An increase in negative supercoiling in bacteria reveals topology-reacting gene clusters and a homeostatic response mediated by the DNA topoisomerase I gene
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ABSTRACT:
ORGANISM(S): Streptococcus pneumoniae R6
SUBMITTER: Antonio Martin-Galiano
PROVIDER: E-GEOD-77749 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:We studied the response to increased DNA-supercoiling in Streptococcus pneumoniae by using seconeolitsine (SCN), a DNA topoisomerase I inhibitor. A homeostatic transcriptional response allowing recovering of supercoiling density was observed in cells treated with subinhibitory SCN concentrations. Supercoiling increases up to 40.7% (6 µM SCN) and 72.9% (8 µM SCN) were reverted to 8.5% and 44.1%, respectively. Likewise, recovery of viability and DNA-supercoiling were observed when cells were treated with those SCN concentrations and the drug was removed. The main DNA topoisomerase gene affected was topA, whose transcription depended on the supercoiling level. A two stage global transcriptomic response with 8 µM seconeolitsine was detected. The early stage (5 and 15 min, 10% of the genome) represented a response induced by increased supercoiling. The second stage represented supercoiling recovery (30 min, 2.0% of the genome). Almost 25% of the early responsive genes formed clusters with coordinated transcriptional regulation. Twelve clusters were evident, with sizes of 6.7 to 31.4 Kb (9 to 22 responsive genes). Strikingly, clusters did not contradict those observed under DNA relaxation, suggesting that bacteria manage supercoiling stress using overlapping responses. This is the first study describing a global transcriptomic response triggered by an increase in DNA supercoiling in bacteria. Total RNA was extracted from S. pneumoniae R6 cultures during exponential growth and used for RNA-Seq. Two independent replicates per condition were performed.
2016-07-07 | E-GEOD-77748 | biostudies-arrayexpress
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2016-08-24 | E-GEOD-85935 | biostudies-arrayexpress
Project description:We studied the response to increased DNA-supercoiling in Streptococcus pneumoniae by using seconeolitsine (SCN), a DNA topoisomerase I inhibitor. A homeostatic transcriptional response allowing recovering of supercoiling density was observed in cells treated with subinhibitory SCN concentrations. Supercoiling increases up to 40.7% (6 µM SCN) and 72.9% (8 µM SCN) were reverted to 8.5% and 44.1%, respectively. Likewise, recovery of viability and DNA-supercoiling were observed when cells were treated with those SCN concentrations and the drug was removed. The main DNA topoisomerase gene affected was topA, whose transcription depended on the supercoiling level. A two stage global transcriptomic response with 8 µM seconeolitsine was detected. The early stage (5 and 15 min, 10% of the genome) represented a response induced by increased supercoiling. The second stage represented supercoiling recovery (30 min, 2.0% of the genome). Almost 25% of the early responsive genes formed clusters with coordinated transcriptional regulation. Twelve clusters were evident, with sizes of 6.7 to 31.4 Kb (9 to 22 responsive genes). Strikingly, clusters did not contradict those observed under DNA relaxation, suggesting that bacteria manage supercoiling stress using overlapping responses. This is the first study describing a global transcriptomic response triggered by an increase in DNA supercoiling in bacteria. RNA was extracted from S. pneumoniae R6 cultures during exponential growth, amplified and used for microarray hibridization. Two independent replicates were performed.
