title: Temporal and spatial characterization of nonsense-mediated mRNA decay.
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 PTC-containing 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.
authors: Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
authors: Tatjana
authors: T
authors: Trcek
authors: Hanae
authors: H
authors: Sato
authors: Robert H
authors: RH
authors: Singer
authors: Lynne E
authors: LE
authors: Maquat
mesh_terms: D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D059365:Nonsense Mediated mRNA Decay; D012333:RNA, Messenger / Q000378:metabolism*; D013997:Time Factors; D055544:beta-Globins / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D055544:beta-Globins