Acetylation of Cytidine in mRNA Promotes Translation Efficiency

Arango, Daniel; Sturgill, David; Alhusaini, Najwa; Dillman, Allissa; Sweet, Thomas J.; Hanson, Gavin; Hosogane, Masaki; Sinclair, Wilson R.; Nanan, Kyster K.; Mandler, Mariana D.; Fox, Stephen D.; Zengeya, Thomas; Andrésson, Þorkell; Meier, Jordan L.; Coller, Jeff; Oberdoerffer, Shalini

doi:10.1016/j.cell.2018.10.030

Cited by 530 publications

(764 citation statements)

References 39 publications

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“…28 NAT10 is an RNA acetyltransferase that has been found to catalyze acetylation of cytidine in ribosomal, transfer, and messenger RNA, forming the minor nucleobase N4-acetylcytidine (ac4C). [29][30][31] The identification of NAT10-CoA interactions by CATNIP is consistent with our previous studies, which have established that NAT10 binds acetyl-CoA and CoA with similar affinities and may be susceptible to metabolic feedback inhibition. 8 Compounding this effect, no related N4-acylations of cytidine (e.g.…”

Section: Evaluating the Dynamic Activity Of Acetyltransferases In Ressupporting

confidence: 90%

A systems chemoproteomic analysis of acyl-CoA/protein interaction networks

Levy

Montgomery

Sardiu

et al. 2019

Preprint

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CoA metabolite interactions, enabling the identification of biological processes susceptible to altered acetyl-CoA levels. Finally, we utilize systems-level analyses to assess the features of novel protein networks that may interact with acyl-CoAs and demonstrate a strategy for high-confidence annotation of direct acetyl-CoA binding proteins and AT enzymes in human proteomes. Overall our studies illustrate the power of integrating chemoproteomics and systems biology analysis methods and provide a novel resource for understanding the diverse signaling roles of acyl-CoAs in biology and disease. Results Validation of CATNIP for the global study of acyl-CoA/protein interactionsIn order to deeply sample acyl-CoA/protein interactions on a proteome-wide scale, we initially set out to integrate CoA-based protein capture methods with LC-MS/MS ( Fig. 2a). In this workflow, whole cell extracts are first incubated with Lys-CoA Sepharose. This affinity matrix enables active site-dependent enrichment of many different classes of CoA-binding proteins, 8 making it ideal for broad profiling studies. Next, enriched proteins are subjected to tryptic digest and analyzed using MudPIT (multidimensional protein identification technology), a proteomics platform that combines strong cation exchange and C18 reverse phase chromatography to pre-fractionate tryptic peptides, followed by ionization and data-dependent MS/MS. 9 The separation afforded by this approach significantly decreases sample complexity, allowing the identification of rare, low abundance peptides from complex proteomic mixtures. To facilitate the identification of acyl-CoA/protein interactions, competition experiments are performed in which proteomes are pre-incubated with a CoA metabolite prior to capture. 10 Decreased enrichment in competition samples compared to controls (as assessed by quantitative spectral counting) signifies that the CoA metabolite interacts with a protein of interest.These interacting proteins can then be further classified into pharmacological or biological networks using either conventional metrics (fold-change, gene ontology, etc) or systems-based analysis tools.As an initial model, we explored the utility of CATNIP to globally profile acetyl-CoA/protein interactions in unfractionated HeLa cell proteomes. Proteomes were pre-incubated with acetyl-CoA or vehicle (buffer) control, followed by enrichment using Lys-CoA Sepharose. These experiments assessed competition at 3, 30, and 300 µM acetyl-CoA, which spans the physiological concentration range of acetyl-CoA in the cytosol and mitochondria. Protein capture in each condition was quantified using distributed normalized spectral abundance factor (dNSAF), a label-free metric that normalizes spectral counts relative to overall protein length ( Fig. S1a-c , Table S1). 11 Each condition was analyzed in triplicate, constituting 12 experiments, >144 hours of instrument time, and over 1.1 million non-redundant peptide spectra collected. We limited our analysis to highconfidence protein identifications (>4...

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Section: Evaluating the Dynamic Activity Of Acetyltransferases In Ressupporting

confidence: 90%

A systems chemoproteomic analysis of acyl-CoA/protein interaction networks

Levy

Montgomery

Sardiu

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

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“…Furthermore, mRNA stability was decreased in NAT10‐deleted HeLa cells indicating that ac 4 C actively promotes mRNA stability and enhances translation. ac 4 C is strongly enriched in the 5′ end of mRNA, however, no effect of ac 4 C on the formation of 48S preinitiation complex was observed by in vitro analysis, demonstrating that ac 4 C does not affect translation initiation . Finally, ribosome‐profiling data show increased ribosome occupancy for acetylated mRNA mediated by NAT10, suggesting that ac 4 C intrinsically promotes translation, a finding further supported in vivo by the quantification of translation products from parental and NAT10‐depleted cells .…”

Section: Internal Mrna Modifications In Translational Regulationmentioning

confidence: 80%

“…Transcriptome‐wide mapping of the ac 4 C, catalysed by acetyltransferase NAT 10, revealed ac 4 C enrichment within coding regions of mRNA . Furthermore, mRNA stability was decreased in NAT10‐deleted HeLa cells indicating that ac 4 C actively promotes mRNA stability and enhances translation.…”

Section: Internal Mrna Modifications In Translational Regulationmentioning

confidence: 99%

The epitranscriptome in translation regulation: mRNA and tRNA modifications as the two sides of the same coin?

