Abstract
The human nuclear poly(A)-binding protein PABPN1 has been implicated in the decay of nuclear noncoding RNAs (ncRNAs). In addition, PABPN1 promotes hyperadenylation by stimulating poly(A)-polymerases (PAPα/γ), but this activity has not previously been linked to the decay of endogenous transcripts. Moreover, the mechanisms underlying target specificity have remained elusive. Here, we inactivated PAP-dependent hyperadenylation in cells by two independent mechanisms and used an RNA-seq approach to identify endogenous targets. We observed the upregulation of various ncRNAs, including snoRNA host genes, primary miRNA transcripts, and promoter upstream antisense RNAs, confirming that hyperadenylation is broadly required for the degradation of PABPN1-targets. In addition, we found that mRNAs with retained introns are susceptible to PABPN1 and PAPα/γ-mediated decay (PPD). Transcripts are targeted for degradation due to inefficient export, which is a consequence of reduced intron number or incomplete splicing. Additional investigation showed that a genetically-encoded poly(A) tail is sufficient to drive decay, suggesting that degradation occurs independently of the canonical cleavage and polyadenylation reaction. Surprisingly, treatment with transcription inhibitors uncouples polyadenylation from decay, leading to runaway hyperadenylation of nuclear decay targets. We conclude that PPD is an important mammalian nuclear RNA decay pathway for the removal of poorly spliced and nuclear-retained transcripts.
Highlights
Eukaryotic messenger RNAs undergo a series of maturation events before they are exported to the cytoplasm and translated
We and others reported that poly(A) tails may stimulate RNA decay in mammalian nuclei. This function is mediated by the concerted actions of the nuclear poly(A) binding protein PABPN1, poly(A)
A global decay function for PAP is validated by the analyses reported here, so we refer to this pathway as PABPN1 and PAPα/γmediated RNA decay (PPD)
Summary
Eukaryotic messenger RNAs (mRNAs) undergo a series of maturation events before they are exported to the cytoplasm and translated. The best-characterized RNA QC pathway is nonsense-mediated mRNA decay (NMD)[3]. NMD targets cytoplasmic mRNAs with premature termination codons (PTCs), a potentially dangerous class of RNAs that produce truncated and possibly dominant-negative proteins. NMD recognizes PTC-containing transcripts upon translation, so each defective transcript still produces one polypeptide. This could be harmful to cells for highly transcribed NMD targets or toxic polypeptides. NMD is stimulated by the presence of a splice junction to identify PTCs, so transcripts from intronless genes will generally not be recognized. Pervasive transcription produces nuclear transcripts that would not be targeted by the cytoplasmic NMD machinery
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