Saccharomyces cerevisiae
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA355262 | ENA |
REPOSITORIES: ENA
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Project description:Accurate identification of NAD-capped RNAs is essential for understanding their biological function. Previous transcriptome-wide methods used to profile NAD-capped RNAs contain inherent limitations of having hindered the accurate identification of NAD caps from eukaryotic RNAs. Herein we introduced two novel orthogonal methods to precisely identify NAD-capped RNAs. One is D-SPAAC, a copper-free click-chemistry-based approach, and the second is an intramolecular ligation-based circNAD to resolve implicit limitations of the previous methods, which enabled us to unravel unforeseen features of NAD RNAs in budding yeast. Contrary to previous reports, we find that 1) cellular NAD RNAs can be full-length and polyadenylated transcripts, 2) transcription start sites for NAD-capped and canonical m7G-capped RNAs are different, and 3) NAD caps can be added post-transcriptionally. Moreover, we uncovered a dichotomy of NAD RNAs in translation where NAD RNAs are detected with mitochondrial ribosomes but not cytoplasmic ribosomes indicating their propensity to be translated in mitochondria.
2023-03-17 | GSE217259 | GEO
Project description:We report the identification of NAD-RNA in the yeast Saccharomyces cerevisiae under two distinct growth mediums.
2016-11-30 | GSE90636 | GEO
Project description:The 5’ end of a eukaryotic mRNA generally has a methyl guanosine cap (m7G cap) that not only protects the mRNA from degradation but also mediates almost all other aspects of gene expression. Some RNAs in E. coli, yeast, and mammals were recently found to contain an NAD+ cap at their 5’ ends. Here we report development of a new method – NAD tagSeq – for transcriptome-wide identification and quantification of NAD+-capped RNAs (NAD-RNAs). The method uses first an enzymatic reaction and then a click chemistry reaction to label NAD-RNAs with a synthetic RNA tag. The tagged RNA molecules can be enriched and directly sequenced using the Oxford Nanopore sequencing technology. NAD tagSeq not only allows more accurate identification and quantification of NAD-RNAs but can also reveal sequences of whole NAD-RNA transcripts. Using NAD tagSeq, we found that NAD-RNAs in Arabidopsis are mostly produced from a few thousand protein-coding genes, with over 60% of them from fewer than 200 genes. The top 2,000 genes that were found to produce the highest numbers of NAD-RNAs were enriched in the gene ontology terms of responses to oxidative stress and other stresses, photosynthesis, and protein synthesis. For some Arabidopsis genes, over 10% of their transcripts could be NAD-capped. The NAD-RNAs in Arabidopsis have similar overall sequence structures to their canonical m7G-capped mRNAs. The identification and quantification of NAD-RNAs and revealing their sequence features provide essential steps toward understanding functions of NAD-RNAs.
2019-07-01 | GSE127755 | GEO
Project description:As the most common mRNA cap, the m7G cap impacts the fate of an mRNA in eukaryotes. The metabolite and redox agent, nicotinamide adenine diphosphate (NAD+), can be used as an initiating nucleotide in RNA synthesis to result in NAD+-capped RNAs. Such RNAs have been identified in bacteria, yeast, and human cells, but it is not known whether they exist in plant transcriptomes. The functions of the NAD+ cap in RNA metabolism or translation are still poorly understood. Here, through NAD captureSeq, we show that NAD+- capped RNAs are widespread in Arabidopsis thaliana. NAD+-capped RNAs are predominantly messenger RNAs encoded by the nuclear and mitochondrial genomes but not the chloroplast genome. NAD-capped transcripts from the nuclear genome appear to be spliced and polyadenylated. Furthermore, although NAD+-capped transcripts constitute a small proportion of the total transcript pool from any gene, they are enriched in the polysomal fraction and associate with translating ribosomes. Our findings implicate the existence of as yet unknown mechanisms of translation initiation on NAD+-capped mRNAs. More importantly, our findings suggest that cellular metabolic and/or redox states may influence, and maybe regulated by, mRNA NAD capping.
2019-06-03 | GSE127002 | GEO
Project description:Eukaryotic messenger RNAs (mRNAs) possess a 5’-end N7-methyl guanosine (m7G) cap that promotes their translation and stability. However, it was recently demonstrated that eukaryotic mRNAs can also carry a 5' end nicotinamide adenine dinucleotide (NAD+) cap that promotes mRNA decay mediated by the NAD+ decapping enzyme DXO1. However, the dynamic regulation of NAD+ capping in plant remains unknown. Here, we describe the global landscape of NAD+-capped RNAs in Arabidopsis thaliana, and demonstrate that DXO1 is responsible for removal of these 5’-end modifications and facilitates mRNA degradation in plant transcriptomes. We also reveal that in the absence of DXO1 NAD+-capped mRNAs are unstable and processed into smRNAs. Furthermore, we find that Abscisic Acid (ABA) remodel the landscape of RNA cap epitransciptome, and the mRNA lost their NAD+ cap contribute to their stability under ABA. Overall, our results support a link between ABA response and RNA NAD+ capping.