Ranjan

Leidel

2019

FEBS Letters

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Translation of mRNA is a highly regulated process that is tightly coordinated with cotranslational protein maturation. Recently, mRNA modifications and tRNA modifications – the so called epitranscriptome – have added a new layer of regulation that is still poorly understood. Both types of modifications can affect codon–anticodon interactions, thereby affecting mRNA translation and protein synthesis in similar ways. Here, we describe an updated view on how the different types of modifications can be mapped, how they affect translation, how they trigger phenotypes and discuss how the combined action of mRNA and tRNA modifications coordinate translation in health and disease.

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“…Recent advances in the field of RNA biology has led to the discovery of a plethora of coding and non-coding (nc) transcripts with novel modes of action, 1 new types of RNA epitranscriptomic modifications, 2,3 and an unforeseen diversity of mechanisms that lead to the production of RNAs. 4 These discoveries prompted the parallel development of modern molecular techniques that provide in-depth insights to the RNA structure and function.…”

Section: Introductionmentioning

confidence: 99%

Unveiling the druggable RNA targets and small molecule therapeutics

Sztuba-Solińska

Chávez-Calvillo

Cline

2019

Bioorganic & Medicinal Chemistry

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Keywords:RNA structure and function RNA interactions Epitranscriptomics Small molecules RNA-based therapeutics RNA in disease Drug target A B S T R A C T The increasing appreciation for the crucial roles of RNAs in infectious and non-infectious human diseases makes them attractive therapeutic targets. Coding and non-coding RNAs frequently fold into complex conformations which, if effectively targeted, offer opportunities to therapeutically modulate numerous cellular processes, including those linked to undruggable protein targets. Despite the considerable skepticism as to whether RNAs can be targeted with small molecule therapeutics, overwhelming evidence suggests the challenges we are currently facing are not outside the realm of possibility. In this review, we highlight the most recent advances in molecular techniques that have sparked a revolution in understanding the RNA structure-to-function relationship. We bring attention to the application of these modern techniques to identify druggable RNA targets and to assess small molecule binding specificity. Finally, we discuss novel screening methodologies that support RNA drug discovery and present examples of therapeutically valuable RNA targets. RNA as a drug targetThe vast diversity of RNAs expanding beyond coding transcripts to several classes of ncRNAs that vary in length, biogenesis, polarity, and putative functions, increases the repertoire of druggable targets. 19,20 Here, specific RNA properties contribute to its attractive, yet challenging makeup as a target molecule. RNA can form complex three-dimensional structures through canonical Watson-Crick base pairing and complex tertiary interactions that are mediated by non-canonical bonds. Such structures can be as intricate and stable as those formed by proteins and can recognize small-molecule ligands, other nucleic acids, and/or proteins with high affinity and specificity. [21][22][23][24] At the same time, the highly dynamic conformation and repetitive character of its surface presents difficulties for drug design. 25,26 In addition, many putative small molecule-binding pockets in RNA are much more polar and solvent exposed than binding sites on proteins, complicating ligand design efforts. Compounding these issues, most target RNAs are expressed at low levels, with the exception of ribosomal (rRNA) and transfer (tRNA) RNAs, which constitute 80-90% and 10-15% of total cellular RNA, respectively. 27 Other abundant RNAs, such as messenger (mRNA), small nuclear (snRNA), and small nucleolar (snoRNA) are present at levels that are about 1-2 orders of magnitude lower than rRNA and tRNA. Certain small RNAs, such as micro (miRNA) and piwi (piRNAs) can be present at very high levels; however, this appears to be cell type dependent. One should keep in mind that if the RNA has catalytic activity or if it acts as a scaffold to regulate https://doi.

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Acetylation of Cytidine in mRNA Promotes Translation Efficiency

Cited by 530 publications

References 39 publications

A systems chemoproteomic analysis of acyl-CoA/protein interaction networks

A systems chemoproteomic analysis of acyl-CoA/protein interaction networks

The epitranscriptome in translation regulation: mRNA and tRNA modifications as the two sides of the same coin?

Unveiling the druggable RNA targets and small molecule therapeutics

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