Format

Send to

Choose Destination
See comment in PubMed Commons below
PLoS One. 2013 Sep 25;8(9):e74791. doi: 10.1371/journal.pone.0074791. eCollection 2013.

SMG6 cleavage generates metastable decay intermediates from nonsense-containing β-globin mRNA.

Author information

1
Center for RNA Biology, The Ohio State University, Columbus, Ohio, United States of America ; Department of Molecular and Cellular Biochemistry, The Ohio State University, Columbus, Ohio, United States of America.

Abstract

mRNAs targeted by endonuclease decay generally disappear without detectable decay intermediates. The exception to this is nonsense-containing human β-globin mRNA, where the destabilization of full-length mRNA is accompanied by the cytoplasmic accumulation of 5'-truncated transcripts in erythroid cells of transgenic mice and in transfected erythroid cell lines. The relationship of the shortened RNAs to the decay process was characterized using an inducible erythroid cell system and an assay for quantifying full-length mRNA and a truncated RNA missing 169 nucleotides from the 5' end. In cells knocked down for Upf1 a reciprocal increase in full-length and decrease in shortened RNA confirmed the role of NMD in this process. Kinetic analysis demonstrated that the 5'-truncated RNAs are metastable intermediates generated during the decay process. SMG6 previously was identified as an endonuclease involved in NMD. Consistent with involvement of SMG6 in the decay process full-length nonsense-containing β-globin mRNA was increased and the Δ169 decay intermediate was decreased in cells knocked down for SMG6. This was reversed by complementation with siRNA-resistant SMG6, but not by SMG6 with inactivating PIN domain mutations. Importantly, none of these altered the phosphorylation state of Upf1. These data provide the first proof for accumulation of stable NMD products by SMG6 endonuclease cleavage.

PMID:
24086375
PMCID:
PMC3783490
DOI:
10.1371/journal.pone.0074791
[Indexed for MEDLINE]
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Public Library of Science Icon for PubMed Central
    Loading ...
    Support Center