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Protein targeting and degradation are coupled for elimination of mislocalized proteins

Nature volume 475, pages 394–397 (2011)Cite this article

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Abstract

A substantial proportion of the genome encodes membrane proteins that are delivered to the endoplasmic reticulum by dedicated targeting pathways1. Membrane proteins that fail targeting must be rapidly degraded to avoid aggregation and disruption of cytosolic protein homeostasis2,3. The mechanisms of mislocalized protein (MLP) degradation are unknown. Here we reconstitute MLP degradation in vitro to identify factors involved in this pathway. We find that nascent membrane proteins tethered to ribosomes are not substrates for ubiquitination unless they are released into the cytosol. Their inappropriate release results in capture by the Bag6 complex, a recently identified ribosome-associating chaperone4. Bag6-complex-mediated capture depends on the presence of unprocessed or non-inserted hydrophobic domains that distinguish MLPs from potential cytosolic proteins. A subset of these Bag6 complex ‘clients’ are transferred to TRC40 for insertion into the membrane, whereas the remainder are rapidly ubiquitinated. Depletion of the Bag6 complex selectively impairs the efficient ubiquitination of MLPs. Thus, by its presence on ribosomes that are synthesizing nascent membrane proteins, the Bag6 complex links targeting and ubiquitination pathways. We propose that such coupling allows the fast tracking of MLPs for degradation without futile engagement of the cytosolic folding machinery.

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Figure 1: Non-translocated PrP is rapidly ubiquitinated.
Figure 2: BAG6 interacts with MLPs through hydrophobic domains.
Figure 3: BAG6 captures MLPs released from the ribosome.
Figure 4: Maximum ubiquitination of MLPs requires BAG6.

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Acknowledgements

We are grateful to E. Whiteman and X. Li for carrying out the initial experiments for parts of this project, S.W. Kang, S. Shao, and Z. Zhang for discussions, P. Sengupta, J. Magadán, and C. Ott for constructs, J. Taunton and J. Garrison for cotransin, S. Shao for comments on the manuscript, and Y. Ye for discussions and sharing results before publication. This work was supported by the Intramural Research Program of the National Institutes of Health (R.S.H.) and a postdoctoral fellowship from The Wenner-Gren Foundations (T.H.).

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Author notes

  1. Heather D. Eshleman & Ramanujan S. Hegde

    Present address: Present addresses: Department of Physiology, University of California, San Francisco, California 94158, USA (H.D.E.); MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 0QH, UK (R.S.H.).,

Authors and Affiliations

  1. Cell Biology and Metabolism Program, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, 20892, Maryland, USA

    Tara Hessa, Ajay Sharma, Malaiyalam Mariappan, Heather D. Eshleman, Erik Gutierrez & Ramanujan S. Hegde

  2. Department of Biology, Johns Hopkins University, Baltimore, 21218, Maryland, USA

    Erik Gutierrez

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Contributions

T.H. performed most of the experiments, with contributions from A.S. (ubiquitination assays in modified lysates), M.M. (defining the substrate specificity of BAG6), H.D.E. (characterizing the Fr-RRL system), E.G. (BAG6 crosslinking analysis) and R.S.H. (in vivo studies). R.S.H. conceived the project, guided the experiments and wrote the paper with input from all of the authors.

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Correspondence to Ramanujan S. Hegde.

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The authors declare no competing financial interests.

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The file contains Supplementary Technical Notes relating to each of the main figures and Supplementary Figures 1-23 with legends. (PDF 9380 kb)

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Hessa, T., Sharma, A., Mariappan, M. et al. Protein targeting and degradation are coupled for elimination of mislocalized proteins. Nature 475, 394–397 (2011). https://doi.org/10.1038/nature10181

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