The eukaryotic translation initiation factor eIF4E elevates steady-state m7G capping of coding and noncoding transcripts
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ABSTRACT:
PROVIDER: PRJNA666357 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR12739715.fastq.gz | Fastqsanger.gz | |||
| SRR12739716.fastq.gz | Fastqsanger.gz | |||
| SRR12739717.fastq.gz | Fastqsanger.gz | |||
| SRR12739718.fastq.gz | Fastqsanger.gz | |||
| SRR12739719.fastq.gz | Fastqsanger.gz |
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Project description:Methyl-7-guanosine (m7G) “capping” of coding and some noncoding RNAs is critical for their maturation and subsequent activity. Here, we discovered that eukaryotic translation initiation factor 4E(eIF4E), itself a cap-binding protein, drives the expression of the capping machinery and the increased capping efficiency of ∼100 coding and noncoding RNAs. This dataset collects transcriptomic data for quantitative cap immunoprecipitation (CapIP) assay in eIF4E-Flag or vector stable U2Os cells.
2020-09-30 | GSE158728 | GEO
Project description:The analysis of capped RNAs by massively parallel sequencing has identified a large number of previously unknown transcripts, some of which are small RNAs and others are 5M-bM-^@M-^Y truncated forms of RefSeq genes. The latter may be generated by endonuclease cleavage or by stalling of Xrn1 at defined sites. With the exception of promoter-proximal transcripts the caps on all of these are added post-transcriptionally by a cytoplasmic capping enzyme complex that includes capping enzyme and a kinase that converts 5M-bM-^@M-^Y-monophosphate ends to a diphosphate capping substrate. We previously described a modified form of capping enzyme with dominant negative activity against cytoplasmic capping (DN-cCE). A tet-inducible form of this was used to identify substrates for cytoplasmic capping by treating cytoplasmic RNA from control and induced cells with and without Xrn1. Surviving RNA was analyzed on Affymetrix Human Exon 1.0 arrays and scored for changes in probe intensity as a function of its position on each RefSeq gene to derive a factor (alpha) that could be compared between sets. Notably, transcriptome-wide changes were not evident unless RNA was treated with Xrn1. This analysis identified 2,666 uncapped mRNAs in uninduced cells, 672 mRNAs that appeared in the uncapped pool in cells expressing DN-cCE, and 835 mRNAs that were in both populations. Changes in cap status of 10 re-capping targets and 5 controls were assessed by 3 independent measures; susceptibility to Xrn1, recovery with a biotin-tagged DNA primer after ligating a complementary RNA oligonucleotide to uncapped 5M-bM-^@M-^Y ends, and binding or exclusion from a high affinity cap-binding matrix comprised of immobilized eIF4E and the corresponding binding domain of eIF4G. 3 biological replicates of 4 different samples comparing XRN1 treatment/non-treatment and Dox induction/non-induction of K294A
2012-12-20 | E-GEOD-36729 | biostudies-arrayexpress
Project description:The analysis of capped RNAs by massively parallel sequencing has identified a large number of previously unknown transcripts, some of which are small RNAs and others are 5’ truncated forms of RefSeq genes. The latter may be generated by endonuclease cleavage or by stalling of Xrn1 at defined sites. With the exception of promoter-proximal transcripts the caps on all of these are added post-transcriptionally by a cytoplasmic capping enzyme complex that includes capping enzyme and a kinase that converts 5’-monophosphate ends to a diphosphate capping substrate. We previously described a modified form of capping enzyme with dominant negative activity against cytoplasmic capping (DN-cCE). A tet-inducible form of this was used to identify substrates for cytoplasmic capping by treating cytoplasmic RNA from control and induced cells with and without Xrn1. Surviving RNA was analyzed on Affymetrix Human Exon 1.0 arrays and scored for changes in probe intensity as a function of its position on each RefSeq gene to derive a factor (alpha) that could be compared between sets. Notably, transcriptome-wide changes were not evident unless RNA was treated with Xrn1. This analysis identified 2,666 uncapped mRNAs in uninduced cells, 672 mRNAs that appeared in the uncapped pool in cells expressing DN-cCE, and 835 mRNAs that were in both populations. Changes in cap status of 10 re-capping targets and 5 controls were assessed by 3 independent measures; susceptibility to Xrn1, recovery with a biotin-tagged DNA primer after ligating a complementary RNA oligonucleotide to uncapped 5’ ends, and binding or exclusion from a high affinity cap-binding matrix comprised of immobilized eIF4E and the corresponding binding domain of eIF4G.
2012-12-20 | GSE36729 | GEO
Project description:Methyl-7-guanosine (m7G) "capping" of coding and some noncoding RNAs is critical for their maturation and subsequent activity. Here, we discovered that eukaryotic translation initiation factor 4E (eIF4E), itself a cap-binding protein, drives the expression of the capping machinery and increased capping efficiency of ∼100 coding and noncoding RNAs. To quantify this, we developed enzymatic (cap quantification; CapQ) and quantitative cap immunoprecipitation (CapIP) methods. The CapQ method has the further advantage that it captures information about capping status independent of the type of 5' cap, i.e., it is not restricted to informing on m7G caps. These methodological advances led to unanticipated revelations: 1) Many RNA populations are inefficiently capped at steady state (∼30 to 50%), and eIF4E overexpression increased this to ∼60 to 100%, depending on the RNA; 2) eIF4E physically associates with noncoding RNAs in the nucleus; and 3) approximately half of eIF4E-capping targets identified are noncoding RNAs. eIF4E's association with noncoding RNAs strongly positions it to act beyond translation. Coding and noncoding capping targets have activities that influence survival, cell morphology, and cell-to-cell interaction. Given that RNA export and translation machineries typically utilize capped RNA substrates, capping regulation provides means to titrate the protein-coding capacity of the transcriptome and, for noncoding RNAs, to regulate their activities. We also discovered a cap sensitivity element (CapSE) which conferred eIF4E-dependent capping sensitivity. Finally, we observed elevated capping for specific RNAs in high-eIF4E leukemia specimens, supporting a role for cap dysregulation in malignancy. In all, levels of capping RNAs can be regulated by eIF4E.
