Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Promoter proximal pausing in response to stress in serum deprived mouse fibroblasts.

ABSTRACT: We have employed short-capped RNA sequencing (sc-RNA-seq) in order to identify genes whose expression is regulated by promoter proximal pausing of RNA Polymerase II (RNAPI) in response to stress stimulation. We used serum-deprived mouse Swiss 3T3 fibroblasts, either untreated (control) or treated with anisomycin to induce the p38/MAP kinase pathway. Serum starved (72 h 0.2% FCS) mouse 3T3 cells were treated with anisomycin (188.5 nM) for 1 h (in duplicates). Untreated, serum-starved cells were used as a control. We isolated nuclear RNA, performed size fractionation followed by isolation of short-capped RNAs (scRNA). scRNAs were subsequently converted into DNA library and sequenced.

ORGANISM(S): Mus musculus

SUBMITTER: Anna Sawicka

PROVIDER: E-GEOD-55775 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

H3S28 phosphorylation is a hallmark of the transcriptional response to cellular stress.

Sawicka Anna A Hartl Dominik D Goiser Malgorzata M Pusch Oliver O Stocsits Roman R RR Tamir Ido M IM Mechtler Karl K Seiser Christian C

Genome research 20140818 11

The selectivity of transcriptional responses to extracellular cues is reflected by the deposition of stimulus-specific chromatin marks. Although histone H3 phosphorylation is a target of numerous signaling pathways, its role in transcriptional regulation remains poorly understood. Here, for the first time, we report a genome-wide analysis of H3S28 phosphorylation in a mammalian system in the context of stress signaling. We found that this mark targets as many as 50% of all stress-induced genes, ...[more]

PMID: 25135956

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Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs. Examination of the effect of ADAR2 on mature miRNA abundance and sequence in adult mouse brain.

Project description:MicroRNAs (miRNAs) are a class of non-coding RNA molecules which have significant gene regulation roles in organisms. The advent of new high throughput sequencing technologies has enabled the revelation of novel miRNAs. Although there are two recent reports on high throughput sequencing analysis of small RNA libraries from different organs of two grapevine wine varieties, there were significant divergence in the number and kinds of miRNAs sequenced in these studies. More sequencing of small RNA libraries is still important for the discovery of novel miRNAs in grapevine. Here, we initially constructed a small RNA library of flower and fruit tissues of a table grapevine cultivar M-bM-^@M-^XSummer BlackM-bM-^@M-^Y and performed sequencing and analysis of sRNAs using the Illumina Solexa platform, expecting to discover more miRNAs related to the development of grapevine flowers and berries and the formation of dessert quality in grapevine berries. Totally, 130 conserved grapevine miRNA (Vv-miRNA) belonging to 28 Vv-miRNA families were validated, and 92 novel potential grapevine-specific ones representing 80 unique ones were first discovered. Forty-two (48.84%) of the novel miRNAs possessed differential semi-quantitative PCR expression profiles in various grapevine tissues that could further confirm their existence in the grapevine, among which twenty were expressed only in grapevine berries, indicating some fruit-specificity. 130 target genes for 46 novel miRNAs could be predicted. The locations of these potential target genes on grapevine chromosomes and their complementary levels with the corresponding miRNAs were also analyzed. Size fractionated small RNAs (16-30 bp) from total RNA extracts was ligated to 5' and 3' adapters, and reverse transcribed. After PCR amplification the sample was subjected to Solexa sequencing. The resultant 35nt sequence data were filtered according to base quality value. The remained sequences were used to trim 5' and 3' adaptors. The clean tags were used for further analysis.

Dataset Information

Promoter proximal pausing in response to stress in serum deprived mouse fibroblasts.

Publications

H3S28 phosphorylation is a hallmark of the transcriptional response to cellular stress.

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