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1.  A Non-Linear Deterministic Model for Regulation of Diauxic Lag on Cellobiose by the Pneumococcal Multidomain Transcriptional Regulator CelR 
PLoS ONE  2012;7(10):e47393.
When grown on glucose and beta-glucosides, S. pneumoniae shows sequential use of sugars resulting in diauxic growth with variable time extent of the lag phase separating the biphasic growth curve. The pneumococcal beta-glucoside uptake locus containing the PTS transporter spr0276-82, is regulated by a multi-domain transcriptional regulator CelR. In this work, we address the contribution of phosphorylation of the phosphorylable cysteine in the EIIB domain of CelR to diauxic lag. Utilising site-directed mutagenesis of the phosphorylable amino acids in the EIIB and EIIA domains of CelR, we show that the EIIB domain activation is linked to the duration of the lag phase. Analysis of mutants for other PTS systems indicates that a second beta-glucoside PTS (spr0505), not able to support growth on cellobiose, is responsible for the lag during diauxic growth. A mathematical model of the process is devised together with a nonlinear identification procedure which provides model parameter estimates characterizing the single phases of bacterial growth. Parameter identification performed on data recorded in appropriate experiments on mutants allows for establishing a relationship between a specific model parameter, the EIIB domain and the time extent of the diauxic lag. The experimental results and the related insights provided by the mathematical model provide evidence that the conflicting activation of the CelR regulator is at the origin of the lag phase during sequential growth on glucose and cellobiose. This data is the first description of diauxic lag regulation involving two PTS and a multidomain regulator and could serve as a promising approach for studying the S. pneumoniae growth process on complex carbon sources as possibly encountered in the human host.
doi:10.1371/journal.pone.0047393
PMCID: PMC3478281  PMID: 23110070
2.  Regulation of neuraminidase expression in Streptococcus pneumoniae 
BMC Microbiology  2012;12:200.
Background
Sialic acid (N-acetylneuraminic acid; NeuNAc) is one of the most important carbohydrates for Streptococcus pneumoniae due of its role as a carbon and energy source, receptor for adhesion and invasion and molecular signal for promotion of biofilm formation, nasopharyngeal carriage and invasion of the lung.
Results
In this work, NeuNAc and its metabolic derivative N-acetyl mannosamine (ManNAc) were used to analyze regulatory mechanisms of the neuraminidase locus expression. Genomic and metabolic comparison to Streptococcus mitis, Streptococcus oralis, Streptococcus gordonii and Streptococcus sanguinis elucidates the metabolic association of the two amino sugars to different parts of the locus coding for the two main pneumococcal neuraminidases and confirms the substrate specificity of the respective ABC transporters. Quantitative gene expression analysis shows repression of the locus by glucose and induction of all predicted transcriptional units by ManNAc and NeuNAc, each inducing with higher efficiency the operon encoding for the transporter with higher specificity for the respective amino sugar. Cytofluorimetric analysis demonstrated enhanced surface exposure of NanA on pneumococci grown in NeuNAc and ManNAc and an activity assay allowed to quantify approximately twelve times as much neuraminidase activity on induced cells as opposed to glucose grown cells.
Conclusions
The present data increase the understanding of metabolic regulation of the nanAB locus and indicate that experiments aimed at the elucidation of the relevance of neuraminidases in pneumococcal virulence should possibly not be carried out on bacteria grown in glucose containing media.
doi:10.1186/1471-2180-12-200
PMCID: PMC3509027  PMID: 22963456
Sialic acid; Metabolic regulation; Carbon catabolite repression
3.  The impact of the competence quorum sensing system on Streptococcus pneumoniae biofilms varies depending on the experimental model 
BMC Microbiology  2011;11:75.
Background
Different models for biofilm in Streptococcus pneumoniae have been described in literature. To permit comparison of experimental data, we characterised the impact of the pneumococcal quorum-sensing competence system on biofilm formation in three models. For this scope, we used two microtiter and one continuous culture biofilm system.
Results
In both microtiter models the competence system influences stability and structure of biofilm in the late attachment phase and synthetic competence stimulating peptide (CSP) restored wild type phenotypes in the comC mutants unable to produce the peptide. Early attachment of single cells to well bottoms was found for both systems to be competence independent, while later phases, including microcolony formation correlated to an intact competence system. The continuous culture biofilm model was not affected by mutations in the competence locus, but deletion of capsule had a significant impact in this model.
Conclusions
Since biofilm remains a largely uncharacterised multi-parameter phenotype it appears to be advisable to exploit more than one model in order to draw conclusion of possible relevance of specific genotypes on pneumococcal physiology.
doi:10.1186/1471-2180-11-75
PMCID: PMC3098770  PMID: 21492426
4.  Biofilm Formation-Gene Expression Relay System in Escherichia coli: Modulation of σS-Dependent Gene Expression by the CsgD Regulatory Protein via σS Protein Stabilization▿ † 
Journal of Bacteriology  2007;189(22):8034-8043.
Bacteria can switch from a single-cell (planktonic) mode to a multicellular community (biofilm) mode via production of cell-cell aggregation and surface adhesion factors. In this report, we present evidence that the CsgD protein, a transcription regulator involved in biofilm formation in Escherichia coli, modulates the expression of the rpoS (σS) regulon. Protein pattern analysis of E. coli cells in stationary phase shows that CsgD affects the expression of several proteins encoded by σS-dependent genes. CsgD regulation of σS-dependent genes takes place at gene transcription level, does not bypass the need for rpoS, and is abolished in an rpoS-null mutant. Consistent with these results, we find that CsgD expression leads to an increase in σS intracellular concentration. Increase in σS cellular amount is mediated by CsgD-dependent transcription activation of iraP, encoding a factor involved in σS protein stabilization. Our results strongly suggest that the CsgD regulatory protein plays a major role as a relay between adhesion factors production and σS-dependent gene expression via σS protein stabilization. Direct coordination between biofilm formation and expression of the rpoS regulon could positively impact important biological processes, such as host colonization or response to environmental stresses.
doi:10.1128/JB.00900-07
PMCID: PMC2168689  PMID: 17873038

Results 1-4 (4)