| S-EPMC7604501 | biostudies-literature
Project description:Eukaryotic messenger RNAs (mRNAs) possess a 5’-end N7-methyl guanosine (m7G) cap that promotes their translation and stability. However, it was recently demonstrated that eukaryotic mRNAs can also carry a 5' end nicotinamide adenine dinucleotide (NAD+) cap that promotes mRNA decay mediated by the NAD+ decapping enzyme DXO1. However, the dynamic regulation of NAD+ capping in plant remains unknown. Here, we describe the global landscape of NAD+-capped RNAs in Arabidopsis thaliana, and demonstrate that DXO1 is responsible for removal of these 5’-end modifications and facilitates mRNA degradation in plant transcriptomes. We also reveal that in the absence of DXO1 NAD+-capped mRNAs are unstable and processed into smRNAs. Furthermore, we find that Abscisic Acid (ABA) remodel the landscape of RNA cap epitransciptome, and the mRNA lost their NAD+ cap contribute to their stability under ABA. Overall, our results support a link between ABA response and RNA NAD+ capping.
2020-11-23 | GSE142389 | GEO
Project description:Eukaryotic messenger RNAs (mRNAs) possess a 5’-end N7-methyl guanosine (m7G) cap that promotes their translation and stability. However, it was recently demonstrated that eukaryotic mRNAs can also carry a 5' end nicotinamide adenine dinucleotide (NAD+) cap that promotes mRNA decay mediated by the NAD+ decapping enzyme DXO1. However, the dynamic regulation of NAD+ capping in plant remains unknown. Here, we describe the global landscape of NAD+-capped RNAs in Arabidopsis thaliana, and demonstrate that DXO1 is responsible for removal of these 5’-end modifications and facilitates mRNA degradation in plant transcriptomes. We also reveal that in the absence of DXO1 NAD+-capped mRNAs are unstable and processed into smRNAs. Furthermore, we find that Abscisic Acid (ABA) remodel the landscape of RNA cap epitransciptome, and the mRNA lost their NAD+ cap contribute to their stability under ABA. Overall, our results support a link between ABA response and RNA NAD+ capping.
2020-11-23 | GSE142388 | GEO
Project description:N7-methylguanosine (m7G) in variable loop region of tRNA stabilizes target tRNA expression, which is catalyzed by METTL1/WDR4 heterodimer. Here, we unveil essential functions of Mettl1 in Drosophila fertility. Mettl1-knockout (Mettl1-KO) decreases elongated spermatid and mature sperm, which is fully rescued by Mettl1-transgene expression, but not catalytic dead Mettl1-transgene, demonstrating that Mettl1-dependent m7G is required for spermatogenesis. Mettl1-KO shows loss of m7G modification on subset of tRNAs and decreased level of tRNA expression. Strikingly, overexpression of translational elongation factor EF1α that can compete with rapid tRNA decay (RTD) pathway in S. cerevisiae, significantly counteract the sterility in Mettl1-KO male, supporting a critical role of m7G tRNA modification in spermatogenesis. Ribo-seq analysis shows that Mettl1-KO elevates ribosome collisions at codons decoded by reduced tRNAs and significantly reduces translation of genes involved in elongated spermatid formation and sperm stability. These findings reveal a developmental role for m7G tRNA modifications and suggest that m7G modification-dependent tRNA stability differs among tissues.
2024-08-16 | GSE241519 | GEO
Project description:Previous study found the presence of internal N7-methylguanosine (m7G) within mammalian mRNA. We performed antibody-based m7G MeRIP-seq for transcriptome-wide mapping the internal m7G sites in U2OS cells.
2023-05-17 | GSE192845 | GEO
Project description:Previous study found the presence of internal N7-methylguanosine (m7G) within mammalian mRNA. We performed antibody-based m7G MeRIP-seq for transcriptome-wide mapping the internal m7G sites in HepG2 cells.
2023-05-17 | GSE192844 | GEO
Project description:The 5’ cap, catalyzed by capping enzyme, RNGTT, is a vital mRNA modification for the functionality of mRNAs and dysregulated in cancer cells. The m6A writer METTL3 is a regulator of mRNA stability and protein translation, and its expression is altered in various cancers, including lung cancer cells. We identified a METTL3 motif in the mRNA of RNGTT and investigated whether METTL3 could regulate the expression of RNGTT. Knock down of METTL3 resulted in destabilizing RNGTT mRNA, reduced protein expression and activity. Sequencing of capped mRNAs and m7G immunoprecipitation identified an underrepresentation of ribosomal protein mRNA.
2024-10-21 | GSE273305 | GEO