Differential redox sensitivity of tRNA dihydrouridylation
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PROVIDER: PRJEB73750 | ENA |
REPOSITORIES: ENA
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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:Oct4 redox sensitivity potentiates reprogramming and differentiation
| PRJNA1007695 | ENA
Project description:The transcription factor Oct4/Pou5f1 is a key component of the regulatory circuitry governing pluripotency. Oct4 is also widely used to generate induced pluripotent stem cells (iPSCs) from somatic cells. These observations provide compelling rationale to understand Oct4’s functions. Here we used domain swapping and mutagenesis to compare Oct4’s reprogramming activity with the paralog Oct1/Pou2f1, identifying a redox-sensitive DNA binding domain cysteine residue (Cys48) as a key determinant of both reprogramming and differentiation. In combination with the Oct4 N-terminus, mutating Oct1’s serine at this position to cysteine is sufficient to confer strong reprogramming activity. Conversely, Oct4C48S strongly reduces reprogramming potential. Oct4C48S renders the protein insensitive to oxidative inhibition of DNA binding. The same mutation prevents oxidation-mediated protein ubiquitylation. An engineered Pou5f1C48S point mutation has little effect on undifferentiated cells embryonic stem cells (ESCs), but upon retinoic acid (RA)-mediated differentiation causes retention of Oct4 expression, decreased proliferation, and increased apoptosis. Transcriptional profiling reveals that the retention of pluripotency also manifests at the RNA level. Pou5f1C48S ESCs also contribute poorly to adult somatic tissues and form less differentiated teratomas. Collectively, the data support a model in which Oct4 redox sensing is a positive reprogramming determinant during one or more reprogramming steps, and mediates Oct4 degradation during differentiation.
2024-04-09 | GSE241332 | GEO
Project description:Differential expression of human tRNA genes drives the abundance of tRNA-derived fragments
| PRJNA473011 | ENA
Project description:Transfer RNA (tRNA) molecules contain a variety of post-transcriptional modifications which are crucial for tRNA stability, translation efficiency, and fidelity. Besides their canonical roles in translation, tRNAs also originate tRNA-derived small RNAs (tsRNAs), a class of small non-coding RNAs with regulatory functions ranging from translation regulation, to gene expression control and cellular stress response. Recent evidence show that tsRNAs are also modified, however the impact of tRNA epitranscriptome deregulation on tsRNAs generation is only now beginning to be uncovered. The 5-methyluridine (m5U) modification at position 54 of cytosolic tRNAs is one of the most common and conserved tRNA modifications among species. This modification is catalyzed by the tRNA methyltransferase TRMT2A, but its biological role remains largely unexplored. Here, we show that TRMT2A knockdown in human cells induces m5U54 tRNA hypomodification, resulting in angiogenin (ANG) dependent tsRNA formation. More specifically, m5U54 hypomodification is followed by ANG overexpression and tRNA cleavage near the anticodon, resulting in accumulation of 5’tRNA-derived stress-induced RNAs (5’tiRNAs), in particular 5’tiRNA-GlyGCC and 5’tiRNA-GluCTC. Additionally, transcriptomic analysis confirms that down-regulation of TRMT2A and consequently m5U54 hypomodification impacts the cellular stress response and RNA stability, which is often correlated with tsRNA generation. Accordingly, exposure to oxidative stress conditions induces TRMT2A down-regulation and tsRNA formation in mammalian cells. These results establish a link between tRNA demethylation and tsRNAs formation and unravel m5U54 as a tRNA cleavage protective mark.
2021-04-20 | GSE164405 | GEO
Project description:We performed tRNAome sequencing to assess the tRNA changes of E.coli under oxidative stress. We found that the global translation inhibition is caused by global down-regulation of almost all tRNA species under oxidative stress. The translation elongation speed is resumed after the cells are fully adapted to the oxidative environment.
2015-05-28 | GSE62995 | GEO
Project description:Diverse chemical modifications fine-tune the function and metabolism of tRNA. Although tRNA modification is universal in all kingdoms of life, profiles of modifications, their functions, and physiological roles have not been elucidated in most organisms including the human pathogen, Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis. To identify physiologically important modifications, we surveyed the tRNA of Mtb, using tRNA sequencing (tRNA-seq). Reverse transcription-derived error signatures in tRNA-seq predicted the sites and presence of 9 modifications. Several chemical treatments prior to tRNA-seq expanded the number of predictable modifications. Deletion of Mtb genes encoding two modifying enzymes, TruB and MnmA, eliminated their respective tRNA modifications, validating the presence of modified sites in tRNA species.
2023-08-11 | GSE240200 | GEO
Project description:The goal of this study was to compare the endogenous tRNA expression levels from cells transduced with a lentiviral vector encopding Cas9 and an sgRNA from a U6 or a Glutamine tRNA promoter. Using high-throughput sequencing methods (Illumina Hi-Seq 2000) we show that endogenous tRNA expression levels are not perturbed by expression of an sgRNA from a human tRNA. tRNA expression profiles were generated from 2 different transduced polyconal cell lines.
2015-09-09 | E-GEOD-69720 | biostudies-arrayexpress
Project description:Checkpoint kinase 1 (CHK1) is critical for cell survival under replication stress (RS). CHK1 inhibitors (CHK1i’s) in combination with chemotherapy have shown promising results in preclinical studies but have displayed minimal efficacy with substantial toxicity in clinical trials. To explore combinatorial strategies that can overcome these limitations, we perform an unbiased high-throughput screen in a non-small cell lung cancer (NSCLC) cell line and identify thioredoxin1 (Trx1), a major component of the mammalian antioxidant-system, as a determinant of CHK1i sensitivity. We establish a role for redox recycling of RRM1, the larger subunit of ribonucleotide reductase (RNR), and a depletion of the deoxynucleotide pool in this Trx1-mediated CHK1i sensitivity. Further, the TrxR inhibitor auranofin, an approved anti-rheumatoid arthritis drug, shows a synergistic interaction with CHK1i via interruption of the deoxynucleotide pool. Together, we show a pharmacological combination to treat NSCLC that relies on a redox regulatory link between the Trx system and mammalian RNR activity.
2024-04-04 | GSE263176 | GEO
Project description:Transfer RNAs (tRNA) are quintessential in deciphering the genetic code; disseminating nucleic acid triplets into correct amino acid identity. While this decoding function is clear, an emerging theme is that tRNA abundance and functionality can powerfully impact protein production rate, folding, activity, and messenger RNA stability. Importantly, however, the expression pattern of tRNAs (in even simple systems) is obliquely known. Limited analysis suggests tRNA levels change during proliferation, differentiation, cancer, and neurodegeneration; possibly mediating changes in translation efficiency and mRNA stability. A major limitation for the field has been the ability to subject tRNA pools to high-throughput analysis as they are highly structured, modified, and of high sequence similarity. Here we present Quantitative Mature tRNA sequencing (QuantM-tRNA seq), an easily implemented high-throughput technique to monitor tRNA abundance and sequence variants (possibly due to RNA modifications). With QuantM-tRNA seq we provide a comprehensive analysis of the tRNA transcriptome from distinct mammalian tissues. We observe dramatic distinctions in isodecoder expression and likely RNA modifications between unique tissues with a particularly strong signature within the central nervous system. Remarkably, despite dramatic changes in tRNA isodecoder gene expression, the overall anticodon pool of each tRNA family is similar. These findings suggest that anticodon pools are buffered via an unknown mechanism to achieve uniform decoding throughout the body.
2020-08-04 | GSE141436 | GEO