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Cellobiose-mediated gene expression in Streptococcus pneumoniae: a repressor function of the novel GntR-type regulator BguR.

Shafeeq S, Kuipers OP, Kloosterman TG - PLoS ONE (2013)

Bottom Line: Its expression was increased in the presence of cellobiose, and decreased in the presence of glucose.A novel GntR-type transcriptional regulator (which we here denote as BguR) was shown to act as a transcriptional repressor of the bgu operon and its repressive effect was relieved in the presence of cellobiose.BguR-dependent repression was demonstrated to be mediated by a 20-bp DNA operator site (5'-AAAAATGTCTAGACAAATTT-3') present in PbguA, as verified by promoter truncation experiments.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.

ABSTRACT
The human pathogen Streptococcus pneumoniae has the ability to use the carbon- and energy source cellobiose due to the presence of a cellobiose-utilizing gene cluster (cel locus) in its genome. This system is regulated by the cellobiose-dependent transcriptional activator CelR, which has been previously shown to contribute to pneumococcal virulence. To get a broader understanding of the response of S. pneumoniae to cellobiose, we compared the pneumococcal transcriptome during growth on glucose as the main carbon source to that with cellobiose as the main carbon source. The expression of various carbon metabolic genes was altered, including a PTS operon (which we here denote as the bgu operon) that has high similarity with the cel locus. In contrast to the cel locus, the bgu operon is conserved in all sequenced strains of S. pneumoniae, indicating an important physiological function in the lifestyle of pneumococci. We next characterized the transcriptional regulation of the bgu operon in more detail. Its expression was increased in the presence of cellobiose, and decreased in the presence of glucose. A novel GntR-type transcriptional regulator (which we here denote as BguR) was shown to act as a transcriptional repressor of the bgu operon and its repressive effect was relieved in the presence of cellobiose. BguR-dependent repression was demonstrated to be mediated by a 20-bp DNA operator site (5'-AAAAATGTCTAGACAAATTT-3') present in PbguA, as verified by promoter truncation experiments. In conclusion, we have identified a new cellobiose-responsive PTS operon, together with its transcriptional regulator in S. pneumoniae.

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Analysis of truncations of PbguA.A schematic overview of the bguA promoter truncations is shown. The table on the right gives the specific β-galactosidase activity of the truncated promoters in GM17 (0.5% glucose+M17) and CM17 (0.5% cellobiose+M17). Standard deviation is given in parentheses. The oval indicates the position of the putative BguR operator site, while the sequence of the BguR operator site is given above.
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pone-0057586-g003: Analysis of truncations of PbguA.A schematic overview of the bguA promoter truncations is shown. The table on the right gives the specific β-galactosidase activity of the truncated promoters in GM17 (0.5% glucose+M17) and CM17 (0.5% cellobiose+M17). Standard deviation is given in parentheses. The oval indicates the position of the putative BguR operator site, while the sequence of the BguR operator site is given above.

Mentions: The data presented above strongly suggest a direct effect of BguR on PbguA. To identify a possible BguR operator sequence, a 5′ promoter truncation study was performed with the bguA promoter. A diagram of the bguA promoter truncation is shown in Fig. 3. Truncation of PbguA near to the predicted -35 core promoter sequence (PbguA-5.4) relieved the repressive action of BguR on PbguA, suggesting the presence of a putative BguR operator in this deleted region of the promoter. Further bioinformatics analysis of this area revealed the presence of a 20-bp palindromic region (5′-AAAAATGTCTAGACAAATTT-3′) that is overlapping with the -35 site and that might act as the BguR operator site. Deletion of half of this predicted operator site (PbguA-5.5) led to high expression of PbguA in CM17 (0.5% Cellobiose+M17) and GM17 (0.5% Glucose+M17) medium in the wild-type (Fig. 3). However, when the PbguA was truncated only a few base pairs upstream of the predicted operator site (PbguA-5.3), expression was similar to that of the full-length promoter (Fig. 3). Therefore, these data suggest that the predicted operator site is functional and acts as the BguR operator site.


