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Proteasome recruitment and activation of the Uch37 deubiquitinating enzyme by Adrm1

Nature Cell Biology volume 8, pages 994–1002 (2006)Cite this article

Abstract

Uch37 is one of the three principal deubiquitinating enzymes (DUBs), and the only ubiquitin carboxy-terminal hydrolase (UCH)-family protease, that is associated with mammalian proteasomes. We show that Uch37 is responsible for the ubiquitin isopeptidase activity in the PA700 (19S) proteasome regulatory complex1. PA700 isopeptidase disassembles Lys 48-linked polyubiquitin specifically from the distal end of the chain, a property that may be used to clear poorly ubiquitinated or unproductively bound substrates from the proteasome. To better understand Uch37 function and the mechanism responsible for its specificity, we investigated how Uch37 is recruited to proteasomes. Uch37 binds through Adrm1, a previously unrecognized orthologue of Saccharomyces cerevisiae Rpn13p, which in turn is bound to the S1 (also known as Rpn2) subunit of the 19S complex. Adrm1 (human Rpn13, hRpn13) binds the carboxy-terminal tail of Uch37, a region that is distinct from the UCH catalytic domain, which we show inhibits Uch37 activity. Following binding, Adrm1 relieves Uch37 autoinhibition, accelerating the hydrolysis of ubiquitin-7-amido-4-methylcoumarin (ubiquitin−AMC). However, neither Uch37 alone nor the Uch37–Adrm1 or Uch37–Adrm1–S1 complexes can hydrolyse di-ubiquitin efficiently; rather, incorporation into the 19S complex is required to enable processing of polyubiquitin chains.

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Figure 1: Human Uch37 is associated with the 26S proteasome in vivo and disassembles di-ubiquitin.
Figure 2: Adrm1 interacts with Uch37 in vivo and in vitro.
Figure 3: Adrm1 is a human homologue of yeast Rpn13p and a subunit of the proteasome.
Figure 4: Adrm1 recruits Uch37 to the proteasome.
Figure 5: Deubiquitination by Uch37 is activated by Adrm1 and in the 19S regulatory complex.

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References

  1. 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  CAS  Google Scholar 

  2. Lam, Y. A., DeMartino, G. N., Pickart, C. M. & Cohen, R. E. Specificity of the ubiquitin isopeptidase in the PA700 regulatory complex of 26 S proteasomes. J. Biol. Chem. 272, 28438–28446 (1997).

    Article  CAS  Google Scholar 

  3. Li, T., Naqvi, N. I., Yang, H. & Teo, T. S. Identification of a 26S proteasome-associated UCH in fission yeast. Biochem. Biophys. Res. Commun. 272, 270–275 (2000).

    Article  CAS  Google Scholar 

  4. Holzl, H. et al. The regulatory complex of Drosophila melanogaster 26S proteasomes. Subunit composition and localization of a deubiquitylating enzyme. J. Cell Biol. 150, 119–130 (2000).

    Article  CAS  Google Scholar 

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

    CAS  PubMed  Google Scholar 

  6. Wolters, D. A., Washburn, M. P. & Yates, J. R., 3rd. An automated multidimensional protein identification technology for shotgun proteomics. Anal. Chem. 73, 5683–5690 (2001).

    Article  CAS  Google Scholar 

  7. 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 

  8. Johnston, S. C., Riddle, S. M., Cohen, R. E. & Hill, C. P. Structural basis for the specificity of ubiquitin C-terminal hydrolases. EMBO J. 18, 3877–3887 (1999).

    Article  CAS  Google Scholar 

  9. Verma, R. et al. Proteasomal proteomics: identification of nucleotide-sensitive proteasome-interacting proteins by mass spectrometric analysis of affinity-purified proteasomes. Mol. Biol. Cell 11, 3425–3439 (2000).

    Article  CAS  Google Scholar 

  10. Leggett, D. S. et al. Multiple associated proteins regulate proteasome structure and function. Mol. Cell 10, 495–507 (2002).

    Article  CAS  Google Scholar 

  11. Yao, T. & Cohen, R. E. A cryptic protease couples deubiquitination and degradation by the proteasome. Nature 419, 403–407 (2002).

    Article  CAS  Google Scholar 

  12. Li, T. et al. Identification of two proteins, S14 and UIP1, that interact with UCH37. FEBS Lett. 488, 201–205 (2001).

    Article  CAS  Google Scholar 

  13. Shimada, S., Ogawa, M., Takahashi, M., Schlom, J. & Greiner, J. W. Molecular cloning and characterization of the complementary DNA of an M(r) 110,000 antigen expressed by human gastric carcinoma cells and upregulated by gamma-interferon. Cancer Res. 54, 3831–3836 (1994).

    CAS  PubMed  Google Scholar 

  14. Ito, T. et al. A comprehensive two-hybrid analysis to explore the yeast protein interactome. Proc. Natl Acad. Sci. USA 98, 4569–4574 (2001).

    Article  CAS  Google Scholar 

  15. Giot, L. et al. A protein interaction map of Drosophila melanogaster. Science 302, 1727–1736 (2003).

    Article  CAS  Google Scholar 

  16. Larsen, C. N., Krantz, B. A. & Wilkinson, K. D. Substrate specificity of deubiquitinating enzymes: ubiquitin C-terminal hydrolases. Biochemistry 37, 3358–3368 (1998).

