Rapid deadenylation triggered by a nonsense codon precedes decay of the RNA body in a mammalian cytoplasmic nonsense-mediated decay pathway.
Nonsense-mediated mRNA decay (NMD) is an RNA surveillance pathway that detects and destroys aberrant mRNAs containing nonsense or premature termination codons (PTCs) in a translation-dependent manner in eukaryotes. In yeast, the NMD pathway bypasses the deadenylation step and directly targets PTC-containing messages for decapping, followed by 5'-to-3' exonuclease digestion of the RNA body. In mammals, most PTC-containing mRNAs are subject to active nucleus-associated NMD. Here, using two distinct transcription-pulsing approaches to monitor mRNA deadenylation and decay kinetics, we demonstrate the existence of an active cytoplasmic NMD pathway in mammalian cells. In this pathway, a nonsense codon triggers accelerated deadenylation that precedes decay of the PTC-containing mRNA body. Transcript is stabilized when accelerated deadenylation is impeded by blocking translation initiation; by ectopically expressing two RNA-binding proteins, UNR and NSAP1; or by ectopically expressing a UPF1 dominant-negative mutant. These results are consistent with the notion that the nonsense codon can function in the cytoplasm by promoting rapid removal of the poly(A) tail as a necessary first step in the decay process.
Chen
Chyi-Ying A
CY
Department of Biochemistry and Molecular Biology, The University of Texas Houston Medical School, Houston, Texas 77030, USA.
Shyu
Ann-Bin
AB
D016475:3T3 Cells; D000249:Adenosine Monophosphate / Q000378:metabolism; D000818:Animals; D018389:Codon, Nonsense; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D005914:Globins / Q000235:genetics; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D008322:Mammals; D051379:Mice; D009154:Mutation; D014176:Protein Biosynthesis; D020365:RNA Helicases; D012323:RNA Processing, Post-Transcriptional / Q000502:physiology*; D020871:RNA Stability / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
C079950:CSDE1 protein, human; D018389:Codon, Nonsense; D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C486049:Syncrip protein, mouse; D015534:Trans-Activators; D014157:Transcription Factors; D000249:Adenosine Monophosphate; D005914:Globins; D020365:RNA Helicases; C101828:UPF1 protein, human