Dataset Information

Identification of ColR binding consensus and prediction of regulon of ColRS two-component system.

ABSTRACT:

Background

Conserved two-component system ColRS of Pseudomonas genus has been implicated in several unrelated phenotypes. For instance, deficiency of P. putida ColRS system results in lowered phenol tolerance, hindrance of transposition of Tn4652 and lysis of a subpopulation of glucose-grown bacteria. In order to discover molecular mechanisms behind these phenotypes, we focused here on identification of downstream components of ColRS signal transduction pathway.

Results

First, highly similar ColR binding sites were mapped upstream of outer membrane protein-encoding oprQ and a putative methyltransferase-encoding PP0903. These two ColR binding sequences were used as an input in computational genome-wide screening for new potential ColR recognition boxes upstream of different genes in P. putida. Biological relevance of a set of in silico predicted ColR-binding sites was analysed in vivo by studying the effect of ColR on transcription from promoters carrying these sites. This analysis disclosed seven novel genes of which six were positively and one negatively regulated by ColR. Interestingly, all promoters tested responded more significantly to the over-expression than to the absence of ColR suggesting that either ColR is limiting or ColS-activating signal is low under the conditions applied. The binding sites of ColR in the promoters analysed were validated by gel mobility shift and/or DNase I footprinting assays. ColR binding consensus was defined according to seven ColR binding motifs mapped by DNase I protection assay and this consensus was used to predict minimal regulon of ColRS system.

Conclusion

Combined usage of experimental and computational approach enabled us to define the binding consensus for response regulator ColR and to discover several new ColR-regulated genes. For instance, genes of outer membrane lipid A 3-O-deacylase PagL and cytoplasmic membrane diacylglycerol kinase DgkA are the members of ColR regulon. Furthermore, over 40 genes were predicted to be putatively controlled by ColRS two-component system in P. putida. It is notable that many of ColR-regulated genes encode membrane-related products thus confirming the previously proposed role of ColRS system in regulation of membrane functionality.

SUBMITTER: Kivistik PA

PROVIDER: S-EPMC2689224 | biostudies-literature | 2009 May

REPOSITORIES: biostudies-literature

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Publications

Identification of ColR binding consensus and prediction of regulon of ColRS two-component system.

Kivistik Paula A PA Kivi Rait R Kivisaar Maia M Hõrak Rita R

BMC molecular biology 20090516

<h4>Background</h4>Conserved two-component system ColRS of Pseudomonas genus has been implicated in several unrelated phenotypes. For instance, deficiency of P. putida ColRS system results in lowered phenol tolerance, hindrance of transposition of Tn4652 and lysis of a subpopulation of glucose-grown bacteria. In order to discover molecular mechanisms behind these phenotypes, we focused here on identification of downstream components of ColRS signal transduction pathway.<h4>Results</h4>First, hig ...[more]

PMID: 19445690

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Project description:BackgroundThe S. mutans LrgA/B holin-like proteins have been shown to affect biofilm formation and oxidative stress tolerance, and are regulated by oxygenation, glucose levels, and by the LytST two-component system. In this study, we sought to determine if LytST was involved in regulating lrgAB expression in response to glucose and oxygenation in S. mutans.ResultsReal-time PCR revealed that growth phase-dependent regulation of lrgAB expression in response to glucose metabolism is mediated by LytST under low-oxygen conditions. However, the effect of LytST on lrgAB expression was less pronounced when cells were grown with aeration. RNA expression profiles in the wild-type and lytS mutant strains were compared using microarrays in early exponential and late exponential phase cells. The expression of 40 and 136 genes in early-exponential and late exponential phase, respectively, was altered in the lytS mutant. Although expression of comYB, encoding a DNA binding-uptake protein, was substantially increased in the lytS mutant, this did not translate to an effect on competence. However, a lrgA mutant displayed a substantial decrease in transformation efficiency, suggestive of a previously-unknown link between LrgA and S. mutans competence development. Finally, increased expression of genes encoding antioxidant and DNA recombination/repair enzymes was observed in the lytS mutant, suggesting that the mutant may be subjected to increased oxidative stress during normal growth. Although the intracellular levels of reaction oxygen species (ROS) appeared similar between wild-type and lytS mutant strains after overnight growth, challenge of these strains with hydrogen peroxide (H2O2) resulted in increased intracellular ROS in the lytS mutant.ConclusionsOverall, these results: (1) Reinforce the importance of LytST in governing lrgAB expression in response to glucose and oxygen, (2) Define a new role for LytST in global gene regulation and resistance to H2O2, and (3) Uncover a potential link between LrgAB and competence development in S. mutans.

Dataset Information

Identification of ColR binding consensus and prediction of regulon of ColRS two-component system.

Background

Results

Conclusion

Publications

Identification of ColR binding consensus and prediction of regulon of ColRS two-component system.

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