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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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Genes differentially expressed due to inactivation of CcpA.A. Venn diagram of the genes significantly differentially transcribed due to loss of CcpA. Transcript levels in D39ΔccpA were compared to D39 wild-type in Glc_M, Gal_M, Glc_TS, Gal_TS as generated by VENNY (http://bioinfogp.cnb.csic.es/tools/venny/index.html). For all intersections, which are not drawn to scale, the numbers of genes are indicated. The total number of genes influenced in each condition tested was: Glc_M, 334 (15.3%), Glc_TS, 418 (19.2%), Gal_M, 299 (13.7%), and Gal_TS 299 (13.7%) in a universe of 2177 genes in the genome of S. pneumoniae. B. Numbers of genes significantly differently transcribed by the ccpA deletion in strain D39 ordered by COG categories. White bars represent M phase of growth, black bars TS phase of growth. [C] Energy production and conversion; [D] Cell cycle control, cell division; chromosome partitioning; [E] Amino acid transport and metabolism; [F] Nucleotide transport and metabolism; [G] Carbohydrate transport and metabolism; [H] Coenzyme transport and metabolism; [I] Lipid transport and metabolism; [J] Translation, ribosomal structure and biogenesis; [K] Transcription; [L] Replication, recombination and repair; [M] Cell wall/membrane/envelope biogenesis; [O] Posttranslational modification, protein turnover, chaperones; [P] Inorganic ion transport and metabolism; [Q] Secondary metabolites biosynthesis, transport and catabolism; [R] General function prediction only; [S] Function unknown; [T] Signal transduction mechanisms; [U] Intracellular trafficking, secretion, and vesicular transport; [V] Defense mechanisms; [X] No prediction. The ratio for each gene is >1.5 or <0.66, but >2 or <0.5 in at least one of the four conditions. The number 2177 represents the total number of amplicons analyzed, covering the entire genome of S. pneumoniae D39. This number is slightly higher than the number of genes in the genome of D39 (2069), since some amplicons were present in versions for different strains (i.e. TIGR4, R6, or D39) or in 2 copies.
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pone-0026707-g001: Genes differentially expressed due to inactivation of CcpA.A. Venn diagram of the genes significantly differentially transcribed due to loss of CcpA. Transcript levels in D39ΔccpA were compared to D39 wild-type in Glc_M, Gal_M, Glc_TS, Gal_TS as generated by VENNY (http://bioinfogp.cnb.csic.es/tools/venny/index.html). For all intersections, which are not drawn to scale, the numbers of genes are indicated. The total number of genes influenced in each condition tested was: Glc_M, 334 (15.3%), Glc_TS, 418 (19.2%), Gal_M, 299 (13.7%), and Gal_TS 299 (13.7%) in a universe of 2177 genes in the genome of S. pneumoniae. B. Numbers of genes significantly differently transcribed by the ccpA deletion in strain D39 ordered by COG categories. White bars represent M phase of growth, black bars TS phase of growth. [C] Energy production and conversion; [D] Cell cycle control, cell division; chromosome partitioning; [E] Amino acid transport and metabolism; [F] Nucleotide transport and metabolism; [G] Carbohydrate transport and metabolism; [H] Coenzyme transport and metabolism; [I] Lipid transport and metabolism; [J] Translation, ribosomal structure and biogenesis; [K] Transcription; [L] Replication, recombination and repair; [M] Cell wall/membrane/envelope biogenesis; [O] Posttranslational modification, protein turnover, chaperones; [P] Inorganic ion transport and metabolism; [Q] Secondary metabolites biosynthesis, transport and catabolism; [R] General function prediction only; [S] Function unknown; [T] Signal transduction mechanisms; [U] Intracellular trafficking, secretion, and vesicular transport; [V] Defense mechanisms; [X] No prediction. The ratio for each gene is >1.5 or <0.66, but >2 or <0.5 in at least one of the four conditions. The number 2177 represents the total number of amplicons analyzed, covering the entire genome of S. pneumoniae D39. This number is slightly higher than the number of genes in the genome of D39 (2069), since some amplicons were present in versions for different strains (i.e. TIGR4, R6, or D39) or in 2 copies.

Mentions: To determine the effect of the ccpA deletion on the transcriptome of S. pneumoniae, the transcriptional profile of D39 wild-type was compared to that of its isogenic ccpA deletion mutant at mid-exponential (M) and transition-to-stationary (TS) phases of growth. In the four conditions tested, Glc_M, Gal_M, Glc_TS, Gal_TS, between 14 and 19% of the ORFs in the genome were significantly, differentially expressed in the ccpA deletion mutant, of which more genes upregulated than downregulated (Fig. 1, see also Table S1 for an overview). This is similar to S. pyogenes (up to 20% genes affected) [8], but higher than S. mutans, L. lactis, and B. subtilis (less than 9% genes affected) [5]–[7]. These findings confirm the role of CcpA as a global regulator with a prevalent repressor function.


CcpA ensures optimal metabolic fitness of Streptococcus pneumoniae.

