Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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RNA-seq of a Streptococcus agalactiae A909 mutant strain deleted for the pleiotropic transcriptional regulator CcpA

ABSTRACT: The commensal bacterium Streptococcus agalactiae is responsible for various infections in a wide variety of hosts including humans. Its broad spectrum of hosts shows its ability to acquire nutrients in variable conditions. The carbon catabolite repression allows bacteria to prioritize the uptake and the catabolism of the environmental sugars. In Gram-positive bacteria, CcpA (catabolite control protein A), a pleiotropic transcriptional regulator, plays a key role in catabolite repression. Studies have shown the involvement of carbon catabolite repression in the adaptation and stress resistance of pathogenic bacteria. The goal of this study is to determine the regulon and the role(s) of CcpA in the physiology and adaptation of S. agalactiae. To this aim, Streptococcus agalactiae strain A909 WT and its isogenic mutant ∆ccpA, obtained by allelic exchange were grown in filter-sterilized chemically defined medium (CDM) supplemented with 0,25% or 1% (w/v) of glucose. Their transcriptomes were compared under these two conditions by using RNA-seq.

INSTRUMENT(S): Illumina HiSeq 2000

ORGANISM(S): Streptococcus agalactiae

SUBMITTER: Isabelle Rosinski-Chupin

PROVIDER: E-MTAB-11639 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:In Bacillus subtilis and its relatives carbon catabolite control, a mechanism enabling to reach maximal efficiency of carbon and energy sources metabolism, is achieved by the global regulator CcpA (carbon catabolite protein A). CcpA in a complex with HPr-Ser-P (seryl-phosphorylated form of histidine-containing protein, HPr) binds to operator sites called catabolite responsive elements, cre. Depending on the cre box position relative to the promoter, the CcpA/HPr-Ser-P complex can either act as a positive or a negative regulator. The cre boxes are highly degenerate semi-palindromes with a lowly conserved consensus sequence. So far, studies aimed at revealing how CcpA can bind such diverse sites were focused on the analysis of single cre boxes. In this study, a genome-wide analysis of cre sites was performed in order to identify differences in cre sequence and position, which determine their binding affinity. The transcriptomes of B. subtilis cultures with three different CcpA expression levels were compared. The higher the amount of CcpA in the cells, the more operons possessing cre sites were differentially regulated. The cre boxes that mediated regulation at low CcpA levels were designated as strong (high affinity) and those which responded only to high amounts of CcpA, as weak (low affinity). Differences in the sequence and position in relation to the transcription start site between strong and weak cre boxes were revealed. Certain residues at specific positions in the cre box as well as, to a certain extent, a more palindromic nature of cre sequences and the location of cre in close vicinity to the transcription start site contribute to the strength of CcpA-dependent regulation. The main factors contributing to cre regulatory efficiencies, enabling subtle differential control of various subregulons of the CcpA regulon, are identified. Bacillus subtilis strain MP902 [strain MP901 (Ptet-ccpA, KmR) carrying pWH119 (Pxyl-tetR, EmR)] was grown in presence of three different concentrations of Ptet inducer, anhydrotetracycline (ATc), leading to three levels of ccpA expression induction. The control culture was grown in absence of ATc. The total RNA for transcriptome analyses was isolated at OD600 = 0.8 from 16 ml culture. Three independent cultures of each strain (target strains and controls) were used, and cells were sampled for microarray experiment.

Dataset Information

RNA-seq of a Streptococcus agalactiae A909 mutant strain deleted for the pleiotropic transcriptional regulator CcpA

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