Letter | Published:

DENR–MCT-1 promotes translation re-initiation downstream of uORFs to control tissue growth

Nature volume 512, pages 208212 (14 August 2014) | Download Citation

Abstract

During cap-dependent eukaryotic translation initiation, ribosomes scan messenger RNA from the 5′ end to the first AUG start codon with favourable sequence context1,2. For many mRNAs this AUG belongs to a short upstream open reading frame (uORF)3, and translation of the main downstream ORF requires re-initiation, an incompletely understood process1,4,5,6. Re-initiation is thought to involve the same factors as standard initiation1,5,7. It is unknown whether any factors specifically affect translation re-initiation without affecting standard cap-dependent translation. Here we uncover the non-canonical initiation factors density regulated protein (DENR) and multiple copies in T-cell lymphoma-1 (MCT-1; also called MCTS1 in humans) as the first selective regulators of eukaryotic re-initiation. mRNAs containing upstream ORFs with strong Kozak sequences selectively require DENR–MCT-1 for their proper translation, yielding a novel class of mRNAs that can be co-regulated and that is enriched for regulatory proteins such as oncogenic kinases. Collectively, our data reveal that cells have a previously unappreciated translational control system with a key role in supporting proliferation and tissue growth.

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Accessions

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Gene Expression Omnibus

Data deposits

Polysome microarray data are deposited at NCBI GEO under accession number GSE54625.

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Acknowledgements

We thank B. Bukau, R. Green and P. Soba for suggestions on the manuscript, V. Benes and T. Bähr-Ivacevic (EMBL Genomics Core Facility) for assistance with microarray experiments, S. Abmayr for anti-Mbc antibody, T. Hsu for anti-Awd antibody, C. Strein and M. Hentze for Drosophila DENR antibodies, P. Jakob for help with testing conditions for S2 in vitro translation extracts, L. Schibalski and J. Grawe for help with cloning, S. Hofmann for help with polysome analysis, and S. Lerch, A. Haffner and M. Schröder for technical assistance. K.K.M. is the recipient of an Alzheimer’s Research Scholarship from the Hans und Ilse Breuer Foundation. P.C.J. is supported in part by a grant from the Fritz Thyssen Foundation to K.E.D. This work was supported in part by a Deutsche Forschungsgemeinschaft (DFG) grant and ERC Starting Grant to A.A.T.

Author information

Author notes

    • Katrin Strassburger
    • , Philipp Christoph Janiesch
    • , Kent E. Duncan
    •  & Aurelio A. Teleman

    These authors contributed equally to this work.

Affiliations

  1. German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany

    • Sibylle Schleich
    • , Katrin Strassburger
    • , Katharina Haneke
    • , Yong-Sheng Cheng
    • , Georg Stoecklin
    •  & Aurelio A. Teleman
  2. Center for Molecular Neurobiology (ZMNH), University Medical Center Hamburg-Eppendorf (UKE), Falkenried 94, 20251 Hamburg, Germany

    • Sibylle Schleich
    • , Philipp Christoph Janiesch
    • , Tatyana Koledachkina
    • , Katharine K. Miller
    • , Katrin Küchler
    •  & Kent E. Duncan
  3. Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), DKFZ-ZMBH Alliance, 69120 Heidelberg, Germany

    • Katharina Haneke
    •  & Georg Stoecklin

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Contributions

S.S. performed most experiments, except as indicated below. K.S. analysed histoblast proliferation, EcR and InR protein levels and signalling in cells and animals, and performed EcR/InR rescue experiments in vivo. P.C.J. assessed proliferation, translation, rRNA and tRNA levels, as well as polysome profiles and genome-wide polysomal profiling of DENR knockdown S2 cells, and performed in vitro translation assays. T.K. established and performed inducible reporter assays in proliferating and quiescent cells (Extended Data Figs 7a–f and 10c). K.K.M. performed DENR–MCT-1 co-immunoprecipitation assays (Extended Data Fig. 10b). K.H. performed DENR RNA immunoprecipitation assays (Extended Data Fig. 4g). Y.-S.C. analysed MCT-1 levels and phosphorylation (Extended Data Fig. 10d′). K.K. established in vitro translation assays from S2 cells. A.A.T. performed the bioinformatic analyses. K.E.D. and A.A.T. jointly designed and coordinated the study and wrote the paper with input from G.S. All authors interpreted data, discussed results and contributed to writing the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Kent E. Duncan or Aurelio A. Teleman.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains a Supplementary Discussion, a complete list of oligos and sequences used, and the number of replicates for each figure panel.

Excel files

  1. 1.

    Supplementary Table 1

    Microarray profiling of polysome gradients from control and DENR knockdown cells.

  2. 2.

    Supplementary Table 2

    stuORF scores for all transcripts.

  3. 3.

    Supplementary Table 3

    Top transcripts with high stuORF scores.

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DOI

https://doi.org/10.1038/nature13401

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