2016-07-07 | E-GEOD-77587 | biostudies-arrayexpress
Project description:DNA methylation in CpG context is fundamental to the epigenetic regulation of gene expression in high eukaryotes. Disorganization of methylation status is implicated in many diseases, cellular differentiation, imprinting, and other biological processes. Techniques that enrich for biologically relevant CpG-rich genomic regions are desired since, depending on the size of an oragnism's methylome, the depth of sequencing required to cover all CpGs can be prohibitively expensive. Currently, restriction-enzyme based Reduced Representation Bisulfite Sequencing and its modified protocols are widely used to study methylation differences. Recently, Agilent Technologies and Roche NimbleGen have aimed to both reduce sequencing costs and capture CpGs of known biological relevance by marketing in-solution custom-capture hybridization platforms. These three methods target approximately 10-13% of the human methylome. For each platform - restriction-enzyme based enhanced reduced representation (ERRBS), capture based Agilent SureSelect Methyl-seq (SSMethylseq), and capture based Roche NimbleGen SeqCap Epi CpGiant (CpGiant) - we used human lung fibroblast cell line IMR90 DNA to make libraries according to each protocol and sequenced to equivalent depth. Overall, SSMethylSeq and CpGiant covered >95% of their designed capture regions whereas ERRBS covered 70% of its expected MspI regions. Methylation levels were concordant across the platforms. The concordance of annotations of CpG units for genomic features, displayed roughly the same proportions of genomic features. SSMethylSeq and CpGiant are most similar and cover marginally more annotated regions than ERRBS. However, the number of CpG units shared by all methods was low, ~26% of any platform. We conclude that captured based methods are largely consistent in terms of covered CpG loci although ERRBS provides comparable data at a significantly reduced price. Furthermore, library preparation for ERRBS can be performed with as little as 75ngs of starting material, whereas micrograms are needed for the capture hybridization techniques. Libraries were made from human lung fibroblast cell line IMR90 DNA for each protocol of ERRBS, Agilent SureSelect Methyl-seq, Roche NimbleGen SeqCap Epi CpGiant, and WGBS, then sequenced as paired-end 100bp on an Illumina HiSeq 2500.
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2016-07-21 | E-GEOD-84651 | biostudies-arrayexpress
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2014-11-13 | E-GEOD-61295 | biostudies-arrayexpress
Project description:We pursued the hypothesis that epigenetic regulators of transcription are involved in both the development and the progression of HPV-associated CIN. Using HELP-tagging, we performed the most comprehensive study to date of DNA methylation in HPV-associated cervical neoplasia, testing ~2 million loci throughout the human genome in biopsies from 78 HPV+ women, identifying changes starting in early CIN and maintained through carcinogenesis. We identified loci at which DNA methylation is consistently altered, beginning early in the course of neoplastic disease and progressing with disease advancement. DNA methylation profiles for cervical biopsies of 19 normals, 20 CIN1, 16 CIN2/3, and 23 cervical cancers.
2016-08-22 | E-GEOD-76986 | biostudies-arrayexpress
Project description:Background: Microorganisms are the major cause of food spoilage during storage, processing and distribution. Pseudomonas fluorescens is a typical spoilage bacterium that contributes to a large extent to the spoilage process of proteinaceous food. RpoS is considered an important global regulator involved in stress survival and virulence in many pathogens. Our previous work revealed that RpoS contributed to the spoilage activities of P. fluorescens by regulating resistance to different stress conditions, extracellular acylated homoserine lactone (AHL) levels, extracellular protease and total volatile basic nitrogen (TVB-N) production. However, RpoS-dependent genes in P. fluorescens remained undefined. Results: RNA-seq transcriptomics analysis combined with quantitative proteomics analysis basing on multiplexed isobaric tandem mass tag (TMT) labeling was performed for the P. fluorescens wild-type strain UK4 and its derivative carrying a rpoS mutation. A total of 375 differentially expressed genes (DEGs) and 212 differentially expressed proteins (DEPs) were identified in these two backgrounds. The DGEs were further verified by qRT-PCR tests, and the genes directly regulated by RpoS were confirmed by 5’-RACE-PCR sequencing. The combining transcriptome and proteome analysis revealed a role of this regulator in several cellular processes, including polysaccharide metabolism, intracellular secretion and extracellular structures, cell well biogenesis, stress responses, ammonia and biogenic amine production, which may contribute to biofilm formation, stress resistance and spoilage activities of P. fluorescens. Moreover, in this work we indeed observed that RpoS contributed to the production of the macrocolony biofilm’s matrix.
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