ALL-tRNAseq enables robust tRNA profiling in tissue samples
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ABSTRACT:
PROVIDER: PRJNA775872 | ENA |
REPOSITORIES: ENA
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Project description:Transfer RNAs (tRNAs) are small adaptor RNAs essential for mRNA translation. Alterations in the cellular tRNA population can directly affect mRNA decoding rates and translational efficiency during cancer development and progression. To evaluate changes in the composition of the tRNA pool, multiple sequencing approaches have been developed to overcome reverse transcription blocks caused by the stable structures of these molecules and their numerous base modifications. It remains however unclear whether current sequencing protocols faithfully capture tRNAs existing in cells or tissues. This is specifically challenging for clinical tissue samples that often present variable RNA qualities. For this reason, we developed ALL-tRNAseq, which combines the highly processive MarathonRT and RNA demethylation, for the robust assessment of tRNA expression, together with a randomized adapter ligation strategy prior to reverse transcription, to assess tRNA fragmentation levels in both cell lines and tissues. Incorporation of tRNA fragments not only informed on sample integrity, but also significantly improved tRNA profiling of tissue samples. Our data showed that our profiling strategy effectively improves classification of oncogenic signatures in glioblastoma and diffuse large B-cell lymphoma tissues, particularly for samples presenting higher levels of RNA fragmentation, further highlighting the utility of ALL-tRNAseq for translational research.
2023-02-24 | GSE186736 | GEO
Project description:Transfer RNAs (tRNAs) are small adaptor RNAs essential for mRNA translation. Alterations in the cellular tRNA population can directly affect mRNA decoding rates and translational efficiency during cancer development and progression. To evaluate changes in the composition of the tRNA pool, multiple sequencing approaches have been developed to overcome reverse transcription blocks caused by the stable structures of these molecules and their numerous base modifications. However, it remains unclear whether current sequencing protocols faithfully capture tRNAs existing in cells or tissues. This is specifically challenging for clinical tissue samples that often present variable RNA qualities. For this reason, we developed ALL-tRNAseq, which combines the highly processive MarathonRT and RNA demethylation for the robust assessment of tRNA expression, together with a randomized adapter ligation strategy prior to reverse transcription to assess tRNA fragmentation levels in both cell lines and tissues. Incorporation of tRNA fragments not only informed on sample integrity but also significantly improved tRNA profiling of tissue samples. Our data showed that our profiling strategy effectively improves classification of oncogenic signatures in glioblastoma and diffuse large B-cell lymphoma tissues, particularly for samples presenting higher levels of RNA fragmentation, further highlighting the utility of ALL-tRNAseq for translational research.
| S-EPMC10111867 | biostudies-literature
Project description:High-resolution quantitative profiling of tRNA abundance and modification status in hematopoietic cells upon Trmt6 deletion by mim-tRNAseq
2024-06-03 | GSE252912 | GEO
Project description:Measurements of cellular tRNA abundance are hampered by pervasive blocks to cDNA synthesis at modified nucleosides and the extensive similarity among tRNA genes. We overcome these limitations with modification-induced misincorporation tRNA sequencing (mim-tRNAseq), which combines a workflow for full-length cDNA library construction from mature tRNA with a simple-to-use computational analysis toolkit. Our method accurately captures tRNA abundance and modification status in multiple eukaryotes and is applicable to any organism with a known genome.
2021-02-12 | GSE152621 | GEO
Project description:We analyze whether tRNA quantifications based on small RNA-seq data are informative enough to distinguish between different human cell lines covering multiple tissue types. We therefore apply both small RNA-seq and Hydro-tRNAseq to HEK293 (kidney), HCT116 (colon), HeLa (cervix), MDA-MB-231 (breast), and BJ fibroblasts. First, the correlations between the two methods of identical samples and computational mapping pipeline range between 0.93 and 0.96 for all cell lines. tRNA quantifications from both protocols are compared and significantly higher Spearman correlations are obtained within matching samples versus mismatching cell lines. In consequence, we demonstrate that small RNA-seq quantifications of sample-specific tRNA profiles show a good agreement with conventional tRNA-seq.
2020-02-28 | GSE137834 | GEO
Project description:Mapping of the epitranscriptome has revealed the chemical diversity of RNA modifications and their functional importance in regulating gene expression. Transfer RNAs (tRNAs) are one of the most modified cellular RNAs, containing on average 10-13 modifications per molecule. These modifications have been shown to be critical for several aspects of tRNA functions, such as decoding, folding, and stability. Here we report that the human RNA methyltransferase TRMT1L associates with components of the Rix1 ribosome biogenesis complex and co-sediments with pre-60S ribosomes. Using eCLIP-Seq, we show that TRMT1L binds to a subset of tRNAs and to the 28S rRNA. Additionally, we demonstrate that TRMT1L is responsible for catalyzing N2, N2-dimethylguanosine (m22G) solely at position 27 of tRNA-Tyr-GUA by Nano-tRNAseq and RNA LC-MS. Surprisingly, TRMT1L depletion also impaired the deposition of acp3U and dihydrouridine on tRNA-Tyr-GUA, Cys-GCA, and Ala-CGC. TRMT1L knockout cells have a marked decrease in tRNA-Tyr-GUA levels, coinciding with a reduction in global translation rates and hypersensitivity of oxidative stress. Our results establish TRMT1L as the elusive methyltransferase catalyzing the m22G27 modification on tRNA Tyr, resolving a long-standing gap of knowledge and highlighting its potential role in a tRNA modification circuit crucial for translation regulation and stress response.
2024-05-08 | GSE266588 | GEO
Project description:tRNA m1A modification regulate hematopoietic stem cell maintenance via mTORC1 signaling [mim-tRNAseq]
| PRJNA1063221 | ENA
Project description:The participation of transfer RNAs (tRNAs) in fundamental aspects of biology and disease necessitates an accurate, experimentally confirmed annotation of tRNA genes, and curation of precursor and mature tRNA sequences. This has been challenging, mainly because RNA secondary structure and nucleotide modifications, together with tRNA gene multiplicity, complicate sequencing and sequencing read mapping efforts. To address these issues, we developed hydro-tRNAseq, a method based on partial alkaline RNA hydrolysis that generates fragments amenable for sequencing. To identify transcribed tRNA genes, we further complemented this approach with Photoactivatable Crosslinking and Immunoprecipitation (PAR-CLIP) of SSB/La, a conserved protein involved in pre-tRNA processing. Our results show that approximately half of all predicted tRNA genes are transcribed in human cells. We also report predominant nucleotide modification sites, their order of introduction, and identify tRNA leader, trailer and intron sequences. By using complementary sequencing-based methodologies, we present a human tRNA atlas, and determine expression levels of mature and processing intermediates of tRNAs in human cells.
2017-07-17 | GSE95683 | GEO
Project description:Valine tRNA biogenesis and bioavailability regulates mitochondrial complex I levels in acute leukemia [tRNAseq]
| PRJNA695557 | ENA
Project description:tRNA quantification of five human cell lines by hydro-tRNAseq and small RNA-seq
| PRJNA573631 | ENA