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CEG1 DEPLETION REVEALS MECHANISMS GOVERNING DEGRADATION OF NON-CAPPED RNAs

ABSTRACT: CEG1 DEPLETION REVEALS MECHANISMS GOVERNING DEGRADATION OF NON-CAPPED RNAs

PROVIDER: PRJNA883220 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR21675721_1.fastq.gz	Fastqsanger.gz
	SRR21675721_2.fastq.gz	Fastqsanger.gz
	SRR21675722_1.fastq.gz	Fastqsanger.gz
	SRR21675722_2.fastq.gz	Fastqsanger.gz
	SRR21675723_1.fastq.gz	Fastqsanger.gz

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CEG1 DEPLETION REVEALS MECHANISMS GOVERNING DEGRADATION OF NON-CAPPED RNAs

Project description:All eukaryotic mRNAs contain a 7-methylguanosine (m7G) cap which serves as a platform that recruits proteins to support essential biological functions such as mRNA processing, nuclear export and cap-dependent translation. Although the caping is one of the first steps of transcription and uncapped mRNA is not functional, the fate and turnover of uncapped transcripts have not been studied extensively. Here, we employed fast nuclear depletion of the capping enzymes in yeast Saccharomyces cerevisiae to uncover the turnover of transcripts that failed to be capped. We found that the degradation of non-capped mRNA is mainly performed by Xrn1, the exonuclease which is predominantly localized in the cytoplasm, and the fate of such transcripts is determined principally by the abundance of their synthesis. Nuclear depletion of the capping enzymes increased the levels of poorly expressed mRNAs and non-coding RNAs and did not affect the distribution of RNA Polymerase II on chromatin. Overall, our data indicate that mRNAs that failed to be capped are not directed to any specific quality-control pathway and are stochastically degraded.

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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.

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