Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Mutant analysis links the translocon and BiP to retrograde protein transport for ER degradation

Nature volume 388, pages 891–895 (1997)Cite this article

Abstract

Proteins enter the secretory pathway through the endoplasmic reticulum1, which delivers properly folded proteins to their site of action2 and contains a quality-control system to monitor and prevent abnormal proteins from being delivered3. Many of these proteins are degraded by the cytoplasmic proteasome4,5,6,7,8, which requires their retrograde transport to the cytoplasm5,6. Based on a co-immunoprecipitation of major histocompatibility complex (MHC) class I heavy-chain breakdown intermediates with the translocon subunit Sec61p (refs 9, 10), it was speculated that Sec61p may be involved in retrograde transport11. Here we present functional evidence from genetic studies that Sec61p mediates retrograde transport of a mutated lumenal yeast carboxypeptidase ycsY (CPY*) in vivo. The endoplasmic reticulum lumenal chaperone BiP (Kar2p) and Sec63p, which are also subunits of the import machinery10,12, are involved in export of CPY* to the cytosol. Thus our results demonstrate that retrograde transport of proteins is mediated by a functional translocon. We consider the export of endoplasmic reticulum-localized proteins to the cytosol by the translocon for proteasome degradation to be a general process in eukaryotic cell biology.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: A functional Sec61p is required for ER degradation.
Figure 2: ER degradation of CPY* is affected by the sec63-1 (a) and kar2-113 (b) mutations.
Figure 3: Control experiments demonstrate that CPY* stabilization in the sec61-2 background is based on a defective export.
Figure 4: Mutant Sec61p is a substrate of ER degradation.

Similar content being viewed by others

References

  1. Pryer, N. K., Wuestehube, L. J. & Schekman, R. Vesicle-mediated protein sorting. Annu. Rev. Biochem. 61, 471–516 (1992).

    Google Scholar 

  2. Gething, M. J. & Sambrook, J. Protein folding in the cell. Nature 355, 33–45 (1992).

    Article  ADS  CAS  Google Scholar 

  3. Kopito, R. R. ER quality control: the cytoplasmic connection. Cell 88, 427–430 (1997).

    Google Scholar 

  4. Biederer, T., Volkwein, C. & Sommer, T. Degradation of subunits of the Sec61p complex, an integral component of the ER membrane, by the ubiquitin-proteasome pathway. EMBO J. 15, 2069–2076 (1996).

    Google Scholar 

  5. Hiller, M. M., Finger, A., Schweiger, M. & Wolf, D. H. ER degradation of a misfolded luminal protein by the cytosolic ubiquitin-proteasome pathway. Science 273, 1725–1728 (1996).

    Google Scholar 

  6. Werner, E. D., Brodsky, J. L. & McCracken, A. A. Proteasome-dependent endoplasmic reticulum-associated protein degradation: an unconventional route to a familiar fate. Proc. Natl Acad. Sci. USA 93, 13797–13801 (1996).

    Google Scholar 

  7. Jensen, T. J.et al. Multiple proteolytic systems, including the proteasome, contribute to CFTR processing. Cell 83, 129–135 (1995).

    Google Scholar 

  8. Halaban, R.et al. Aberrant retention of tyrosinase in the endoplasmic reticulum mediates accelerated degradation of the enzyme and contributes to the dedifferentiated phenotype of amelanotic melanoma cells. Proc. Natl Acad. Sci. USA 94, 6210–6215 (1997).

    Google Scholar 

  9. Walter, P. & Johnson, A. E. Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annu. Rev. Cell Biol. 10, 87–119 (1994).

    Google Scholar 

  10. Rapoport, T. A., Jungnickel, B. & Kutay, U. Protein transport across the eukaryotic endoplasmic reticulum and bacterial inner membranes. Annu. Rev. Biochem. 65, 271–303 (1996).

    Google Scholar 

  11. Wiertz, E. J. H. J.et al. Sec61-mediated transfer of a membrane protein from the endoplasmic reticulum to the proteasome for destruction. Nature 384, 432–438 (1996).

    Article  ADS  CAS  Google Scholar 

  12. Lyman, S. K. & Schekman, R. Binding of secretory precursor polypeptides to a translocon subcomplex is regulated by BiP. Cell 88, 85–96 (1997).

    Google Scholar 

  13. Finger, A., Knop, M. & Wolf, D. H. Analysis of two mutated vacuolar proteins reveals a degradation pathway in the endoplasmic reticulum or a related compartment of yeast. Eur. J. Biochem. 218, 565–574 (1993).

    Google Scholar 

  14. Stirling, C. S., Rothblatt, J., Hosobuchi, M., Deshaies, R. & Schekman, R. Protein translocation mutants defective in the insertion of integral membrane proteins into the endoplasmic reticulum. Mol. Cell. Biol. 3, 129–142 (1992).

    Google Scholar 

  15. Panzner, S., Dreier, L., Hartmann, E., Kostka, S. & Rapoport, T. A. Posttranslational protein transport in yeast reconstituted with a purified complex of Sec proteins and Kar2p. Cell 81, 561–570 (1995).

