The function of intragenic tRNA R-loop
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ABSTRACT:
PROVIDER: PRJNA640382 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Intragenic tRNAs-promoted R-loops formation orchestrate Arabidopsis oxidative stress survival
2021-09-01 | GSE152776 | GEO
Project description:Expression of selenoproteins requires the co-translational incorporation of selenocysteine (Sec) in response to an in-frame UGA codon. The machinery of UGA/Sec re-coding is complex and many factors affect the hierarchy of expression among selenoproteins, including modification of tRNA[Ser]Sec. Its hyper-modification in the anticodon stem loop is influenced by selenium bioavailability, and a mutation in adenosine 37 (A37) that abrogates isopentenylation, has a profound effect on selenoprotein expression in mice. Patients with mutations in tRNA-isopentenyl-transferase (TRIT1) show a severe neurological disorder and hence we wondered whether mutations in TRIT1 negatively affected the expression of selenoproteins. Fibroblasts from a patient carrying a pathogenic R323Q mutation in TRIT1 in homozygosity did not show decreased selenoprotein expression, although recombinant TRIT1R323Q had significantly reduced activity in vitro towards anticodon stem-loop substrates. We thus engineered mice conditionally deficient in Trit1 in hepatocytes and neurons. Selenoprotein expression as assessed by western blotting, 75Se metabolic labeling, and ribosomal profiling was not decreased despite the general reduction of N6-isopentenyl-adenosine in tRNAs. We show that 5-methylcarboxymethylation and 2’O-methylation of U34 occur independently of isopentenylation of A37 in tRNA[Ser]Sec. Reanalyzing previously published ribosomal profiling datasets, we demonstrate that (i) failure of 5-carboxymethylation at U34 is associated with reduced expression of GPX1, but not GPX4, and that (ii) FTSJ1 is not the elusive U34-2’O-methyltransferase involved in the methylation of tRNA[Ser]Sec.
2021-11-17 | GSE183923 | GEO
Project description:Genomic rearrangements may cause both Mendelian and complex disorders. Currently, several major mechanisms causing genomic rearrangements have been proposed such as non-allelic homologous recombination (NAHR), non-homologous end joining (NHEJ), fork stalling and template switching (FoSTeS) and microhomology-mediated break-induced replication (MMBIR). However, to what extent these mechanisms contribute to gene-specific pathogenic copy-number changes (CNCs) remains understudied. Furthermore, only few studies resolved these pathogenic alterations at nucleotide-level resolution. Accordingly, our aim is to explore which mechanisms contribute to a large, unique set of locus-specific non-recurrent genomic rearrangements causing the genetic neurocutaneous disorder neurofibromatosis type 1 (NF1). Through breakpoint-spanning PCR as well as array Comparative Genomic Hybridization (aCGH), we have identified the breakpoints and characterized the likely rearrangement mechanism of the NF1 intragenic CNCs in 78 unrelated patients. Unlike the most typical recurrent rearrangements mediated by flanking low copy repeats (LCRs), NF1 intragenic CNCs have diverse breakpoint locations, and are characterized by different rearrangement mechanisms. We propose the DNA replication-based mechanisms comprising FoSTeS/MMBIR and serial replication stalling to be the predominant mechanism leading to NF1 intragenic CNCs. In addition to the loop of a 197-bp palindrome located in intron 40, four Alu elements located in intron 1, 2, 3 and 50 were also identified as significant intragenic rearrangement hotspots within the NF1 gene. However, no clear genotype-phenotype correlations could be identified among the NF1 patients carrying NF1 intragenic CNCs.
