High-resolution quantitative profiling of tRNA abundance and modification status in eukaryotes by mim-tRNAseq
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ABSTRACT:
PROVIDER: PRJNA639839 | ENA |
REPOSITORIES: ENA
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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:High-resolution quantitative profiling of tRNA abundance and modification status in hematopoietic cells upon Trmt6 deletion by mim-tRNAseq
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Project description:To investigate differential mitochondrial tRNA gene expression in wild type (WT) and mutant (E423P) mitochondrial RNA polymerase (POLRMT) overexpression flies, we performed Drsophila mitochondrial tRNA-seq according to the Hydro-tRNAseq method. tRNA-seq reads were subjected to 3' - adapter filtering , 3' - adapter trimming and quality control. The tRNAs expression levels were measured by tag count. For each tRNA sequence-based profile, the mapped reads number used to estimate the expression level of each tRNA. The tRNAs expression profiling was calculated based on uniquely mapped reads and including mapped reads, respectively. We found no difference in the relative abundance of mitochondrial individual tRNAs between WT and E423P overexpression flies.
2021-01-06 | GSE164324 | GEO
Project description:Respiratory viruses such as SARS-CoV-2 are pathogens that can reach pandemic proportions. The early human response to respiratory viruses are in the nasal cavity and nasopharyngeal regions. Defining biomarkers of disease trajectory at the time of a positive test is important for opting for treatment and monitoring decisions. We hypothesize that the nasopharyngeal tRNA profiles can be used to predict symptom severity resulting from SARS-CoV-2 infection. We carried out multiplex small RNA sequencing (MSR-seq) on nasopharyngeal swaps to measure the human tRNA response to SARS-CoV-2 infection. Our result simultaneously measured full-length tRNA abundance, tRNA modifications, and tRNA fragmentation among individuals that went on to develop no/mild or severe symptoms. We identified distinct tRNA signatures that can predict the clinical outcome of SARS-CoV-2 infected individual at the time of a positive test. These results highlight the utility of using host tRNA properties as biomarkers for clinical applications.
2022-10-01 | GSE208612 | 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 endogenously modified tRNA with a comprehensive and user-friendly computational analysis toolkit. Our method accurately captures tRNA abundance and modification status in yeast, fly, and human cells and is applicable to any organism with a known genome. We applied mim-tRNAseq to discover a dramatic heterogeneity of tRNA isodecoder pools among diverse human cell lines and a surprising interdependence of modifications at distinct sites within the same tRNA transcript.
| S-EPMC8062790 | biostudies-literature
Project description:The human genome encodes hundreds of tRNA genes but their individual contribution to the tRNA pool is not fully understood. Deep sequencing of tRNA transcripts (tRNA-Seq) can estimate tRNA abundance at single gene resolution, but tRNA structures and post-transcriptional modifications impair these analyses. Here we present a bioinformatics strategy to investigate differential tRNA gene expression and use it to compare tRNA-Seq datasets from cultured human cells and human brain. We find that sequencing caveats affect quantitation of only a subset of human tRNA genes. Unexpectedly, we detect several cases where the differences in tRNA expression among samples do not involve variations at the level of isoacceptor tRNA sets (tRNAs charged with the same amino acid but using different anticodons); but rather among tRNA genes within the same isodecoder set (tRNAs having the same anticodon sequence). Because isodecoder tRNAs are functionally equal in terms of genetic translation, their differential expression may be related to non-canonical tRNA functions. We show that several instances of differential tRNA gene expression result in changes in the abundance of tRNA-derived fragments (tRFs) but not of mature tRNAs. Examples of differentially expressed tRFs include: PIWI-associated RNAs, tRFs present in tissue samples but not in cells cultured in vitro, and somatic tissue-specific tRFs. Our data support that differential expression of tRNA genes regulate non-canonical tRNA functions performed by tRFs.
2019-04-17 | GSE114904 | GEO
Project description:Dynamic changes in tRNA modifications and abundance during T-cell activation [tRNA-seq]
| PRJNA694919 | ENA
Project description:Differential expression of human tRNA genes drives the abundance of tRNA-derived fragments
| PRJNA473011 | ENA
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:RNase P is essential to perform the 5’ maturation of tRNA precursors. Beyond the ancestral form of RNase P containing a ribozyme, protein-only RNase P enzymes termed PRORP were identified in eukaryotes. In human mitochondria, PRORP forms a complex with two protein partners to become functional. In plants, although PRORP enzymes are active alone, we investigate their interaction network to understand their integration with gene expression pathways. Here we investigate functional interactions involving the Arabidopsis nuclear RNase P PRORP2. We show, using an immuno-affinity strategy, that PRORP2 makes a complex with the tRNA methyl transferases TRM1A and B in vivo. Beyond RNase P, these enzymes can also interact with RNase Z. We show that TRM1A/B localize in the nucleus and find that their double knock out mutation results in a severe macroscopic phenotype. Using a combination of immuno-detections, mass spectrometry and a transcriptome wide tRNAseq approach, we observe that TRM1A/B are responsible for the m2,2G26 modification of 70% of cytosolic tRNAs in vivo. We use the transcriptome wide tRNAseq approach as well as RNA blot hybridizations to show that RNase P activity is impaired in TRM1A/B mutants for specific tRNAs, in particular, tRNAs containing a m2,2G modification at position 26 that are strongly down-regulated in TRM1A/B mutants. Altogether, results indicate that the m2,2G adding enzymes TRM1A/B functionally cooperate with nuclear RNase P in vivo for the early steps of cytosolic tRNAs biogenesis.
2023-10-20 | PXD040022 | Pride