Cellobiose-mediated gene expression in Streptococcus pneumoniae: a repressor function of the novel GntR-type regulator BguR.

Shafeeq S, Kuipers OP, Kloosterman TG - PLoS ONE (2013)

Analysis of truncations of PbguA.A schematic overview of the bguA promoter truncations is shown. The table on the right gives the specific β-galactosidase activity of the truncated promoters in GM17 (0.5% glucose+M17) and CM17 (0.5% cellobiose+M17). Standard deviation is given in parentheses. The oval indicates the position of the putative BguR operator site, while the sequence of the BguR operator site is given above.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585215&req=5

pone-0057586-g003: Analysis of truncations of PbguA.A schematic overview of the bguA promoter truncations is shown. The table on the right gives the specific β-galactosidase activity of the truncated promoters in GM17 (0.5% glucose+M17) and CM17 (0.5% cellobiose+M17). Standard deviation is given in parentheses. The oval indicates the position of the putative BguR operator site, while the sequence of the BguR operator site is given above.
Mentions: The data presented above strongly suggest a direct effect of BguR on PbguA. To identify a possible BguR operator sequence, a 5′ promoter truncation study was performed with the bguA promoter. A diagram of the bguA promoter truncation is shown in Fig. 3. Truncation of PbguA near to the predicted -35 core promoter sequence (PbguA-5.4) relieved the repressive action of BguR on PbguA, suggesting the presence of a putative BguR operator in this deleted region of the promoter. Further bioinformatics analysis of this area revealed the presence of a 20-bp palindromic region (5′-AAAAATGTCTAGACAAATTT-3′) that is overlapping with the -35 site and that might act as the BguR operator site. Deletion of half of this predicted operator site (PbguA-5.5) led to high expression of PbguA in CM17 (0.5% Cellobiose+M17) and GM17 (0.5% Glucose+M17) medium in the wild-type (Fig. 3). However, when the PbguA was truncated only a few base pairs upstream of the predicted operator site (PbguA-5.3), expression was similar to that of the full-length promoter (Fig. 3). Therefore, these data suggest that the predicted operator site is functional and acts as the BguR operator site.

Bottom Line: Its expression was increased in the presence of cellobiose, and decreased in the presence of glucose.A novel GntR-type transcriptional regulator (which we here denote as BguR) was shown to act as a transcriptional repressor of the bgu operon and its repressive effect was relieved in the presence of cellobiose.BguR-dependent repression was demonstrated to be mediated by a 20-bp DNA operator site (5'-AAAAATGTCTAGACAAATTT-3') present in PbguA, as verified by promoter truncation experiments.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.

ABSTRACT
The human pathogen Streptococcus pneumoniae has the ability to use the carbon- and energy source cellobiose due to the presence of a cellobiose-utilizing gene cluster (cel locus) in its genome. This system is regulated by the cellobiose-dependent transcriptional activator CelR, which has been previously shown to contribute to pneumococcal virulence. To get a broader understanding of the response of S. pneumoniae to cellobiose, we compared the pneumococcal transcriptome during growth on glucose as the main carbon source to that with cellobiose as the main carbon source. The expression of various carbon metabolic genes was altered, including a PTS operon (which we here denote as the bgu operon) that has high similarity with the cel locus. In contrast to the cel locus, the bgu operon is conserved in all sequenced strains of S. pneumoniae, indicating an important physiological function in the lifestyle of pneumococci. We next characterized the transcriptional regulation of the bgu operon in more detail. Its expression was increased in the presence of cellobiose, and decreased in the presence of glucose. A novel GntR-type transcriptional regulator (which we here denote as BguR) was shown to act as a transcriptional repressor of the bgu operon and its repressive effect was relieved in the presence of cellobiose. BguR-dependent repression was demonstrated to be mediated by a 20-bp DNA operator site (5'-AAAAATGTCTAGACAAATTT-3') present in PbguA, as verified by promoter truncation experiments. In conclusion, we have identified a new cellobiose-responsive PTS operon, together with its transcriptional regulator in S. pneumoniae.

Show MeSH
Related in: MedlinePlus