Synechocystis PrhaBAD Inducible Promoter
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PROVIDER: PRJEB87132 | ENA |
REPOSITORIES: ENA
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Project description:The small protein AtpΘ (encoded by gene atpT) identified in cyanobacteria becomes maximum expressed during low-energy conditions such as during darkness. In the model cyanobacterium Synechocystis sp. PCC 6803 (Synechocystis 6803), AtpΘ was demonstrated to inhibit the ATP hydrolysis activity of the F0F1 ATP synthase. Here, the regulation of atpT gene expression was studied in greater detail. The darkness-induced activation of the atpT promoter indicated the existence of regulatory factors. To identify such factors, DNA-protein affinity precipitation was performed using biotinylated DNA fragments. These fragments contained either the promoter-5’UTR (PatpT66-5’UTR) as bait or, as negative control, only the 5’UTR (atpT 5’UTR). Each DNA fragment was incubated with total protein samples isolated from exponential phase Synechocystis 6803 which had been kept in the light or 12 h in darkness prior to protein preparation. The resulting protein samples were then analyzed by mass spectrometry.
2022-04-14 | PXD027801 | Pride
Project description:Comparison of gene expression of WT Synechocystis cells with cells overexpressing the sRNA PsrR1 in order to detect direct targets of the sRNA PsrR1. We monitored gene expression of an Synechocystis PCC6083 overexpressor strain (psrR1+) (Mitschke et al., 2011) harboring a self-replicating plasmid from which PsrR1 is transcribed under control of the copper responsive PpetJ promoter and a control strain (WT) harboring an empty plasmid 0h and 24 h after copper depletion. For the timepoint (0h) we sampled biological replicates and for timepoint (24h) we sampled biological triplicates.
2012-10-10 | E-GEOD-41411 | biostudies-arrayexpress
Project description:The fission yeast Schizosaccharomyces pombe lacks a diverse toolkit of inducible promoters for experimental manipulation. Available inducible promoters suffer from slow induction kinetics, limited control of expression levels and/or a requirement for defined growth medium. In particular, no S. pombe inducible promoter systems exhibit a linear dose response, which would allow expression to be tuned to specific levels. We have adapted a fast, orthogonal promoter system with a large dynamic range and a linear dose response, based on _-estradiol-regulated function of the human estrogen receptor, for use in S. pombe. We show that this promoter system, termed Z3EV, turns on quickly, can reach a maximal induction of 20 fold, and exhibits a linear dose response over its entire induction range, with few off target effects. We demonstrate the utility of this system by regulating the mitotic inhibitor Wee1 to create a strain in which cell size is regulated by _-estradiol concentration. This promoter system will be of great utility for experimentally regulating gene expression in fission yeast.
2016-12-13 | GSE92246 | GEO
Project description:Cyanobacteria, photoautotrophic prokaryotes, contribute significantly to the global photosynthesis and require large amounts of the essential micronutrient iron in order to maintain their Fe-rich photosynthetic apparatus. Here we use the model organism Synechocystis sp. PCC 6803 (later referred to Synechocystis) in both, standard and iron stress conditions, to study transcription and post-transcription regulation in iron deprivation. Although iron is one of the most abundant metals on earth, it is not soluble under aerobic conditions. Thus Synechocystis had to find ways to overcome iron deficiency. At the same time, however, free intracellular iron needs to be kept at permissive levels, as it becomes toxic under aerobic conditions by producing reactive oxygen species. For these reasons, complex regulatory networks have evolved to tightly control intracellular iron concentrations, ensuring its essential function yet avoiding cellular damage (Pierre Cornelis et al). In a previous study, we investigated iron deprivation in Synechocystis using customised amplification library for the analysis of global gene expression in the unicellular cyanobacterium (Hernández-Prieto et al, 2012; Georg et al, 2017),(Georg et al, 2017) but it is still little known about RNA stability in this organism. We now extend this study through a transcriptome wide half-life analysis in Synechocystis grown under standard and iron-limiting conditions using oligonucleotide microarrays that detect both protein-coding and non-coding transcripts (ncRNA). We used the antibiotic Rifampicin to stop the transcription. Samples were taken at time points 0 min (before the addition of rifampicin) and in a time series of 2 min, 4 min, 8 min, 16 min, 32 min and 64 min after its addition.
