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Functional coordination and HuR-mediated regulation of mRNA stability during T cell activation

ABSTRACT: We evaluated changes in mRNA stability and transcription using 4sU metabolic pulse labeling across a four hour time course following activation of Jurkat T cells with PMA and PHA

ORGANISM(S): Homo sapiens

PROVIDER: GSE71382 | GEO | 2015/10/28

SECONDARY ACCESSION(S): PRJNA291072

REPOSITORIES: GEO

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Functional coordination and HuR-mediated regulation of mRNA stability during T cell activation

Project description:We evaluated changes in mRNA stability and transcription using 4sU metabolic pulse labeling across a four hour time course following activation of Jurkat T cells with PMA and PHA Measurement of total mRNA (T) and 4sU labeled mRNA (IP) in three biological replicates at five time points: prior to activation (U) and the first four hours after activation (1-4)

2015-10-28 | E-GEOD-71382 | biostudies-arrayexpress

Thiouracil Pulse-chase, Tachyzoites and Bradyzoites mRNA stability data

Project description:Pulse chase measurements using thiouracil (DTU) labeling via UPRT and chasing with uracil Data from tachyzoites is labeled "DTU Pulse Chase". Two independent pulse chase experiments were performed in tachyzoites, pulse chase 1 and 2. Duplicate arrays at each timepoint were performed for pulse chase 2 (2 a and b). Data from bradyzoites are labeled "DTU Bradyzoite Pulse Chase". Two independent pulse chase experiments were performed in bradyzoites and a single set of arrays were performed for each experiment. Just one chase timepoint was used in the bradyzoite experiments, the 2 hour chase. An RNA stablity experiment design type examines stability and/or decay of RNA transcripts. Keywords: RNA_stability_design

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Thiouracil Pulse-chase, Tachyzoites and Bradyzoites mRNA stability data

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Alternative polyadenylation isoform abundance and stability of HEK293T cells

Project description:mRNA isoform abundance and stability in HEK293T cells were examined by 4sU labeling and fractionation

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A high resolution sequencing and modeling identifies distinct dynamic RNA regulatory strategies

Project description:To monitor the relative regulatory contributions of RNA production, processing and degradation, we sampled RNA from mouse DCs every 15 minutes, for the first 3 hours of their response to LPS (13 samples in total), following a short (10 minute) metabolic labeling pulse with 4sU preceding the sampled time point. We isolated RNA from each of these samples in two ways. First, to comprehensively measure total RNA regardless of its transcription time, we isolated RNA depleted of rRNA. Second, to measure only newly made RNA, we isolated 4sU-labeled RNA, which could only have been transcribed during the 10 minute labeling pulse and is thus enriched for short-lived transcripts, including mRNA precursors and processing intermediates. We deeply sequenced each isolated RNA population to provide the necessary depth to study exons, introns, and splicing junctions across the transcriptome. We showed our approach general utility by also applying it to lincRNAs and early zebrafish development data Time-course mRNA profiles of mouse DCs that were stimulated with LPS, and labeled with a short (10 minute) 4sU pulse (0-3 hours, 15 min. intervals, 13 time points).

2014-12-16 | E-GEOD-56977 | biostudies-arrayexpress

A high resolution sequencing and modeling identifies distinct dynamic RNA regulatory strategies

2014-12-16 | GSE56977 | GEO

Effect of 4sU labeling concentrations on quantification bias in nucleotide conversion RNA-seq data

Project description:To investigate the effect of increasing 4sU labeling concentrations on quantification bias in nucleotide conversion RNA-seq, we labeled 3 different cell lines with increasing concentrations of 4sU. We then measured 4sU dropout in 4sU samples compared to 4sU naive samples.

2024-02-06 | GSE229504 | GEO

Effect of 4sU labeling durations on quantification bias in nucleotide conversion RNA-seq data

Project description:To investigate the effect of labeling durations on quantification bias in nucleotide conversion RNA-seq, we labeled NIH cells with 4sU. We then measured 4sU dropout in 4sU samples compared to 4sU naive samples.

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Unstressed HeLa cells and ELAVL1/HuR knock down conditions: polyA RNA-Seq, small RNA-Seq, and PAR-CLIP

Project description:Post-transcriptional gene regulation relies on hundreds of RNA binding proteins (RBPs) but the function of most RBPs is unknown. The human RBP HuR/ELAVL1 is a conserved mRNA stability regulator. We used PAR-CLIP, a method based on RNA-protein crosslinking, to identify transcriptome wide ~26,000 HuR binding sites. These sites were on average highly conserved, enriched for HuR binding motifs and mainly located in 3' untranslated regions. Surprisingly, many sites were intronic, implicating HuR in splicing. Upon HuR knock down, mRNA levels and protein synthesis of thousands of target genes was down regulated, validating functionality. HuR and miRNA binding sites tended to reside nearby but generally did not overlap. Additionally, HuR knock down triggered strong and specific up regulation of miR-7. In summary, we identified thousands of direct and functional HuR targets, found a human miRNA controlled by HuR, and propose a role for HuR in splicing. PolyA mRNA was extracted from anti HuR siRNA treated and mock transfected HeLa cells to identify changes in mRNA expression and splicing. 2x100 paired end sequencing was performed according to the protocol on the Illumina HiSeq. PARCLIP was performed as in Hafner et. Al 2010 but with an antibody against endogenous HuR (3A2, Santa Cruz, sc-5261) in unstressed HeLa cells. We used, independently, 4-thiouridine (4SU) and 6-thioguanosine (6SG) to assess a possible nucleotide bias. As our proteomics measurements required labeling of cells in a special medium we also performed PAR-CLIP on cells grown in SILAC medium. Altogether we performed three PAR-CLIP experiments: 4SU labeling in standard DMEM medium, 4SU labeling in SILAC medium (as a replicate) and 6SG labeling in SILAC medium. Small RNA was extracted from anti HuR siRNA treated and mock transfected HeLa cells to identify changes in mRNA expression. Sequencing was performed on Illumina GAII using the standard sRNA 36cycle protocol.

2011-06-29 | E-GEOD-29943 | biostudies-arrayexpress

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