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:

A cryptic protease couples deubiquitination and degradation by the proteasome

Nature volume 419, pages 403–407 (2002)Cite this article

Abstract

The 26S proteasome is responsible for most intracellular proteolysis in eukaryotes1,2. Efficient substrate recognition relies on conjugation of substrates with multiple ubiquitin molecules and recognition of the polyubiquitin moiety by the 19S regulatory complex—a multisubunit assembly that is bound to either end of the cylindrical 20S proteasome core. Only unfolded proteins can pass through narrow axial channels into the central proteolytic chamber of the 20S core, so the attached polyubiquitin chain must be released to allow full translocation of the substrate polypeptide. Whereas unfolding is rate-limiting for the degradation of some substrates and appears to involve chaperone-like activities associated with the proteasome3,4,5, the importance and mechanism of degradation-associated deubiquitination has remained unclear. Here we report that the POH1 (also known as Rpn11 in yeast) subunit of the 19S complex is responsible for substrate deubiquitination during proteasomal degradation. The inability to remove ubiquitin can be rate-limiting for degradation in vitro and is lethal to yeast. Unlike all other known deubiquitinating enzymes (DUBs) that are cysteine proteases6,7, POH1 appears to be a Zn2+-dependent protease.

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: Proteasomal degradation of UbOM is targeted by means of the unfolded ovomucoid moiety.
Figure 2: A ubiquitin-aldehyde (Ubal)-insensitive DUB couples release of ubiquitin with substrate degradation (western blots).
Figure 3: Alignment of the MPN domain of MPN family proteins.
Figure 4: Evidence that Rpn11/POH1 is a Zn2+-dependent DUB and characterization of rpn11 mutants.

Similar content being viewed by others

References

  1. Hershko, A. & Ciechanover, A. The ubiquitin system. Annu. Rev. Biochem. 67, 425–479 (1998)

    CAS  Google Scholar 

  2. Zwickl, P., Baumeister, W. & Steven, A. Dis-assembly lines: the proteasome and related ATPase-assisted proteases. Curr. Opin. Struct. Biol. 10, 242–250 (2000)

    Article  CAS  Google Scholar 

  3. Thrower, J. S., Hoffman, L., Rechsteiner, M. & Pickart, C. M. Recognition of the polyubiquitin proteolytic signal. EMBO J. 19, 94–102 (2000)

    Article  CAS  Google Scholar 

  4. Braun, B. C. et al. The base of the proteasome regulatory particle exhibits chaperone-like activity. Nature Cell Biol. 1, 221–226 (1999)

    Article  ADS  CAS  Google Scholar 

  5. Liu, C. et al. Conformational remodeling of proteasomal substrates by PA700, the 19S regulatory complex of the 26S proteasome. J. Biol. Chem. 277, 26815–26820 (2002)

    Article  CAS  Google Scholar 

  6. D'Andrea, A. & Pellman, D. Deubiquitinating enzymes: a new class of biological regulators. Crit. Rev. Biochem. Mol. Biol. 33, 337–352 (1998)

    Article  CAS  Google Scholar 

  7. Amerik, A. Y., Li, S. J. & Hochstrasser, M. Analysis of the deubiquitinating enzymes of the yeast Saccharomyces cerevisiae. Biol. Chem. 381, 981–992 (2000)

    Article  CAS  Google Scholar 

  8. DeKoster, G. T. & Robertson, A. D. Thermodynamics of unfolding for Kazal-type serine protease inhibitors: entropic stabilization of ovomucoid first domain by glycosylation. Biochemistry 36, 2323–2331 (1997)

    Article  CAS  Google Scholar 

  9. Johnson, E. S., Ma, P. C., Ota, I. M. & Varshavsky, A. A proteolytic pathway that recognizes ubiquitin as a degradation signal. J. Biol. Chem. 270, 17442–17456 (1995)

    Article  CAS  Google Scholar 

  10. Hershko, A. & Rose, I. A. Ubiquitin-aldehyde: a general inhibitor of ubiquitin-recycling processes. Proc. Natl Acad. Sci. USA 84, 1829–1833 (1987)

    Article  ADS  CAS  Google Scholar 

  11. Lam, Y. A., Xu, W., DeMartino, G. N. & Cohen, R. E. Editing of ubiquitin conjugates by an isopeptidase in the 26S proteasome. Nature 385, 737–740 (1997)

    Article  ADS  CAS  Google Scholar 

  12. Wintrode, P. L., Makhatadze, G. I. & Privalov, P. L. Thermodynamics of ubiquitin unfolding. Proteins 18, 246–253 (1994)

    Article  CAS  Google Scholar 

  13. Borodovsky, A. et al. A novel active site-directed probe specific for deubiquitylating enzymes reveals proteasome association of USP14. EMBO J. 20, 5187–5196 (2001)

    Article  CAS  Google Scholar 

  14. Lam, Y. A., Lawson, T. G., Velayutham, M., Zweier, J. L. & Pickart, C. M. A proteasomal ATPase subunit recognizes the polyubiquitin degradation signal. Nature 416, 763–767 (2002)