2022-07-30 | GSE209879 | GEO
Project description:In cyanobacteria DNA supercoiling varies over the diurnal light/dark cycle and is integrated with temporal programs of transcription and replication. We manipulated DNA supercoiling in Synechocystis sp. PCC 6803 by CRISPRi-based knockdown of gyrase subunits gyrA, gyrB and overexpression of topoisomerase I (TopoI) topA and analyzed the transcriptional response to gyrase knock-downs (endpoint in triplicate) and topoisomerase I overexpression (endpoint in triplicate, and 19 time points time series before and after induction) in Synechocystis sp. PCC 6803 via RNA-seq of coding RNA. In detail, Illumina Ribo-Zero Plus rRNA Depletion Kit was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was evaluated with the RNA Pico 6000 kit on the Agilent 2100 Bioanalyzer. RNA was free of detectable rRNA. Preparation of cDNA libraries was performed according to the manufacturer’s instructions for the TruSeq stranded mRNA kit (Illumina, San Diego, CA, United States). Subsequently, each cDNA library was sequenced on an Illumina NextSeq 500 system (2 x 75 nt PE high output v2.5).
2021-12-06 | E-MTAB-10949 | biostudies-arrayexpress
Project description:The transcriptome approach was used to characterize the global change (2 Fe concentrations) in Synechocystis gene expression in response to continuous iron starvation, compared to standard medium (17 microM). For iron depletion, exponentially growing cells were washed in Fe-free medium, and grown for 48 hrs under standard conditions in liquid medium cotaining either 1 mM or 2 mM of ferric ammonium citrate. Then, cells were washed, and resuspended in Fe-free medium (Fe contaminations 0.5 microM) and grown for another 48 hrs period. For each concentration point, total RNA were isolated from stressed and unstressed cells, reverse-transcribed, differentially labelled (dye swapped), hybridized together (stressed versus unstressed samples) and analyzed with DNA glass microarrays (two slides per each concentration point) (Custom-commercial array : IntelliGeneTM CyanoCHIP version 2.0, TAKARA). To identify differentially expressed genes, the median of the normalized ratio of Cy5/Cy3 intensity was calculated for each spot of the replicated dye-swap. The results of the analysis were carefully examined to exclude the dye effect between the 2 Cy-swapped arrays. Keywords: stress responses
2005-12-07 | GSE3717 | GEO
Project description:Cyanobacteria are oxygenic photoautotrophs responsible for a substantial proportion of nitrogen fixation and primary production in the hydrosphere. Non-nitrogen fixing cyanobacteria, such as Synechocystis sp. PCC 6803, depend of the availability of nitrogenized species to survive. Therefore, an intricate regulatory network around the transcriptional factor NtcA maintains the homeostasis of nitrogen in these organisms. The mechanisms controlling NtcA activity are well understood but a comprehensive study of its regulon is missing in Synechocystis. To define NtcA regulon during the early stage of nitrogen starvation, we have performed chromatin immunoprecipitation followed by sequencing (ChiP-seq), in parallel with genome level transcriptome analysis (RNA-seq). By combining both methods we assigned 51 activated and 28 repressed genes directly by NtcA. Most of direct targets included genes involved in nitrogen and carbon metabolism and photosynthesis. NtcA regulon also included 8 ncRNAs, of which ncr0710, Syr6 and NsiR7 were experimentally validated. Intriguingly, we identified several NtcA intragenic binding sites suggesting that NtcA can modulate transcriptional expression by binding along the whole transcript and not only in the promoter region as previously though. Finally, the transcriptional implication of PipX was analyzed in some NtcA-targets genes, revealing that PipX assists NtcA in the global nitrogen regulation in Synechocystis.
2017-10-01 | GSE97282 | GEO
Project description:Synechocystis Raw sequence reads
| PRJNA431100 | ENA