Comparative tRNA-seq for rapid profiling of tRNA modifications in a non-model organism
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ABSTRACT:
PROVIDER: PRJNA615798 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR11431928.fastq.gz | Fastqsanger.gz | |||
| SRR11431929.fastq.gz | Fastqsanger.gz | |||
| SRR11431930.fastq.gz | Fastqsanger.gz | |||
| SRR11431931.fastq.gz | Fastqsanger.gz | |||
| SRR11431932.fastq.gz | Fastqsanger.gz |
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Project description:tRNAs are heavily decorated with post-transcriptional modifications (tRNA modifications). Profile of tRNA modifications in non-model organisms are largely uncharacterized. Here using high-throughput sequencing, sites and frequency of tRNA modifications are predicted in Vibrio cholerae and Escherichia coli. During cDNA synthesis, some modifications cause misincorporation of a wrong base or termination of reverse transcription (RT). Using these RT-derived signatures we aim to explore organism-specific modifications and to track changes in modification frequency among different cellular conditions.
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Project description:A microarray tiling approach was utilized to assay tRNA modifications by comparing hybridization affinity of tRNA-modification mutants to wild-type across all annotated yeast tRNAs. Keywords: Assaying noncoding RNA modifications
2005-10-28 | GSE3503 | 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.
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Project description:Profiling tRNA modifications in Mycobacterium tuberculosis
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Project description:Abstract: tRNAs are highly modified in the elbow region and harbor 5-methyluridine at position 54 and pseudouridine at position 55 in the T arm, which are generated by the enzymes TrmA and TruB, respectively. Although all elongator tRNAs contain these modifications across all domains of life, the cellular relevance of these modifications and their corresponding modifying enzymes remains elusive. In addition to modifying every tRNA, Escherichia coli TrmA and TruB have both been shown to fold tRNA independently of its modification activity acting as tRNA chaperones, and strains lacking trmA or truB are outcompeted by wildtype. To identify how TrmA and TruB contribute to cellular fitness, we have systematically assessed the effects of deleting trmA and/or trmB in E. coli. Since tRNA folding is a pre-requisite for tRNA aminoacylation, we determined cellular aminoacylation levels revealing a global decrease in aminoacylation for all tRNAs in ΔtrmA and ΔtruB. Moreover, the absence of 5-methyluridine 54 or pseudouridine 55 alters tRNA modification at other positions: whereas acp3U47 is decreased, thiouridine levels are increased. Understanding the importance of TrmA and TruB for tRNA aminoacylation and modification, we then analyzed how these global tRNA changes in ΔtrmA and ΔtruB strains affect translation. Whereas global protein synthesis is not significantly changed in ΔtrmA and ΔtruB, the abundances of many specific proteins are altered, and transcriptomics experiments suggest that the dysregulation of many proteins is controlled at the translational level. In conclusion, we demonstrate that universally conserved modifications of the tRNA elbow are critical for global tRNA function by enhancing other tRNA modifications, tRNA folding, tRNA aminoacylation and translation of specific genes thereby contributing to cellular fitness.
2023-05-31 | GSE233667 | GEO
Project description:The tRNA pool determines the efficiency, throughput, and accuracy of translation. Previous studies have identified dynamic changes in the tRNA supply and mRNA demand during cancerous proliferation. Yet, dynamic changes may occur also during physiologically normal proliferation, and these are less characterized. We examined the tRNA and mRNA pools of T-cells during their vigorous proliferation and differentiation upon triggering of the T cell antigen receptor. We observe a global signature of switch in demand for codon at the early proliferation phase of the response, accompanied by corresponding changes in tRNA expression levels. In the later phase, upon differentiation of the T cells, the response of the tRNA pool is relaxed back to basal level, potentially restraining excessive proliferation. Sequencing of tRNAs allowed us to also evaluate their diverse base-modifications. We found that two types of tRNA modifications, Wybutosine and ms2t6A, are reduced dramatically during T-cell activation. These modifications occur in the anti-codon loops of two tRNAs that decode “slippery codons”, that are prone to ribosomal frameshifting. Attenuation of these frameshift-protective modifications is expected to increase proteome-wide frameshifting during T-cell proliferation. Indeed, human cell lines deleted of a Wybutosine writer showed increased ribosomal frameshifting, as detected with a reporter that consists of a critical frameshifting site taken from the HIV gag-pol slippery codon motif. These results may explain HIV’s specificity to proliferating T-Cells since it requires ribosomal frameshift exactly on this codon for infection. The changes in tRNA expression and modifications uncover a new layer of translation regulation during T-cell proliferation and exposes a potential trade-off between cellular growth and translation fidelity.
2021-10-13 | GSE165537 | GEO
Project description:All elongator tRNAs harbor 5-methyluridine at position 54 and pseudouridine at position 55 in the T arm, which are generated by the enzymes TrmA and TruB, respectively. Escherichia coli TrmA and TruB have both been shown to act as tRNA chaperones, and strains lacking trmA or truB are outcompeted by wildtype. Here, we investigate how TrmA and TruB contribute to cellular fitness. Deletion of trmA and truB in E. coli causes a global decrease in aminoacylation and alters other tRNA modification such as acp3U47 and 4-thiouridine. Whereas global protein synthesis is not significantly changed in ΔtrmA and ΔtruB, the expression of many specific proteins is altered at the translational level. In conclusion, we demonstrate that universal modifications of the tRNA T arm are critical for global tRNA function by enhancing other tRNA modifications, tRNA folding, tRNA aminoacylation, and translation of specific genes thereby improving cellular fitness and explaining their conservation.
2024-06-05 | GSE237609 | GEO
Project description:Covalent nucleotide modifications in noncoding RNAs such as tRNAs affect a plethora of biological processes, with new functions continuing to be discovered for even well-known tRNA modifications. To systematically compare the functions of a large set of ncRNA modifications in gene regulation, we carried out ribosome profiling and RNA-Seq in budding yeast for 57 nonessential genes involved in tRNA modification.
2017-06-29 | GSE100626 | GEO
Project description:tRNAs are known to be extensively decorated with various nucleotide modifications, which can modulate the biological properties of tRNAs, such as promoting translation efficiency or fidelity, influencing codon preference and stabilizing tertiary structure of tRNAs. Dynamic alteration of the tRNA modifications, particularly under stress, are conserved gene regulatory mechanisms employed by cells across the kingdoms of life. The malaria-causing Plasmodium falciparum, which has a highly AT-biased genome, is reliant on a non-redundant set of tRNA genes to decode distinct and codon-skewed transcriptomes across its developmental stages. This genomic feature highlights the regulatory potential of tRNA modifications and warrants studies on how the tRNA epitranscriptome can impact gene regulatory function in this parasite. However, systematic studies of tRNA modifications in P. falciparum remain sporadic. In this study, we present a sequencing-based analysis of tRNA modifications in P. falciparum through optimizing the TGIRT-tRNA sequencing approach. The optimized protocol significantly reduces reverse transcription (RT) stop-induced short reads and enhances full-length cDNA synthesis, enabling the identification and stoichiometric estimation of a select nucleotide modifications at single-base resolution. We were able to identify seven types of conserved tRNA modifications and discovered two non-canonical signatures at A59 and U73 positions on specific nuclear-encoded tRNAs, which provides new insights into previously unreported modification sites. Our findings further reveal the dynamic nature of tRNA modifications during stage progression and under stress conditions.
2025-05-18 | GSE297063 | GEO
Project description:Dynamic changes in tRNA modifications and abundance during T-cell activation [tRNA-seq]
| PRJNA694919 | ENA