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Nanopore sequencing reveals endogenous NMD-targeted isoforms in human cells

ABSTRACT: Nanopore sequencing reveals endogenous NMD-targeted isoforms in human cells

PROVIDER: PRJEB44747 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR5880581.fastq.gz	Fastqsanger.gz
	ERR5880582.fastq.gz	Fastqsanger.gz
	ERR5880583.fastq.gz	Fastqsanger.gz
	ERR5880584.fastq.gz	Fastqsanger.gz
	ERR5880585.fastq.gz	Fastqsanger.gz

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Similar Datasets

Nanopore sequencing reveals endogenous NMD-targeted isoforms in human cells

Project description:Nonsense-mediated mRNA decay (NMD) is a eukaryotic, translation-dependent degradation pathway that targets mRNAs with premature termination codons and also regulates the expression of some mRNAs that encode full-length proteins. Although many genes express NMD-sensitive transcripts, identifying them based on short-read sequencing data remains a challenge. To identify and analyze endogenous targets of NMD, we applied cDNA Nanopore sequencing and short-read sequencing to human cells with varying expression levels of NMD factors. Our approach detects full-length NMD substrates that are highly unstable and increase in levels or even only appear when NMD is inhibited. Among the many new NMD-targeted isoforms that our analysis identified, most derive from alternative exon usage. The isoform-aware analysis revealed many genes with significant changes in splicing but no significant changes in overall expression levels upon NMD knockdown. NMD-sensitive mRNAs have more exons in the 3΄UTR and for mRNAs with a termination codon in the last exon. The length of the 3΄UTR per se does not correlate with NMD sensitivity. Analysis of splicing signals reveals isoforms where NMD has been co-opted in the regulation of gene expression, but a main function is most likely to rid the transcriptome of isoforms resulting from spurious splicing events. Long-read sequencing enabled the identification of many novel NMD-sensitive mRNAs and revealed both known and unexpected features concerning their biogenesis and their biological role. Our data provide a highly valuable resource of human NMD transcript targets for future genomic and transcriptomic applications.

2021-07-20 | E-MTAB-10452 | biostudies-arrayexpress

Nanopore sequencing reveals endogenous NMD-targeted isoforms in human cells.

Project description:BackgroundNonsense-mediated mRNA decay (NMD) is a eukaryotic, translation-dependent degradation pathway that targets mRNAs with premature termination codons and also regulates the expression of some mRNAs that encode full-length proteins. Although many genes express NMD-sensitive transcripts, identifying them based on short-read sequencing data remains a challenge.ResultsTo identify and analyze endogenous targets of NMD, we apply cDNA Nanopore sequencing and short-read sequencing to human cells with varying expression levels of NMD factors. Our approach detects full-length NMD substrates that are highly unstable and increase in levels or even only appear when NMD is inhibited. Among the many new NMD-targeted isoforms that our analysis identifies, most derive from alternative exon usage. The isoform-aware analysis reveals many genes with significant changes in splicing but no significant changes in overall expression levels upon NMD knockdown. NMD-sensitive mRNAs have more exons in the 3΄UTR and, for those mRNAs with a termination codon in the last exon, the length of the 3΄UTR per se does not correlate with NMD sensitivity. Analysis of splicing signals reveals isoforms where NMD has been co-opted in the regulation of gene expression, though the main function of NMD seems to be ridding the transcriptome of isoforms resulting from spurious splicing events.ConclusionsLong-read sequencing enables the identification of many novel NMD-sensitive mRNAs and reveals both known and unexpected features concerning their biogenesis and their biological role. Our data provide a highly valuable resource of human NMD transcript targets for future genomic and transcriptomic applications.

| S-EPMC8361881 | biostudies-literature

Genome-wide analysis of miRNA-targeted cellular NMD substrates in HeLa cell

Project description:Analysis of miRNA-targeted cellular NMD substrates in HeLa cell. The hypothesis tested in the present study was that endogenous NMD substrates containing long 3' untranslated region may targeted for miRNA. Results provide important information expanding the roles of miRISC in the posttranscriptional regulation of gene expression: a new cross-talk between miRNA-mediated gene silencing and NMD. ABSTRACT: Imperfect base-pairing between microRNA (miRNA) and the 3â-untranslated region (3âUTR) of target mRNA triggers translational repression of the target mRNA. Here, we provide evidence that human Argonaute 2 (Ago2) targets cap-binding protein (CBP)80/20- and exon junction complex (EJC)-bound mRNAs and inhibits nonsense-mediated mRNA decay (NMD), which is tightly restricted to CBP80/20-bound mRNAs. Furthermore, microarray analyses reveal that a subset of cellular transcripts, which are expected to be targeted for NMD, is stabilized by miRNA-mediated gene silencing. The regulation of NMD by miRNAs will shed light on a new post-transcriptional regulation mechanism of gene expression in mammalian cells Total RNA obtained from HeLa cells with downregulation of Ago2 or Ago2/UPF1 by siRNA. The up- or down-regulated transcripts were compared to control siRNA treated HeLa cell RNA extract. Significant transcripts were confirmed by replication.

2010-04-09 | E-GEOD-16170 | biostudies-arrayexpress

Genome-wide analysis of miRNA-targeted cellular NMD substrates in HeLa cell

Project description:Analysis of miRNA-targeted cellular NMD substrates in HeLa cell. The hypothesis tested in the present study was that endogenous NMD substrates containing long 3' untranslated region may targeted for miRNA. Results provide important information expanding the roles of miRISC in the posttranscriptional regulation of gene expression: a new cross-talk between miRNA-mediated gene silencing and NMD. ABSTRACT: Imperfect base-pairing between microRNA (miRNA) and the 3’-untranslated region (3’UTR) of target mRNA triggers translational repression of the target mRNA. Here, we provide evidence that human Argonaute 2 (Ago2) targets cap-binding protein (CBP)80/20- and exon junction complex (EJC)-bound mRNAs and inhibits nonsense-mediated mRNA decay (NMD), which is tightly restricted to CBP80/20-bound mRNAs. Furthermore, microarray analyses reveal that a subset of cellular transcripts, which are expected to be targeted for NMD, is stabilized by miRNA-mediated gene silencing. The regulation of NMD by miRNAs will shed light on a new post-transcriptional regulation mechanism of gene expression in mammalian cells

2010-03-08 | GSE16170 | GEO

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF2_NPCs]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF2_NPCs]

| PRJNA1097476 | ENA

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [pUPF1_RIPseq_ESCs]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [pUPF1_RIPseq_ESCs]

| PRJNA1097472 | ENA

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [Upf3b_mice]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [Upf3b_mice]

| PRJNA1097475 | ENA

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF2_ESCs]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF2_ESCs]

| PRJNA1097477 | ENA

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [pUPF1_RIPseq_NPCs]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [pUPF1_RIPseq_NPCs]

| PRJNA1097478 | ENA

Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF3B_NPCs]

Project description:Genome-wide identification of endogenous NMD-targeted RNAs in human stem cells [UPF3B_NPCs]

| PRJNA1097481 | ENA

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