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Concerted action of poly(A) nucleases and decapping enzyme in mammalian mRNA turnover

Nature Structural & Molecular Biology volume 12, pages 10541063 (2005) | Download Citation

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Abstract

In mammalian cells, the enzymatic pathways involved in cytoplasmic mRNA decay are incompletely defined. In this study, we have used two approaches to disrupt activities of deadenylating and/or decapping enzymes to monitor effects on mRNA decay kinetics and trap decay intermediates. Our results show that deadenylation is the key first step that triggers decay of both wild-type stable and nonsense codon–containing unstable β-globin mRNAs in mouse NIH3T3 fibroblasts. PAN2 and CCR4 are the major poly(A) nucleases active in cytoplasmic deadenylation that have biphasic kinetics, with PAN2 initiating deadenylation followed by CCR4-mediated poly(A) shortening. DCP2-mediated decapping takes place after deadenylation and may serve as a backup mechanism for triggering mRNA decay when initial deadenylation by PAN2 is compromised. Our findings reveal a functional link between deadenylation and decapping and help to define in vivo pathways for mammalian cytoplasmic mRNA decay.

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Acknowledgements

We thank R. Kulmacz, J. Lever and A. van Hoof for critical reading of the manuscript and their comments and Y. Tao for technical assistance with the heterokaryon experiments. This work was supported by a grant from the US National Institutes of Health (GM46454) to A.-B.S.

Author information

Author notes

    • Akio Yamashita
    •  & Tsung-Cheng Chang

    These authors contributed equally to this work.

Affiliations

  1. Department of Biochemistry and Molecular Biology, The University of Texas Medical School, Houston, Texas 77030, USA.

    • Akio Yamashita
    • , Tsung-Cheng Chang
    • , Yukiko Yamashita
    • , Wenmiao Zhu
    • , Zhenping Zhong
    • , Chyi-Ying A Chen
    •  & Ann-Bin Shyu

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The authors declare no competing financial interests.

Corresponding author

Correspondence to Ann-Bin Shyu.

Supplementary information

PDF files

  1. 1.

    Supplementary Fig. 1

    Catalytically inactive poly(A) nuclease mutants remain able to associate with their corresponding partners in vivo.

  2. 2.

    Supplementary Fig. 2

    Semi-log plots showing mRNA decay kinetics.

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DOI

https://doi.org/10.1038/nsmb1016

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