Time-resolved NMR monitoring of tRNA maturation

Gato, Alexandre; Heiss, MM; Catala, Marjorie; Kellner, Stefanie; Tisné, Carine

doi:10.1101/588814

Cited by 9 publications

(35 citation statements)

References 66 publications

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“…LC-MS/MS analyses can reproducibly quantify individual RNA modifications in purified tRNAs as shown by analysing six LC-MS/MS measurements on commercially available tRNA-Phe from Saccharomyces cerevisiae (supplementary Figure 3A and B confirming the analysis of S.c . tRNA-Phe published in [ 53 ]). 5ʹ tsRNA-Gly GCC/CCC and 5ʹ tsRNA-Glu CUC/UUC along with their remaining parental full-length tRNAs were purified from biological triplicate experiments after either iAs exposure or ecANG expression followed by LC-MS/MS analyses ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Production and purification of endogenously modified tRNA-derived small RNAs

et al. 2020

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2020): Production and purification of endogenously modified tRNA-derived small RNAs, RNA Biology, ABSTRACT During particular stress conditions, transfer RNAs (tRNAs) become substrates of stress-induced endonucleases, resulting in the production of distinct tRNA-derived small RNAs (tsRNAs). These small RNAs have been implicated in a wide range of biological processes, but how isoacceptor and even isodecoder-specific tsRNAs act at the molecular level is still poorly understood. Importantly, stress-induced tRNA cleavage affects only a few tRNAs of a given isoacceptor or isodecoder, raising the question as to how such limited molecule numbers could exert measurable biological impact. While the molecular function of individual tsRNAs is likely mediated through association with other molecules, addressing the interactome of specific tsRNAs has only been attempted by using synthetic RNA sequences. Since tRNAs carry post-transcriptional modifications, tsRNAs are likely modified but the extent of their modifications remains largely unknown. Here, we developed a biochemical framework for the production and purification of specific tsRNAs using human cells. Preparative scale purification of tsRNAs from biological sources should facilitate experimentally addressing as to how exactly these small RNAs mediate the multitude of reported molecular functions. ARTICLE HISTORY

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Section: Resultsmentioning

confidence: 99%

Production and purification of endogenously modified tRNA-derived small RNAs

et al. 2020

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“…This might be favored by an incomplete set of modifications within Trm11 tRNA substrates, which may increase their intrinsic flexibility. Indeed, a recent study aimed at monitoring the addition of tRNA modifications using yeast cell extracts revealed that many of those occur in the T-arm, either concomitantly with m 2 G 10 or even later ( 72 ). This model further supports the ruler model first proposed for ThiI and then for Trm11 and TrmN/Trm14 MTases ( 38 , 42 ).…”

Section: Discussionmentioning

confidence: 99%

Structural and functional insights into Archaeoglobus fulgidus m2G10 tRNA methyltransferase Trm11 and its Trm112 activator

Wang

Tran

Jactel

et al. 2020

Nucleic Acids Research

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tRNAs play a central role during the translation process and are heavily post-transcriptionally modified to ensure optimal and faithful mRNA decoding. These epitranscriptomics marks are added by largely conserved proteins and defects in the function of some of these enzymes are responsible for neurodevelopmental disorders and cancers. Here, we focus on the Trm11 enzyme, which forms N2-methylguanosine (m2G) at position 10 of several tRNAs in both archaea and eukaryotes. While eukaryotic Trm11 enzyme is only active as a complex with Trm112, an allosteric activator of methyltransferases modifying factors (RNAs and proteins) involved in mRNA translation, former studies have shown that some archaeal Trm11 proteins are active on their own. As these studies were performed on Trm11 enzymes originating from archaeal organisms lacking TRM112 gene, we have characterized Trm11 (AfTrm11) from the Archaeoglobus fulgidus archaeon, which genome encodes for a Trm112 protein (AfTrm112). We show that AfTrm11 interacts directly with AfTrm112 similarly to eukaryotic enzymes and that although AfTrm11 is active as a single protein, its enzymatic activity is strongly enhanced by AfTrm112. We finally describe the first crystal structures of the AfTrm11-Trm112 complex and of Trm11, alone or bound to the methyltransferase inhibitor sinefungin.

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“…tRNAs are the most heavily modified RNAs (1) . tRNAs are modified post-transcriptionally and the biosynthesis of modified nucleosides often requires different modification enzymes acting sequentially at distinct steps of tRNA maturation (2,3) . The complex mechanisms underlying the stepwise modification of tRNAs were largely deciphered in the yeast S. cerevisiae , as well as in studies conducted in prokaryotes and Archaea .…”

Section: Introductionmentioning

confidence: 99%

tRNA 2’-O-methylation modulates small RNA silencing and life span in Drosophila

Angelova

Dimitrova

Silva

et al. 2019

Preprint

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2'-O-methylation (Nm) represents one of the most common RNA modifications. Nm affects RNA structure and function with crucial roles in various RNA-mediated processes ranging from RNA silencing, translation, self versus non-self recognition to viral defense mechanisms. Here, we identify two novel Nm methyltransferases (Nm-MTases) in Drosophila melanogaster (CG7009 and CG5220) as functional orthologs of yeast TRM7 and human FTSJ1, respectively. Genetic knockout studies together with MALDI-TOF mass spectrometry and RiboMethSeq mapping revealed that CG7009 is responsible for methylating the wobble position in tRNA Phe , tRNA Trp and tRNA Leu , while subsequently, CG5220 methylates position C32 in the same tRNAs and targets also additional tRNAs. CG7009 or CG5220 mutant animals were viable and fertile but exhibited various phenotypes such as life span reduction, small RNA pathways dysfunction and increased sensitivity to RNA virus infections. Our results provide the first detailed characterization of two TRM7 family members in Drosophila and uncover a molecular link between enzymes catalysing Nm at specific tRNAs and small RNA-induced gene silencing pathways.

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Time-resolved NMR monitoring of tRNA maturation

Cited by 9 publications

References 66 publications

Production and purification of endogenously modified tRNA-derived small RNAs

Production and purification of endogenously modified tRNA-derived small RNAs

Structural and functional insights into Archaeoglobus fulgidus m2G10 tRNA methyltransferase Trm11 and its Trm112 activator

tRNA 2’-O-methylation modulates small RNA silencing and life span in Drosophila

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