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1.
Mol Microbiol ; 89(2): 334-49, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23710838

ABSTRACT

The two-component regulatory system CiaRH of Streptococcus pneumoniae controls 25 genes, five of which specify homologous small non-coding csRNAs (cia-dependent small RNAs). The csRNAs were predicted to act regulatory as base-pairing sRNAs, but their targets have not been identified. By csRNA gene inactivations we established that the major phenotypes associated with a hyperactive CiaRH system, enhanced ß-lactam resistance and prevention of genetic competence, are dependent on the csRNAs. Computational target predictions and evaluations by translational fusions identified six genes to be under csRNA control: spr0081, spr0371, spr0159, spr0551, spr1097 and spr2043(comC). Measuring the effect of single csRNAs on three targets indicated that they acted additively. One of the targets, comC(spr2043), encoding the precursor of the competence stimulating pheromone CSP, constitutes a link of CiaRH to competence control. Partially disrupting predicted csRNA-comC complementarity led to strongly diminished repression by the csRNAs and to transformability in a strain with a hyperactive CiaRH. Thus, a hyperactive CiaRH system prevents competence development by csRNA-dependent post-transcriptional repression of CSP production. The csRNAs are also involved in competence regulation in the wild-type strain R6, but their activity is only apparent in the absence of the protease gene htrA, another CiaRH regulon member.


Subject(s)
Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Gene Expression Regulation, Bacterial , Protein Kinases/genetics , RNA, Small Untranslated/genetics , Bacterial Proteins/chemistry , Gene Targeting , Promoter Regions, Genetic , Protein Kinases/chemistry , Protein Kinases/metabolism , RNA, Bacterial/chemistry , RNA, Bacterial/genetics , RNA, Small Untranslated/chemistry , RNA, Small Untranslated/metabolism , Regulon , Streptococcus pneumoniae/drug effects , Streptococcus pneumoniae/genetics , Streptococcus pneumoniae/growth & development , beta-Lactam Resistance/genetics
2.
J Mol Microbiol Biotechnol ; 20(2): 96-104, 2011 Apr.
Article in English | MEDLINE | ID: mdl-21422763

ABSTRACT

The two-component regulatory system CiaRH of Streptococcus pneumoniae affects a variety of processes such as competence development, autolysis, bacteriocin production, host colonization, and virulence. While the targets of the regulator CiaR are known, the role of phosphorylation in CiaR regulation has not been defined. To address this issue, the presumed phosphorylation site of CiaR, aspartic acid at position 51, was replaced by alanine. The mutant CiaRD51A protein was no longer able to activate CiaR-dependent promoters, strongly suggesting that the phosphorylated form of CiaR is active in regulation. However, depending on the growth medium, inactivation of the kinase gene ciaH resulted in a subtle increase of CiaR-dependent promoter activities or in a strong reduction. Therefore, CiaH may act as a kinase or phosphatase and CiaR is apparently able to obtain its phosphate independently of CiaH. On the other hand, promoter measurements in cells with an intact CiaRH system demonstrated a high, nearly constitutive, expression level of the CiaR regulon independent from the growth medium. Thus, in contrast to many other two-component regulatory systems, CiaRH has apparently evolved to maintain high levels of gene expression under a variety of conditions rather than responding strongly to a signal.


Subject(s)
Bacterial Proteins/metabolism , Gene Expression Regulation, Bacterial , Protein Kinases/metabolism , Streptococcus pneumoniae/genetics , Streptococcus pneumoniae/metabolism , Amino Acid Substitution , Aspartic Acid/genetics , Aspartic Acid/metabolism , DNA, Bacterial/metabolism , Electrophoretic Mobility Shift Assay , Gene Expression Profiling , Genes, Reporter , Histidine Kinase , Mutagenesis, Site-Directed , Phosphorylation , Promoter Regions, Genetic , Protein Binding , Signal Transduction , Streptococcus pneumoniae/growth & development , Stress, Physiological , beta-Galactosidase/genetics , beta-Galactosidase/metabolism
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