or Bacillus subtilis. Furthermore, our findings raise the possibility that GBC-like molecules play a major role in the growth of most if not all beta-hemolytic streptococci.

Show MeSH
Related in: MedlinePlus