LEAD-m6A-seq data of HeLa mRNA
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ABSTRACT:
PROVIDER: PRJNA665055 | ENA |
REPOSITORIES: ENA
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Project description:We applied LEAD-m6A-seq to 11 sites of HeLa mRNA to probe m6A modification at the target sites.
2020-11-01 | GSE158389 | GEO
Project description:We report the application of m6A-RIP-seq to indentify the m6A modifications variation in genes invovled in EMT. Briefly, total polyadenylated RNA was isolated from HeLa cells treated with or without 10 ng/ml TGF-β for 3 days by use of TRIZOL reagent followed the using of FastTrack MAGMaxi mRNA isolation kit (Invitrogen). RNA fragmentation, m6A-seq, and library preparation were performed according to instructions of manufacture and the previously published protocol (Wang et al., 2015). NEBNext Ultra Directional RNA Library Prep Kit (New England BioLabs, Ipswich, MA) was used for library preparation. Each experiment was conducted in two biological replicates. m6A-seq data were analyzed according to the protocols described before (Wang et al., 2015). Significant peaks with FDR < 0.05 were annotated to RefSeq database (hg19). Sequence motifs were identified by using Homer. Gene expression was calculated by Cufflinks using the sequencing reads from input samples. Cuffdiff was used to find the DE genes.
2019-03-25 | GSE112795 | GEO
Project description:m6A-RIP-seq of mRNA in HeLa cells undergoing EMT
| PRJNA448996 | ENA
Project description:RNA modifications are integral to regulation of RNA metabolism. One such abundant mRNA modification is m6A, which impacts various aspects of RNA metabolism including splicing, transport and degradation. Current knowledge about proteins recruited to m6A to carry out these molecular processes is still limited. Here we describe a comprehensive and systematic mass spectrometry-based screening of m6A interactors in various cell types and species. Amongst the main findings, we identified G3BP1 as a protein, which is repelled by m6A and which positively regulates mRNA stability in an m6A regulated manner. Furthermore, we identified FMR1 as a novel, RNA sequence context dependent m6A reader, thus revealing a connection between an mRNA modification and an autism spectrum disorder. Collectively, our data represents a rich resource for the community and sheds further light on the complex interplay between m6A, m6A interactors and mRNA homeostasis.
2017-08-22 | PXD006498 | Pride
Project description:N6-methyladenosine (m6A) is one of the most abundant modifications in eukaryotic RNA. Recent mapping of m6A methylomes in mammals, yeast, and plants as well as characterization of m6A methyltransferases, demethylases, and binding proteins have revealed regulatory functions of this dynamic RNA modification. In bacteria, although m6A is present in ribosomal RNA (rRNA), its occurrence in messenger RNA (mRNA) still remains elusive. Here, we used liquid chromatography-mass spectrometry (LC-MS) to calculate the m6A/A ratio in mRNA from a wide range of bacterial species, which demonstrates that m6A is an abundant mRNA modification in tested bacteria. Subsequent transcriptome-wide m6A profiling in Escherichia coli and Pseudomonas aeruginosa revealed a conserved distinct m6A pattern that is significantly different from that in eukaryotes. Most m6A peaks are located inside open reading frames (ORF), and carry a unique consensus motif (GCCAU). Functional enrichment analysis of bacterial m6A peaks indicates that the majority of m6A-modified transcripts are associated with respiration, amino acids metabolism, stress response, and small RNAs genes, suggesting potential regulatory roles of m6A in these pathways. m6A profiling in E.coli and P.aeruginosa mRNA
2015-07-08 | E-GEOD-65347 | biostudies-arrayexpress
Project description:N6-methyladenosine (m6A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m6A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila and human cells. In Drosophila, its depletion results in reduced m6A levels and altered m6A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerisation and interaction with other members of the m6A machinery, while its catalytic activity is dispensable. Finally, we demonstrate that the loss of Hakai destabilizes several subunits of the methyltransferase complex, resulting in impaired m6A deposition. Our work adds new functional and molecular insights into the mechanism of the m6A mRNA writer complex.
2021-04-29 | PXD022294 | Pride
Project description:Circadian transcriptional rhythms are necessary for lipid metabolic homeostasis. Disruptions can lead to metabolic diseases. Whether epigenetic N6-methyladenosine (m6A) mRNA methylation impacts circadian regulation of lipid metabolism is unclear. Here, we show m6A mRNA methylation oscillations in murine liver depend upon a functional circadian clock. Hepatic deletion of Bmal1 increased m6A mRNA methylation, particularly of PPaRα. Inhibition of m6A methylation via knockdown of m6A methyltransferase METTL3 decreased PPaRα m6A abundance and increased PPaRα mRNA lifetime and expression, reducing lipid accumulation in cells in vitro. Our data suggest YTH domain family 2 (YTHDF2, a m6A binding protein) binds to PPaRα, prolonging its lifetime and mRNA expression. Reactive oxygen species accumulation increased PPaRα transcript m6A levels, revealing a possible mechanism for circadian clock disruption on m6A mRNA methylation. These data suggest m6A RNA methylation is important for circadian clock regulation of downstream genes and lipid metabolism that impacts metabolic outcome.
2018-06-21 | GSE102620 | GEO
Project description:Circadian transcriptional rhythms are necessary for lipid metabolic homeostasis. Disruptions can lead to metabolic diseases. Whether epigenetic N6-methyladenosine (m6A) mRNA methylation impacts circadian regulation of lipid metabolism is unclear. Here, we show m6A mRNA methylation oscillations in murine liver depend upon a functional circadian clock. Hepatic deletion of Bmal1 increased m6A mRNA methylation, particularly of PPaRα. Inhibition of m6A methylation via knockdown of m6A methyltransferase METTL3 decreased PPaRα m6A abundance and increased PPaRα mRNA lifetime and expression, reducing lipid accumulation in cells in vitro. Our data suggest YTH domain family 2 (YTHDF2, a m6A binding protein) binds to PPaRα, prolonging its lifetime and mRNA expression. Reactive oxygen species accumulation increased PPaRα transcript m6A levels, revealing a possible mechanism for circadian clock disruption on m6A mRNA methylation. These data suggest m6A RNA methylation is important for circadian clock regulation of downstream genes and lipid metabolism that impacts metabolic outcome.
2018-10-25 | GSE102619 | GEO
Project description:HeLa cell chromatin associated nascent pre-mRNA (CA-RNA) contains many unspliced introns and m6A in exons, but very rarely in introns. m6A methylation of exons in CA-RNA frequently occur before splicing of exons and is essentially completed upon the release of mRNA into the nucleoplasm. m6A modifications are virtually the same in the newly synthesized CA-RNA, nucleoplasmic mRNA in transit and in the steady state cytoplasmic mRNA. This result suggests quantitatively little cytoplasmic methylation or demethylation. Only ~10% of m6As in CA-RNA are within 50 nucleotides of 5' or 3' splice sites arguing against a common splicing function of methylation. Both HeLa and mouse embryonic stem cell mRNAs showed a distinct increase of m6As, some in clusters, in short-lived mRNAs. Thus there is an unanticipated observation that m6A is added to exons presumably coincident with or soon after exon definition in nascent pre-mRNA, linking nuclear methylation to cytoplasmic mRNA stability.
2017-06-23 | GSE86336 | GEO
Project description:To investigate the effect of HSATIII lncRNA on m6A modification, we performed m6A-RIP(RNA immuno precipitation) RNA-seq from heat shock-exposed HeLa cells upon HSATIII knockdown.
2021-04-30 | GSE164200 | GEO