2020-11-23 | GSE142389 | GEO
Project description:Eukaryotic messenger RNAs (mRNAs) possess a 5’-end N7-methyl guanosine (m7G) cap that promotes their translation and stability. However, it was recently demonstrated that eukaryotic mRNAs can also carry a 5' end nicotinamide adenine dinucleotide (NAD+) cap that promotes mRNA decay mediated by the NAD+ decapping enzyme DXO1. However, the dynamic regulation of NAD+ capping in plant remains unknown. Here, we describe the global landscape of NAD+-capped RNAs in Arabidopsis thaliana, and demonstrate that DXO1 is responsible for removal of these 5’-end modifications and facilitates mRNA degradation in plant transcriptomes. We also reveal that in the absence of DXO1 NAD+-capped mRNAs are unstable and processed into smRNAs. Furthermore, we find that Abscisic Acid (ABA) remodel the landscape of RNA cap epitransciptome, and the mRNA lost their NAD+ cap contribute to their stability under ABA. Overall, our results support a link between ABA response and RNA NAD+ capping.
2020-11-23 | GSE142388 | GEO
Project description:Spliced messages constitute one-fourth of expressed mRNAs in the yeast Saccharomyces cerevisiae, and most mRNAs in metazoans. Splicing requires 5' splice site (5'SS), branch point (BP), and 3' splice site (3'SS) elements, but the role of the BP in splicing control is poorly understood because BP identification remains difficult. We developed a high-throughput method, Branch-seq, to map BP and 5'SS of isolated RNA lariats. Applied to S. cerevisiae, Branch-seq detected 76% of expressed, annotated BPs and identified a comparable number of novel BPs. We used RNA-seq to confirm associated 3'SS locations, identifying some 200 novel splice junctions, including an AT-AC intron. We show that several yeast introns use two or even three different BPs, with effects on 3'SS choice, protein coding potential, or RNA stability and identify novel introns whose splicing changes during meiosis or in response to stress. Together, these findings reveal BP-based regulation and demonstrate unanticipated complexity of splicing in yeast.
2016-08-01 | GSE68022 | GEO
Project description:The hub metabolite, nicotinamide adenine dinucleotide (NAD), can be used as an initiating nucleotide in RNA synthesis to result in NAD-capped RNAs (NAD-RNA). Since NAD has been heightened as one of the most essential modulators in aging and various age-related diseases, its attachment to RNA might indicate a yet-to-be discovered mechanism that impacts adult life-course. However, the unknown identity of NAD-linked RNAs in adult and aging tissues has hindered functional studies. Here, we introduce ONE-seq method to identify the RNA transcripts that contain NAD cap. ONE-seq has been optimized to use only one-step chemo-enzymatic biotinylation, followed by streptavidin capture and the nudix phosphohydrolase NudC-catalyzed elution, to specifically recover NAD-capped RNAs for epitranscriptome and gene-specific analyses. Our data describes more than a thousand of previously unknown NAD-RNAs in the mouse liver and reveals epitranscriptome-wide dynamics of NAD-RNAs with age.ONE-seq empowers the identification of NAD-capped RNAs that are responsive to distinct physiological states, facilitating functional investigation into this modification.
2022-01-27 | GSE194271 | GEO
Project description:The 5´-end 7-methylguanosine cap structure has long been known as a signature feature of eukaryotic cellular and viral mRNAs that confers mRNA stability and efficient translation. Recent findings in diverse organisms have demonstrated that RNAs can additionally possess a non-canonical cap structure consisting of a nicotinamide adenosine dinucleotide (NAD+) at their 5´ end in place of m7G. It has been shown that 5´ end-NAD+ cap promotes rapid decay of the RNA at least in part by the DXO family of proteins in mammalian cells. This observation led to the hypothesis that mammalian cells harbor additional deNADding enzymes that may function in distinct pathways. Here we report Nudt12 efficiently removes NAD+ caps and functions as alternate cellular deNADding enzyme that targets NAD+-capped RNAs distinct from DXO. Importantly, with the use of an NAD-Cap Detection (NAD-CapD) approach that utilizes enzymatic properties to release intact NAD+/NADH from the 5´ end of NAD-capped cellular RNAs and a colorimetric NAD Quantitation to detect released NAD+/NADH, we can follow total cellular NAD+ cap levels. Removal of Nudt12 or DXO deNADding enzymes from cells significantly increased levels of NAD+-capped cellular RNAs. Moreover, fungal Rai1 and Dxo1, previously demonstrated to possess deNADding activity in vitro, can also function as deNADding enzymes in yeast cells. Double disruption of Rai1 and Dxo1 in yeast cells lead to accumulation of NAD+-capped RNAs, indicating that both enzymes function to clear NAD+ from the 5´ end of RNAs. Finally, our findings established that alterations in cellular NAD+ levels impact NAD+-capped RNA levels implying NAD+ capping is a modulated process that may be linked to the metabolic state of the cell.
2019-05-20 | GSE110801 | GEO
Project description:This project aims to identify novel RNA binding proteins in the baker's yeast, Saccharomyces cerevisiae. Since interactions between RNAs and proteins may be transient, yeast cells were crosslinked with UV light at 254 nm which promotes the covalent link between proteins and RNAs. After this, polyadenylated mRNAs were purified via oligo(dT) coupled to magentic beads under stringet conditions. Finally, samples were subjected to mass spectrometry analysis. To rule out the possibility of RNA-independent binding we also analysed other samples: i) samples digested with RNase one; ii) samples where we performed competition assays with polyadenylic acid.
2015-10-26 | PXD002293 | Pride