Cotranscriptional effect of a premature termination codon revealed by live-cell imaging

Valeria De Turris, Pamela Nicholson, Rodolfo Zamudio Orozco, Robert H. Singer, Oliver Mühlemann

Research output: Contribution to journalArticle

34 Scopus citations

Abstract

Aberrant mRNAs with premature translation termination codons (PTCs) are recognized and eliminated by the nonsensemediated mRNA decay (NMD) pathway in eukaryotes. We employed a novel live-cell imaging approach to investigate the kinetics of mRNA synthesis and release at the transcription site of PTC-containing (PTC+) and PTC-free (PTC-) immunoglobulin-μ reporter genes. Fluorescence recovery after photobleaching (FRAP) and photoconversion analyses revealed that PTC+ transcripts are specifically retained at the transcription site. Remarkably, the retained PTC+ transcripts are mainly unspliced, and this RNA retention is dependent upon two important NMD factors, UPF1 and SMG6, since their depletion led to the release of the PTC+ transcripts. Finally, ChIP analysis showed a physical association of UPF1 and SMG6 with both the PTC+ and the PTC- reporter genes in vivo. Collectively, our data support a mechanism for regulation of PTC+ transcripts at the transcription site. Published by Cold Spring Harbor Laboratory Press.

Original languageEnglish (US)
Pages (from-to)2094-2107
Number of pages14
JournalRNA
Volume17
Issue number12
DOIs
StatePublished - Dec 1 2011

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Keywords

  • Live-cell imaging
  • NMD
  • Retention
  • SMG6
  • Splicing
  • UPF1

ASJC Scopus subject areas

  • Molecular Biology

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