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CcpA ensures optimal metabolic fitness of Streptococcus pneumoniae.

Carvalho SM, Kloosterman TG, Kuipers OP, Neves AR - PLoS ONE (2011)

Bottom Line: In agreement, CcpA influenced the level of many intracellular metabolites potentially involved in metabolic regulation.Our data strengthen the view that a true understanding of cell physiology demands thorough analyses at different cellular levels.Moreover, integration of transcriptional and metabolic data uncovered a link between CcpA and the association of surface molecules (e.g. capsule) to the cell wall.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal.

ABSTRACT
In gram-positive bacteria, the transcriptional regulator CcpA is at the core of catabolite control mechanisms. In the human pathogen Streptococcus pneumoniae, links between CcpA and virulence have been established, but its role as a master regulator in different nutritional environments remains to be elucidated. Thus, we performed whole-transcriptome and metabolic analyses of S. pneumoniae D39 and its isogenic ccpA mutant during growth on glucose or galactose, rapidly and slowly metabolized carbohydrates presumably encountered by the bacterium in different host niches. CcpA affected the expression of up to 19% of the genome covering multiple cellular processes, including virulence, regulatory networks and central metabolism. Its prevalent function as a repressor was observed on glucose, but unexpectedly also on galactose. Carbohydrate-dependent CcpA regulation was also observed, as for the tagatose 6-phosphate pathway genes, which were activated by galactose and repressed by glucose. Metabolite analyses revealed that two pathways for galactose catabolism are functionally active, despite repression of the Leloir genes by CcpA. Surprisingly, galactose-induced mixed-acid fermentation apparently required CcpA, since genes involved in this type of metabolism were mostly under CcpA-repression. These findings indicate that the role of CcpA extends beyond transcriptional regulation, which seemingly is overlaid by other regulatory mechanisms. In agreement, CcpA influenced the level of many intracellular metabolites potentially involved in metabolic regulation. Our data strengthen the view that a true understanding of cell physiology demands thorough analyses at different cellular levels. Moreover, integration of transcriptional and metabolic data uncovered a link between CcpA and the association of surface molecules (e.g. capsule) to the cell wall. Hence, CcpA may play a key role in mediating the interaction of S. pneumoniae with its host. Overall, our results support the hypothesis that S. pneumoniae optimizes basic metabolic processes, likely enhancing in vivo fitness, in a CcpA-mediated manner.

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Related in: MedlinePlus

Amount of capsule polysaccharide in D39 and its isogenic ccpA mutant.Estimation of capsule was performed based on the determination of its glucuronic acid content in strains D39 and D39ΔccpA in mid-exponential (white bars, OD600 of 0.35±0.02) and transition-to-stationary (black bars, OD600 of 1.3±0.1) cultures grown in CDM containing 56±1 mM Glc (white background) or 57±1 mM Gal (light grey background). Hatched bars indicate loose capsule polysaccharide. All the determinations were done twice in two independent cultures and the values are means ± SD.
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pone-0026707-g006: Amount of capsule polysaccharide in D39 and its isogenic ccpA mutant.Estimation of capsule was performed based on the determination of its glucuronic acid content in strains D39 and D39ΔccpA in mid-exponential (white bars, OD600 of 0.35±0.02) and transition-to-stationary (black bars, OD600 of 1.3±0.1) cultures grown in CDM containing 56±1 mM Glc (white background) or 57±1 mM Gal (light grey background). Hatched bars indicate loose capsule polysaccharide. All the determinations were done twice in two independent cultures and the values are means ± SD.

Mentions: It has previously been reported that CcpA-deficient mutants show attenuated virulence, and this behaviour was associated with lower expression of genes committed to the synthesis of the pneumococcal capsule [11]. Our metabolome data showed significant differences in the pools of the capsule cytoplasmic precursors UDP-glucose and α-G1P. Therefore, we deemed it important to measure the amounts of capsule produced in D39 and its ccpA mutant (Fig. 6). Curiously, inactivation of ccpA did not affect the total amount of capsule either on Glc- or Gal-CDM. However, in Gal-containing medium the amount of capsule produced was higher for both strains. Furthermore, a major difference in Gal-CDM was the amount of loose capsule (not attached to the cell wall), which accounted for about 94% of the total polysaccharide in D39ΔccpA (Fig. 6). In the parent strain, only a small fraction (∼13%) of loose capsule was detected, exclusively in the TS phase of growth.