    Google Scholar 

  16. Finke, K.et al. Asecond trimeric complex containing homologs of the Sec61p complex functions in protein transport across the ER membrane of S. cerevisiae. EMBO J. 15, 1482–1494 (1996).

    Google Scholar 

  17. Feldheim, D., Rothblatt, J. & Schekman, R. Topology and functional domains of Sec63p, an endoplasmic reticulum membrane protein required for secretory protein translocation. Mol. Cell. Biol. 12, 3288–3296 (1992).

    Google Scholar 

  18. Corsi, A. K. & Schekman, R. The lumenal domain of Sec63p stimulates the ATPase activity of BiP and mediates BiP recruitment to the translocon in Saccharomyces cerevisiae. J. Cell Biol. 137, 1483–1493 (1997).

    Google Scholar 

  19. Cyr, D. M., Langer, T. & Douglas, M. G. DnaJ-like proteins: molecular chaperones and specific regulators of Hsp70. Trends Biochem. Sci. 19, 176–181 (1994).

    Google Scholar 

  20. Hanein, D.et al. Oligomeric rings of the Sec61p complex induced by ligands required for protein translocation. Cell 87, 721–732 (1996).

    Google Scholar 

  21. Scidmore, M. A., Okamura, H. H. & Rose, M. D. Genetic interactions between KAR2 and SEC63, encoding eukaryotic homologues of DnaK and DnaJ in the endoplasmic reticulum. Mol. Biol. Cell 4, 1145–1159 (1993).

    Google Scholar 

  22. Sanders, S. L., Whitfield, K. M., Vogel, J. P., Rose, M. D. & Schekman, R. Sec61p and BiP directly facilitate polypeptide translocation into the ER. Cell 69, 353–365 (1992).

    Google Scholar 

  23. Esnault, Y., Blondel, M. -O., Deshaies, R. J., Schekman, R. & Képès, F. The yeast SSS1 gene is essential for secretory protein translocation and encodes a conserved protein of the endoplasmic reticulum. EMBO J. 12, 4083–4093 (1993).

    Google Scholar 

  24. Ng, D. T., Brown, J. D. & Walter, P. Signal sequences specify the targeting route to the endoplasmic reticulum membrane. J. Cell Biol. 134, 269–278 (1996).

    Google Scholar 

  25. Chen, P., Johnson, P., Sommer, T., Jentsch, S. & Hochstrasser, M. Multiple ubiquitin-conjugating enzymes participate in the in vivo degradation of the yeast MAT α2 repressor. Cell 74, 357–369 (1993).

    Google Scholar 

  26. Knop, M., Finger, A., Braun, T., Hellmuth, K. & Wolf, D. H. Der1, a novel protein specifically requires for endoplasmic reticulum degradation in yeast. EMBO J. 15, 753–763 (1996).

    Google Scholar 

  27. Hampton, R. Y., Gardner, R. G. & Rine, J. Role of 26S proteasome and HRD genes in the degradation of 3-hydroxy-3-methylglutaryl-CoA reductase, an integral endoplasmic reticulum membrane protein. Mol. Biol. Cell 7, 2029–2044 (1994).

    Google Scholar 

  28. Ausubel, F. M.et al. Current Protocols in Molecular Biology(Greene, New York, (1992)).

    Google Scholar 

  29. Rose, M. D., Misra, L. M. & Vogel, J. P. Kar2, a karyogamy gene, is the yeast homolog of the mammalian BiP/GRP78 gene. Cell 57, 1211–1221 (1989).

    Google Scholar 

Download references

Acknowledgements

We thank M. Hochstrasser (Deg1-β-galactosidase fusion), M. D. Rose (kar2-113 allele), T. A. Rapoport (Sec61p antibodies), H. K. Rudolph and R. Schekman (Kar2 antibodies), M. Knop (polyclonal CPY antibodies) and S. Rupp (proteinase yscA antibodies) for providing gene constructs and affinity purified antibodies; and S. Jäger and M. Hämmerle for discussions. This work was supported by the Bundesministerium für Forschung und Technologie and the Fonds der Chemischen Industrie, Frankfurt.

Author information

Authors and Affiliations

  1. Institut für Biochemie der Universität Stuttgart, Pfaffenwaldring 55, D-70569, Stuttgart, Germany

    Richard K. Plemper, Sigrun Böhmler, Javier Bordallo & Dieter H. Wolf

  2. Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rössle Strasse 10, D-13122, Berlin, Germany

    Thomas Sommer

Authors
  1. Richard K. Plemper
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sigrun Böhmler
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Javier Bordallo
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Thomas Sommer
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Dieter H. Wolf
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Plemper, R., Böhmler, S., Bordallo, J. et al. Mutant analysis links the translocon and BiP to retrograde protein transport for ER degradation. Nature 388, 891–895 (1997). https://doi.org/10.1038/42276

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/42276

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing