Temporal and spatial characterization of nonsense-mediated mRNA decay

Tatjana Trcek, Hanae Sato, Robert H. Singer, Lynne E. Maquat

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

Nonsense-mediated mRNA decay (NMD) is a quality control mechanism responsible for "surveying" mRNAs during translation and degrading those that harbor a premature termination codon (PTC). Currently the intracellular spatial location of NMD and the kinetics of its decay step in mammalian cells are under debate. To address these issues, we used single-RNA fluorescent in situ hybridization (FISH) and measured the NMD of PTCcontaining β-globin mRNA in intact single cells after the induction of β-globin gene transcription. This approach preserves temporal and spatial information of the NMD process, both of which would be lost in an ensemble study. We determined that decay of the majority of PTC-containing β-globin mRNA occurs soon after its export into the cytoplasm, with a half-life of <1 min; the remainder is degraded with a half-life of >12 h, similar to the half-life of normal PTC-free β-globin mRNA, indicating that it had evaded NMD. Importantly, NMD does not occur within the nucleoplasm, thus countering the long-debated idea of nuclear degradation of PTC-containing transcripts. We provide a spatial and temporal model for the biphasic decay of NMD targets.

Original languageEnglish (US)
Pages (from-to)541-551
Number of pages11
JournalGenes and Development
Volume27
Issue number5
DOIs
StatePublished - Mar 1 2013

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Nonsense Mediated mRNA Decay
Globins
Nonsense Codon
Messenger RNA
Half-Life
Protein Biosynthesis
Fluorescence In Situ Hybridization
Quality Control
Cytoplasm
RNA

Keywords

  • mRNA decay
  • mRNA quality control
  • Single-RNA FISH

ASJC Scopus subject areas

  • Genetics
  • Developmental Biology

Cite this

Temporal and spatial characterization of nonsense-mediated mRNA decay. / Trcek, Tatjana; Sato, Hanae; Singer, Robert H.; Maquat, Lynne E.

In: Genes and Development, Vol. 27, No. 5, 01.03.2013, p. 541-551.

Research output: Contribution to journalArticle

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