2015-07-30 | GSE64869 | GEO
Project description:Genomic rearrangements may cause both Mendelian and complex disorders. Currently, several major mechanisms causing genomic rearrangements have been proposed such as non-allelic homologous recombination (NAHR), non-homologous end joining (NHEJ), fork stalling and template switching (FoSTeS) and microhomology-mediated break-induced replication (MMBIR). However, to what extent these mechanisms contribute to gene-specific pathogenic copy-number changes (CNCs) remains understudied. Furthermore, only few studies resolved these pathogenic alterations at nucleotide-level resolution. Accordingly, our aim is to explore which mechanisms contribute to a large, unique set of locus-specific non-recurrent genomic rearrangements causing the genetic neurocutaneous disorder neurofibromatosis type 1 (NF1). Through breakpoint-spanning PCR as well as array Comparative Genomic Hybridization (aCGH), we have identified the breakpoints and characterized the likely rearrangement mechanism of the NF1 intragenic CNCs in 78 unrelated patients. Unlike the most typical recurrent rearrangements mediated by flanking low copy repeats (LCRs), NF1 intragenic CNCs have diverse breakpoint locations, and are characterized by different rearrangement mechanisms. We propose the DNA replication-based mechanisms comprising FoSTeS/MMBIR and serial replication stalling to be the predominant mechanism leading to NF1 intragenic CNCs. In addition to the loop of a 197-bp palindrome located in intron 40, four Alu elements located in intron 1, 2, 3 and 50 were also identified as significant intragenic rearrangement hotspots within the NF1 gene. However, no clear genotype-phenotype correlations could be identified among the NF1 patients carrying NF1 intragenic CNCs. Patient DNA samples with non-overlapping CNCs, as estimated by MLPA, were labeled with Cy3 and Cy5 fluorophores respectively, and hybridized onto the microarray. Alternatively, patient DNA was hybridized versus unrelated individual blood DNA. No hybridizations using biological replicates were performed. In total 6 samples were included. Please note that our experimental setup included a hybridization in which two DNA samples with non-overlapping deletions, from two NF1 patients, were hybridized in one experiment (sample codes: 1253 and 1403). In this specific case, assigning test or reference function to the samples was a matter of arbitrary choice. However, if needed, sample 1253 can be denoted as test, and sample 1403 can be denoted as reference in this experiment.
2015-07-30 | E-GEOD-64869 | biostudies-arrayexpress
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 cancer cells selectively promote translation of specific oncogenic transcripts to facilitate cancer survival and progression, while the underlying mechanisms are poorly understood. N7-methylguanosine (m7G) tRNA modification and its methyltransferase complex METTL1/WDR4 are significantly up-regulated in intrahepatic cholangiocarcinoma (ICC) and associated with poor prognosis. We developed tRNA reduction and cleavage sequencing (TRAC-Seq) to reveal the m7G tRNA methylome inICC cell line and ribosome nascent-chain complex-bound mRNAs sequencing(RNC-seq) and ribosome profiling(Ribo-seq) to study the differential translated genes and reveal the ribosome pausing. A subset of 22 tRNAs is modified at a ‘RAGGU’ motif within the variable loop. We observe increased ribosome occupancy at the corresponding codons in the Mettl1 knockdown ICC cell line implying widespread effects on tRNA function, ribosome pausing, and mRNA translation. Translation of cell cycle genes and EGFR signaling pathway genes is particularly affected. Our study uncovers the important physiological function and mechanism of METTL1-mediated m7G tRNA modification in the regulation of cancer progression.
2021-08-07 | GSE161319 | GEO
Project description:Effects of tRNA[Ser]Sec anticodon stem loop modifications on selenoprotein expression
| PRJNA762342 | ENA
Project description:The goal of this study is to investigate a chromatin-based mechanism that suppresses intragenic initiation of RNAPII transcription in A.thaliana. We demonstrate that RNAPII transcription across gene promoters represses their function in plants. The FACT histone chaperone complex is required for this repression mechanism by co-transcriptionally enforcing elevated levels of H3K4me1 at repressed intragenic promoters;
2018-04-26 | GSE113677 | 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:tRNAs are subject to numerous modifications including methylation. Mutations in the human N7-methylguanosine (m7G) methyltransferase complex METTL1-WDR4 cause primordial dwarfism and brain malformation yet the molecular and cellular function in mammals is not well understood. We developed m7G methylated tRNA immunoprecipitation sequencing (MeRIP-Seq) and tRNA reduction and cleavage sequencing (TRAC-Seq) to reveal the m7G tRNA methylome in mouse embryonic stem cells (mESCs). A subset of 22 tRNAs are modified at a ‘RAGGU’ motif within the variable loop. We observe increased ribosome occupancy at the corresponding codons in Mettl1 knockout mESCs implying widespread effects on tRNA function, ribosome pausing, and mRNA translation. Translation of cell cycle genes and those associated with brain abnormalities is particularly affected. Mettl1 or Wdr4 knockout mESCs display defective self-renewal and neural differentiation. Our study uncovers the complexity of the mammalian m7G tRNA methylome and highlights its essential role in ESCs with links to human disease.
2018-07-01 | GSE112670 | GEO