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

Genes differentially expressed due to inactivation of CcpA.A. Venn diagram of the genes significantly differentially transcribed due to loss of CcpA. Transcript levels in D39ΔccpA were compared to D39 wild-type in Glc_M, Gal_M, Glc_TS, Gal_TS as generated by VENNY (http://bioinfogp.cnb.csic.es/tools/venny/index.html). For all intersections, which are not drawn to scale, the numbers of genes are indicated. The total number of genes influenced in each condition tested was: Glc_M, 334 (15.3%), Glc_TS, 418 (19.2%), Gal_M, 299 (13.7%), and Gal_TS 299 (13.7%) in a universe of 2177 genes in the genome of S. pneumoniae. B. Numbers of genes significantly differently transcribed by the ccpA deletion in strain D39 ordered by COG categories. White bars represent M phase of growth, black bars TS phase of growth. [C] Energy production and conversion; [D] Cell cycle control, cell division; chromosome partitioning; [E] Amino acid transport and metabolism; [F] Nucleotide transport and metabolism; [G] Carbohydrate transport and metabolism; [H] Coenzyme transport and metabolism; [I] Lipid transport and metabolism; [J] Translation, ribosomal structure and biogenesis; [K] Transcription; [L] Replication, recombination and repair; [M] Cell wall/membrane/envelope biogenesis; [O] Posttranslational modification, protein turnover, chaperones; [P] Inorganic ion transport and metabolism; [Q] Secondary metabolites biosynthesis, transport and catabolism; [R] General function prediction only; [S] Function unknown; [T] Signal transduction mechanisms; [U] Intracellular trafficking, secretion, and vesicular transport; [V] Defense mechanisms; [X] No prediction. The ratio for each gene is >1.5 or <0.66, but >2 or <0.5 in at least one of the four conditions. The number 2177 represents the total number of amplicons analyzed, covering the entire genome of S. pneumoniae D39. This number is slightly higher than the number of genes in the genome of D39 (2069), since some amplicons were present in versions for different strains (i.e. TIGR4, R6, or D39) or in 2 copies.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198803&req=5

pone-0026707-g001: Genes differentially expressed due to inactivation of CcpA.A. Venn diagram of the genes significantly differentially transcribed due to loss of CcpA. Transcript levels in D39ΔccpA were compared to D39 wild-type in Glc_M, Gal_M, Glc_TS, Gal_TS as generated by VENNY (http://bioinfogp.cnb.csic.es/tools/venny/index.html). For all intersections, which are not drawn to scale, the numbers of genes are indicated. The total number of genes influenced in each condition tested was: Glc_M, 334 (15.3%), Glc_TS, 418 (19.2%), Gal_M, 299 (13.7%), and Gal_TS 299 (13.7%) in a universe of 2177 genes in the genome of S. pneumoniae. B. Numbers of genes significantly differently transcribed by the ccpA deletion in strain D39 ordered by COG categories. White bars represent M phase of growth, black bars TS phase of growth. [C] Energy production and conversion; [D] Cell cycle control, cell division; chromosome partitioning; [E] Amino acid transport and metabolism; [F] Nucleotide transport and metabolism; [G] Carbohydrate transport and metabolism; [H] Coenzyme transport and metabolism; [I] Lipid transport and metabolism; [J] Translation, ribosomal structure and biogenesis; [K] Transcription; [L] Replication, recombination and repair; [M] Cell wall/membrane/envelope biogenesis; [O] Posttranslational modification, protein turnover, chaperones; [P] Inorganic ion transport and metabolism; [Q] Secondary metabolites biosynthesis, transport and catabolism; [R] General function prediction only; [S] Function unknown; [T] Signal transduction mechanisms; [U] Intracellular trafficking, secretion, and vesicular transport; [V] Defense mechanisms; [X] No prediction. The ratio for each gene is >1.5 or <0.66, but >2 or <0.5 in at least one of the four conditions. The number 2177 represents the total number of amplicons analyzed, covering the entire genome of S. pneumoniae D39. This number is slightly higher than the number of genes in the genome of D39 (2069), since some amplicons were present in versions for different strains (i.e. TIGR4, R6, or D39) or in 2 copies.
Mentions: To determine the effect of the ccpA deletion on the transcriptome of S. pneumoniae, the transcriptional profile of D39 wild-type was compared to that of its isogenic ccpA deletion mutant at mid-exponential (M) and transition-to-stationary (TS) phases of growth. In the four conditions tested, Glc_M, Gal_M, Glc_TS, Gal_TS, between 14 and 19% of the ORFs in the genome were significantly, differentially expressed in the ccpA deletion mutant, of which more genes upregulated than downregulated (Fig. 1, see also Table S1 for an overview). This is similar to S. pyogenes (up to 20% genes affected) [8], but higher than S. mutans, L. lactis, and B. subtilis (less than 9% genes affected) [5]–[7]. These findings confirm the role of CcpA as a global regulator with a prevalent repressor function.

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