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Analysis of RNA decay factor mediated RNA stability contributions on RNA abundance.

ABSTRACT: Histone epigenome data determined by chromatin immunoprecipitation sequencing (ChIP-seq) is used in identifying transcript regions and estimating expression levels. However, this estimation does not always correlate with eventual RNA expression levels measured by RNA sequencing (RNA-seq). Part of the inconsistency may arise from the variance in RNA stability, where the transcripts that are more or less abundant than predicted RNA expression from histone epigenome data are inferred to be more or less stable. However, there is little systematic analysis to validate this assumption. Here, we used stability data of whole transcriptome measured by 5'-bromouridine immunoprecipitation chase sequencing (BRIC-seq), which enabled us to determine the half-lives of whole transcripts including lincRNAs, and we integrated BRIC-seq with ChIP-seq to achieve better estimation of the eventual transcript levels and to understand the importance of post-transcriptional regulation that determine the eventual transcript levels.We identified discrepancies between the RNA abundance estimated by ChIP-seq and measured RNA expression from RNA-seq; for number of genes and estimated that the expression level of 865 genes was controlled at the level of RNA stability in HeLa cells. ENCODE data analysis supported the idea that RNA stability control aids to determine transcript levels in multiple cell types. We identified UPF1, EXOSC5 and STAU1, well-studied RNA degradation factors, as controlling factors for 8% of cases. Computational simulations reasonably explained the changes of eventual mRNA levels attributable to the changes in the rates of mRNA half-lives. In addition, we propose a feedback circuit that includes the regulated degradation of mRNAs encoding transcription factors to maintain the steady state level of RNA abundance. Intriguingly, these regulatory mechanisms were distinct between mRNAs and lincRNAs.Integrative analysis of ChIP-seq, RNA-seq and our BRIC-seq showed that transcriptional regulation and RNA degradation are independently regulated. In addition, RNA stability is an important determinant of eventual transcript levels. RNA binding proteins, such as UPF1, STAU1 and EXOSC5 may play active roles in such controls.

SUBMITTER: Maekawa S

PROVIDER: S-EPMC4359779 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Analysis of RNA decay factor mediated RNA stability contributions on RNA abundance.

Maekawa Sho S Imamachi Naoto N Irie Takuma T Tani Hidenori H Matsumoto Kyoko K Mizutani Rena R Imamura Katsutoshi K Kakeda Miho M Yada Tetsushi T Sugano Sumio S Suzuki Yutaka Y Akimitsu Nobuyoshi N

BMC genomics 20150306

<h4>Background</h4>Histone epigenome data determined by chromatin immunoprecipitation sequencing (ChIP-seq) is used in identifying transcript regions and estimating expression levels. However, this estimation does not always correlate with eventual RNA expression levels measured by RNA sequencing (RNA-seq). Part of the inconsistency may arise from the variance in RNA stability, where the transcripts that are more or less abundant than predicted RNA expression from histone epigenome data are infe ...[more]

PMID: 25879614

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Project description:Nonsense-mediated decay (NMD) is a host RNA control pathway that removes aberrant transcripts with long 3' untranslated regions (UTRs) due to premature termination codons (PTCs) that arise through mutation or defective splicing. To maximize coding potential, RNA viruses often contain internally located stop codons that should also be prime targets for NMD. Using an agroinfiltration-based NMD assay in Nicotiana benthamiana, we identified two segments conferring NMD-resistance in the carmovirus Turnip crinkle virus (TCV) genome. The ribosome readthrough structure just downstream of the TCV p28 termination codon stabilized an NMD-sensitive reporter as did a frameshifting element from umbravirus Pea enation mosaic virus. In addition, a 51-nt unstructured region (USR) at the beginning of the TCV 3' UTR increased NMD-resistance 3-fold when inserted into an unrelated NMD-sensitive 3' UTR. Several additional carmovirus 3' UTRs also conferred varying levels of NMD resistance depending on the construct despite no sequence similarity in the analogous region. Instead, these regions displayed a marked lack of RNA structure immediately following the NMD-targeted stop codon. NMD-resistance was only slightly reduced by conversion of 19 pyrimidines in the USR to purines, but resistance was abolished when a 2-nt mutation was introduced downstream of the USR that substantially increased the secondary structure in the USR through formation of a stable hairpin. The same 2-nt mutation also enhanced the NMD susceptibility of a subgenomic RNA expressed independently of the genomic RNA. The conserved lack of RNA structure among most carmoviruses at the 5' end of their 3' UTR could serve to enhance subgenomic RNA stability, which would increase expression of the encoded capsid protein that also functions as the RNA silencing suppressor. These results demonstrate that the TCV genome has features that are inherently NMD-resistant and these strategies could be widespread among RNA viruses and NMD-resistant host mRNAs with long 3' UTRs.

Dataset Information

Analysis of RNA decay factor mediated RNA stability contributions on RNA abundance.

Publications

Analysis of RNA decay factor mediated RNA stability contributions on RNA abundance.

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