    Article  ADS  CAS  Google Scholar 

  15. Rubin, D. M., Glickman, M. H., Larsen, C. N., Dhruvakumar, S. & Finley, D. Active site mutants in the six regulatory particle ATPases reveal multiple roles for ATP in the proteasome. EMBO J. 17, 4909–4919 (1998)

    Article  CAS  Google Scholar 

  16. Leggett, D. S. et al. Multiple associated proteins regulate proteasome structure and function. Mol. Cell (in the press)

  17. Papa, F. R. & Hochstrasser, M. The yeast DOA4 gene encodes a deubiquitinating enzyme related to a product of the human tre-2 oncogene. Nature 366, 313–319 (1993)

    Article  ADS  CAS  Google Scholar 

  18. Papa, F. R., Amerik, A. Y. & Hochstrasser, M. Interaction of the Doa4 deubiquitinating enzyme with the yeast 26S proteasome. Mol. Biol. Cell 10, 741–756 (1999)

    Article  CAS  Google Scholar 

  19. Glickman, M. H. et al. A subcomplex of the proteasome regulatory particle required for ubiquitin-conjugate degradation and related to the COP9-signalosome and eIF3. Cell 94, 615–623 (1998)

    Article  CAS  Google Scholar 

  20. Lyapina, S. et al. Promotion of NEDD-CUL1 conjugate cleavage by COP9 signalosome. Science 292, 1382–1385 (2001)

    Article  ADS  CAS  Google Scholar 

  21. Hofmann, K. & Bucher, P. The PCI domain: a common theme in three multiprotein complexes. Trends Biochem. Sci. 23, 204–205 (1998)

    Article  CAS  Google Scholar 

  22. Glickman, M. H., Rubin, D. M., Fried, V. A. & Finley, D. The regulatory particle of the Saccharomyces cerevisiae proteasome. Mol. Cell Biol. 18, 3149–3162 (1998)

    Article  CAS  Google Scholar 

  23. Alberts, I. L., Nadassy, K. & Wodak, S. J. Analysis of zinc binding sites in protein crystal structures. Protein Sci. 7, 1700–1716 (1998)

    Article  CAS  Google Scholar 

  24. Eytan, E., Armon, T., Heller, H., Beck, S. & Hershko, A. Ubiquitin C-terminal hydrolase activity associated with the 26 S protease complex. J. Biol. Chem. 268, 4668–4674 (1993)

    CAS  PubMed  Google Scholar 

  25. Wilkinson, K. D. Regulation of ubiquitin-dependent processes by deubiquitinating enzymes. FASEB J. 11, 1245–1256 (1997)

    Article  CAS  Google Scholar 

  26. Ponting, C. P., Aravind, L., Schultz, J., Bork, P. & Koonin, E. V. Eukaryotic signalling domain homologues in archaea and bacteria. Ancient ancestry and horizontal gene transfer. J. Mol. Biol. 289, 729–745 (1999)

    Article  CAS  Google Scholar 

  27. Thick, J., Mak, Y. F., Metcalfe, J., Beatty, D. & Taylor, A. M. A gene on chromosome Xq28 associated with T-cell prolymphocytic leukemia in two patients with ataxia telangiectasia. Leukemia 8, 564–573 (1994)

    CAS  PubMed  Google Scholar 

  28. Beers, E. P. & Callis, J. Utility of polyhistidine-tagged ubiquitin in the purification of ubiquitin-protein conjugates and as an affinity ligand for the purification of ubiquitin-specific hydrolases. J. Biol. Chem. 268, 21645–21649 (1993)

    CAS  PubMed  Google Scholar 

  29. Hoffman, L., Pratt, G. & Rechsteiner, M. Multiple forms of the 20S multicatalytic and the 26S ubiquitin/ATP- dependent proteases from rabbit reticulocyte lysate. J. Biol. Chem. 267, 22362–22368 (1992)

    CAS  PubMed  Google Scholar 

  30. Piotrowski, J. et al. Inhibition of the 26S proteasome by polyubiquitin chains synthesized to have defined lengths. J. Biol. Chem. 272, 23712–23721 (1997)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank D. Finley and D. Leggett for yeast strains, advice on proteasome purification, and a sample of yeast proteasomes; G. DeMartino for bovine PA700; A. Robertson for ovomucoid protein; C. Pickart for the Ub(V76)DHFR plasmid; K. Wilkinson, H. Ploegh and A. Borodovsky for UbVS, recombinant Ubp6 protein and anti-human Ubp6; C. Pickart, A. Lam and K. Wilkinson for discussions; and L. Weisman and C. Pickart for comments on the manuscript. This work was supported by a grant from the NIH and a UI Biocatalysis and Bioprocessing Fellowship to T.Y.

Author information

Authors and Affiliations

  1. Department of Biochemistry, University of Iowa, 51 Newton Road, Iowa City, Iowa, 52242, USA

    Tingting Yao & Robert E. Cohen

Authors
  1. Tingting Yao
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Robert E. Cohen
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Robert E. Cohen.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yao, T., Cohen, R. A cryptic protease couples deubiquitination and degradation by the proteasome. Nature 419, 403–407 (2002). https://doi.org/10.1038/nature01071

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature01071

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