CcpA ensures optimal metabolic fitness of Streptococcus pneumoniae.

Carvalho SM, Kloosterman TG, Kuipers OP, Neves AR - PLoS ONE (2011)

Amount of capsule polysaccharide in D39 and its isogenic ccpA mutant.Estimation of capsule was performed based on the determination of its glucuronic acid content in strains D39 and D39ΔccpA in mid-exponential (white bars, OD600 of 0.35±0.02) and transition-to-stationary (black bars, OD600 of 1.3±0.1) cultures grown in CDM containing 56±1 mM Glc (white background) or 57±1 mM Gal (light grey background). Hatched bars indicate loose capsule polysaccharide. All the determinations were done twice in two independent cultures and the values are means ± SD.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198803&req=5

pone-0026707-g006: Amount of capsule polysaccharide in D39 and its isogenic ccpA mutant.Estimation of capsule was performed based on the determination of its glucuronic acid content in strains D39 and D39ΔccpA in mid-exponential (white bars, OD600 of 0.35±0.02) and transition-to-stationary (black bars, OD600 of 1.3±0.1) cultures grown in CDM containing 56±1 mM Glc (white background) or 57±1 mM Gal (light grey background). Hatched bars indicate loose capsule polysaccharide. All the determinations were done twice in two independent cultures and the values are means ± SD.
Mentions: It has previously been reported that CcpA-deficient mutants show attenuated virulence, and this behaviour was associated with lower expression of genes committed to the synthesis of the pneumococcal capsule [11]. Our metabolome data showed significant differences in the pools of the capsule cytoplasmic precursors UDP-glucose and α-G1P. Therefore, we deemed it important to measure the amounts of capsule produced in D39 and its ccpA mutant (Fig. 6). Curiously, inactivation of ccpA did not affect the total amount of capsule either on Glc- or Gal-CDM. However, in Gal-containing medium the amount of capsule produced was higher for both strains. Furthermore, a major difference in Gal-CDM was the amount of loose capsule (not attached to the cell wall), which accounted for about 94% of the total polysaccharide in D39ΔccpA (Fig. 6). In the parent strain, only a small fraction (∼13%) of loose capsule was detected, exclusively in the TS phase of growth.

Bottom Line: In agreement, CcpA influenced the level of many intracellular metabolites potentially involved in metabolic regulation.Our data strengthen the view that a true understanding of cell physiology demands thorough analyses at different cellular levels.Moreover, integration of transcriptional and metabolic data uncovered a link between CcpA and the association of surface molecules (e.g. capsule) to the cell wall.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal.

ABSTRACT
In gram-positive bacteria, the transcriptional regulator CcpA is at the core of catabolite control mechanisms. In the human pathogen Streptococcus pneumoniae, links between CcpA and virulence have been established, but its role as a master regulator in different nutritional environments remains to be elucidated. Thus, we performed whole-transcriptome and metabolic analyses of S. pneumoniae D39 and its isogenic ccpA mutant during growth on glucose or galactose, rapidly and slowly metabolized carbohydrates presumably encountered by the bacterium in different host niches. CcpA affected the expression of up to 19% of the genome covering multiple cellular processes, including virulence, regulatory networks and central metabolism. Its prevalent function as a repressor was observed on glucose, but unexpectedly also on galactose. Carbohydrate-dependent CcpA regulation was also observed, as for the tagatose 6-phosphate pathway genes, which were activated by galactose and repressed by glucose. Metabolite analyses revealed that two pathways for galactose catabolism are functionally active, despite repression of the Leloir genes by CcpA. Surprisingly, galactose-induced mixed-acid fermentation apparently required CcpA, since genes involved in this type of metabolism were mostly under CcpA-repression. These findings indicate that the role of CcpA extends beyond transcriptional regulation, which seemingly is overlaid by other regulatory mechanisms. In agreement, CcpA influenced the level of many intracellular metabolites potentially involved in metabolic regulation. Our data strengthen the view that a true understanding of cell physiology demands thorough analyses at different cellular levels. Moreover, integration of transcriptional and metabolic data uncovered a link between CcpA and the association of surface molecules (e.g. capsule) to the cell wall. Hence, CcpA may play a key role in mediating the interaction of S. pneumoniae with its host. Overall, our results support the hypothesis that S. pneumoniae optimizes basic metabolic processes, likely enhancing in vivo fitness, in a CcpA-mediated manner.

Show MeSH
Related in: MedlinePlus