Rapid deadenylation triggered by a nonsense codon precedes decay of the RNA body in a mammalian cytoplasmic nonsense-mediated decay pathway

Mol Cell Biol. 2003 Jul;23(14):4805-13. doi: 10.1128/MCB.23.14.4805-4813.2003.

Abstract

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.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 3T3 Cells
  • Adenosine Monophosphate / metabolism
  • Animals
  • Codon, Nonsense*
  • Cytoplasm / genetics
  • Cytoplasm / metabolism*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Gene Expression Regulation
  • Genes, Dominant
  • Globins / genetics
  • Heterogeneous-Nuclear Ribonucleoproteins*
  • Mammals
  • Mice
  • Mutation
  • Protein Biosynthesis
  • RNA Helicases
  • RNA Processing, Post-Transcriptional / physiology*
  • RNA Stability / physiology
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism
  • Trans-Activators*
  • Transcription Factors / genetics
  • Transcription Factors / metabolism

Substances

  • CSDE1 protein, human
  • Codon, Nonsense
  • DNA-Binding Proteins
  • Heterogeneous-Nuclear Ribonucleoproteins
  • RNA, Messenger
  • RNA-Binding Proteins
  • Syncrip protein, mouse
  • Trans-Activators
  • Transcription Factors
  • Adenosine Monophosphate
  • Globins
  • RNA Helicases
  • UPF1 protein, human