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Transcriptome analysis of the Bacillus subtilis ccpA topA(S478P) strain

ABSTRACT: In this study the isogenic Bacillus subtilis mutant strains ccpA topA+ and ccpA topA(S478P) were analyzed. The S478P suppressor mutation occurred when the ccpA mutant was grown on minimal medium supplemented with glucose and ammonium as single sources of carbon and nitrogen, respectively. Under these conditions, the ccpA mutant is unable to produce enough glutamate for growth, since gltAB, encoding the glutamate synthase, is not expressed. In order to get an insight how the S478P mutation in the DNA topoisomerase I affects expression on a global level, the two strains were subjected to a microarray analysis. Bacteria were cultivated in minimal medium supplemented with glucose and glutamate. The microarray data show that the topA(S478P) mutation results in a global re-direction of the central carbon metabolism that includes glutamate biosynthesis.

ORGANISM(S): Bacillus subtilis subsp. subtilis str. 168

PROVIDER: GSE88793 | GEO | 2017/11/01

SECONDARY ACCESSION(S): PRJNA348556

REPOSITORIES: GEO

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Project description:Background The catabolite control protein A (CcpA) is a member of the LacI/GalR family of transcriptional regulators controlling carbon-metabolism pathways in low-GC Gram positive bacteria. It functions as a catabolite repressor or activator, allowing the bacteria to utilize the preferred carbon source over secondary carbon sources. This study is the first CcpA-dependent transcriptome and proteome analysis in S. aureus wild type and ccpA-deleted mutant, focussing on short-time effects of glucose under stable pH conditions. Results The addition of glucose to exponentially growing S. aureus increased enzymes of glycolytic pathway, indicating a higher glycolytic activity, while proteins required for the complete oxidation in the TCA cycle were repressed via CcpA. Phosphotransacetylase and acetate kinase, converting acetylCoA to acetate with a concomitant substrate-level phosphorylation were neither regulated by glucose nor by CcpA. Most CcpA directly repressed genes were involved in utilization of amino acids as secondary carbon sources. More genes were found to be differentially expressed by CcpA in a glucose-independent manner than in the classical, glucose dependent way, suggesting that glucose-independent regulation by CcpA may be of particular importance in S. aureus. In the presence of glucose, CcpA was found to regulate expression of genes involved in metabolism, but that of genes coding for virulence determinants. Conclusions This study identified the CcpA regulon of exponentially growing S. aureus, for the first time. As in other bacteria, the CcpA-regulon of S. aureus comprised a large amount of metabolic genes but also some 50 genes associated with virulence. CcpA seemed to work in a glucose- as well as glucose-independent way. The transcriptomes of strain Newman and its isogenic ccpA-deleted mutant were determined in early exponential growth and 30 min after the addition of 10 mM glucose, under controlled pH conditions. In the absence of glucose, the wild type grew slightly faster than the mutant, reaching an OD600 of 1 approximately 20 min earlier than the mutant. Adding 10 mM glucose at OD600 1 increased the growth rate of the wild type but had only a minor effect on that of the mutant. 60 min after glucose addition, glucose was depleted down to 0.3 mM by the wild type, while still 3 mM glucose was left in the culture of the mutant. Despite increased glucose consumption rates in the wild type, acetate production was only slightly enhanced compared to the mutant. No lactate was excreted at any time point sampled. Acidification of the medium upon glucose metabolism was prevented by buffering, maintaining a pH of 7.5 for both strains and under both growth conditions for at least 2 h after glucose addition, allowing to rule out any pH effects.

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Transcriptome analysis of the Bacillus subtilis ccpA topA(S478P) strain

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