    Article  CAS  Google Scholar 

  17. Johnston, S. C., Larsen, C. N., Cook, W. J., Wilkinson, K. D. & Hill, C. P. Crystal structure of a deubiquitinating enzyme (human UCH-L3) at 1.8 A resolution. EMBO J. 16, 3787–3796 (1997).

    Article  CAS  Google Scholar 

  18. Lamerant, N. & Kieda, C. Adhesion properties of adhesion-regulating molecule 1 protein on endothelial cells. FEBS J. 272, 1833–1844 (2005).

    Article  CAS  Google Scholar 

  19. Sumegi, M., Hunyadi-Gulyas, E., Medzihradszky, K. F. & Udvardy, A. 26S proteasome subunits are O-linked N-acetylglucosamine-modified in Drosophila melanogaster. Biochem. Biophys. Res. Commun. 312, 1284–1289 (2003).

    Article  CAS  Google Scholar 

  20. Simins, A. B., Weighardt, H., Weidner, K. M., Weidle, U. H. & Holzmann, B. Functional cloning of ARM-1, an adhesion-regulating molecule upregulated in metastatic tumor cells. Clin. Exp. Metastasis 17, 641–648 (1999).

    Article  CAS  Google Scholar 

  21. Pilarsky, C., Wenzig, M., Specht, T., Saeger, H. D. & Grutzmann, R. Identification and validation of commonly overexpressed genes in solid tumors by comparison of microarray data. Neoplasia 6, 744–750 (2004).

    Article  CAS  Google Scholar 

  22. Shimada, S., Ogawa, M., Schlom, J. & Greiner, J. W. Identification of a novel tumor-associated Mr 110,000 gene product in human gastric carcinoma cells that is immunologically related to carcinoembryonic antigen. Cancer Res. 51, 5694–5703 (1991).

    CAS  PubMed  Google Scholar 

  23. Jensen, D. E. et al. BAP1: a novel ubiquitin hydrolase which binds to the BRCA1 RING finger and enhances BRCA1-mediated cell growth suppression. Oncogene 16, 1097–1112 (1998).

    Article  CAS  Google Scholar 

  24. Lee, K. K., Florens, L., Swanson, S. K., Washburn, M. P. & Workman, J. L. The deubiquitylation activity of ubiquitinp8 is dependent upon Sgf11 and its association with the SAGA complex. Mol. Cell. Biol. 25, 1173–1182 (2005).

    Article  CAS  Google Scholar 

  25. Das, C. et al. Structural basis for conformational plasticity of the Parkinson's disease-associated ubiquitin hydrolase UCH-L1. Proc. Natl Acad. Sci. USA 103, 4675–4680 (2006).

    Article  CAS  Google Scholar 

  26. Misaghi, S. et al. Structure of the ubiquitin hydrolase UCH-L3 complexed with a suicide substrate. J. Biol. Chem. 280, 1512–1520 (2005).

    Article  CAS  Google Scholar 

  27. Pace, C. N., Vajdos, F., Fee, L., Grimsley, G. & Gray, T. How to measure and predict the molar absorption coefficient of a protein. Protein Sci. 4, 2411–2423 (1995).

    Article  CAS  Google Scholar 

  28. Dignam, J. D., Lebovitz, R. M. & Roeder, R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 11, 1475–1489 (1983).

    Article  CAS  Google Scholar 

  29. James, P., Halladay, J. & Craig, E. A. Genomic libraries and a host strain designed for highly efficient two-hybrid selection in yeast. Genetics 144, 1425–1436 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Altschul, S. F. et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 (1997).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank C. Gorbea and M. Rechsteiner for plasmids containing 19S complex cDNAs, A. Borodovsky and H. Ploegh for UbVS, B. Plapp for advice on fitting kinetic data and C. Slaughter for mass spectrometry of Uch37 peptides. This work was supported, in part, by the National Institutes of Health grants R01 GM37666 (to R.E.C.) and R37 GM41628 (to R.C.C.). T.Y. is a fellow of the Leukemia and Lymphoma Society.

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Author notes

  1. Wei Xu

    Present address: McArdle Laboratory for Cancer Research, Madison, WI 53706, USA

  2. Robert E. Cohen

    Present address: Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, USA

Authors and Affiliations

  1. Stowers Institute for Medical Research, Kansas City, 64110, MO, USA

    Tingting Yao, Laurence Florens, Selene K. Swanson, Michael P. Washburn, Ronald C. Conaway & Joan Weliky Conaway

  2. Department of Biochemistry, Carver College of Medicine, University of Iowa, Iowa City, 52242, IA, USA

    Tingting Yao, Ling Song, Wei Xu & Robert E. Cohen

  3. Department of Physiology, University of Texas Southwestern Medical Center, Dallas, 75390, TX, USA

    George N. DeMartino

  4. Department of Biochemistry and Molecular Biology, Kansas University Medical Center, Kansas City, 66160, KS, USA

    Ronald C. Conaway & Joan Weliky Conaway

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Yao, T., Song, L., Xu, W. et al. Proteasome recruitment and activation of the Uch37 deubiquitinating enzyme by Adrm1. Nat Cell Biol 8, 994–1002 (2006). https://doi.org/10.1038/ncb1460

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