Halfpipe: a tool for analyzing metabolic labeling RNA-seq data to quantify RNA half-lives
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PROVIDER: PRJNA1166201 | ENA |
REPOSITORIES: ENA
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Project description:We introduce Halfpipe, a tool for analyzing RNA-seq data from metabolic RNA labeling experiments. Its main features are the absolute quantification of 4sU-labeling-induced T>C conversions in the data, calculating the proportion of newly synthesized transcripts, and estimating (compartment-specific) RNA half-lives. Halfpipe excels at correcting critical biases caused by typically low labeling efficiency. We measure and compare the RNA metabolism in the G1 phase and during the mitosis of synchronized human cells. We find that RNA half-lives of constantly expressed RNAs are similar in mitosis and G1 phase, suggesting that RNA stability of those genes is constant throughout the cell cycle. Our estimates correlate well with literature values and with known RNA sequence features.
2025-02-26 | GSE278274 | GEO
Project description:Global RNA half lives in cyanobacteria
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Project description:Dinoflagellates possess many physiological processes that appear to be under post-transcriptional control. However, the extent to which their genes are regulated post-transcriptionally remains unresolved. To gain insight into the role of differential mRNA stability in dinoflagellates, we biosynthetically labeled RNA with 4-thiouracil to isolate newly transcribed and pre-existing RNA pools in Karenia brevis. These isolated fractions were then used for analysis of global mRNA stability by hybridization to a K. brevis microarray. Global K. brevis mRNA half-lives were calculated from the ratio of newly transcribed/pre-existing RNA for 7086 array features using the online software HALO (Half-life Organizer). Overall, mRNA half-lives were substantially longer than reported in other organisms studied at the global level, ranging from 42 minutes to greater than 3 days, with a median of 33 hours. Thirteen percent of messages showed a half-life of 3 days, demonstrating their stability throughtout the course of the cell cycle and divison. Consistent with well-documented trends observed in other organisms, housekeeping processes, including energy metabolism and transport, were significantly enriched in the most highly stable messages. Shorter-lived transcripts included a higher proportion of transcriptional regulation, stress response, and other response/regulatory processes. Log phase cultures (n=6) were exposed to 0.2 mM 4-thiouracil for 2h to biosynthetically label newly transcribed RNA and total RNA was extracted. Following extraction, RNA was biotinylated to allow for purification of the thiolated newly synthesized RNA from the total RNA pool using streptavidin coated magnetic beads. From each replicate 3 pools of RNA, total RNA, pre-exisiting RNA, and newly synthesized RNA, were Cy3 labeled and hybridized to microarrys in a one color format. Based on the appearance of bioanlayzer profiles, total and pre-existing RNA were treated as total RNA, while newly synthesized RNA was treated as mRNA in the labeling protocol.
2013-08-01 | E-GEOD-46174 | biostudies-arrayexpress
Project description:We calculated global RNA half lives for all genes in the cyanobacteria Synechococcus sp. strain PCC 7002. Samples from three replicates of the WT strain were taken before and following the addition of the antibiotic rifampicin to stop nascent transcription. RNA spike-ins were added for normalization and using the number of RNA-sequencing reads we were able to calculate half lives for genes, transcripts, and for each position on the genome.
2019-01-01 | GSE109174 | GEO
Project description:Dinoflagellates possess many physiological processes that appear to be under post-transcriptional control. However, the extent to which their genes are regulated post-transcriptionally remains unresolved. To gain insight into the role of differential mRNA stability in dinoflagellates, we biosynthetically labeled RNA with 4-thiouracil to isolate newly transcribed and pre-existing RNA pools in Karenia brevis. These isolated fractions were then used for analysis of global mRNA stability by hybridization to a K. brevis microarray. Global K. brevis mRNA half-lives were calculated from the ratio of newly transcribed/pre-existing RNA for 7086 array features using the online software HALO (Half-life Organizer). Overall, mRNA half-lives were substantially longer than reported in other organisms studied at the global level, ranging from 42 minutes to greater than 3 days, with a median of 33 hours. Thirteen percent of messages showed a half-life of 3 days, demonstrating their stability throughtout the course of the cell cycle and divison. Consistent with well-documented trends observed in other organisms, housekeeping processes, including energy metabolism and transport, were significantly enriched in the most highly stable messages. Shorter-lived transcripts included a higher proportion of transcriptional regulation, stress response, and other response/regulatory processes.
2013-08-01 | GSE46174 | GEO
Project description:We obtained proteome wide measurements of protein half-lives in M. pneumoniae by performing pulse-chase experiments following SILAC methods. We estimated degradation rates of individual proteins by measuring the increase in light protein compared to heavy protein labeling along the growth curve.
2020-11-24 | PXD016386 | Pride
Project description:This work aimed at analysing mRNA half lives in drug resistant and in drug sensitive strains of C. albicans (Gu4 and Gu5 azole susceptible and resistant clinical isolates (franz et al., 1999)), using the transcriptional inhibitor thiolutine.
2011-10-01 | E-TABM-1179 | biostudies-arrayexpress
Project description:We calculated half-life values of mRNAs quantified by RNA-Seq by a suitable method of normalization. We determined the half-lives of more than 2200 mRNAs in the Stenotrophomonas maltophilia D457 wild-type strain and in an isogenic RNase G deficient mutant. Median half-lives were 2,74 and 3 min in the wild-type and the rng-deficient mutant respectively. We found an overall enhancement of half-life times of mRNAs when the gene encoding RNase G is lacking, showing that many RNAs are targets of RNase G in S. maltophilia. For achieving such goal, we propose a method for the normalization of RNA-Seq based studies on global bacterial mRNA decay.
2017-09-06 | GSE103467 | GEO
Project description:Determining mRNA half-lives on a transcriptome-wide scale
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Project description:Determination of mRNA half-lives in embryonic mouse skin
| PRJEB21639 | ENA