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:

The conserved protein DCN-1/Dcn1p is required for cullin neddylation in C. elegans and S. cerevisiae

Nature volume 435, pages 1257–1261 (2005)Cite this article

Abstract

SCF-type E3 ubiquitin ligases are multi-protein complexes required for polyubiquitination and subsequent degradation of target proteins by the 26S proteasome1. Cullins, together with the RING-finger protein Rbx1, form the catalytic core of the ligase, and recruit the substrate-recognition module1,2,3,4. Cycles of covalent modification of cullins by the ubiquitin-like molecule Nedd8 (neddylation)5 and removal of Nedd8 by the COP9 signalosome (deneddylation) positively regulate E3 ligase activity6,7. Here we report the identification and analysis of a widely conserved protein that is required for cullin neddylation in the nematode Caenorhabditis elegans and the yeast Saccharomyces cerevisiae. C. elegans DCN-1 and S. cerevisiae Dcn1p (defective in cullin neddylation) are characterized by a novel UBA-like ubiquitin-binding domain and a DUF298 domain of unknown function. Consistent with their requirements for neddylation, DCN-1 and Dcn1p directly bind Nedd8 and physically associate with cullins in both species. Moreover, overexpression of Dcn1p in yeast results in the accumulation of Nedd8-modified cullin Cdc53p. Both in vivo and in vitro experiments indicate that Dcn1p does not inhibit deneddylation of Cdc53p by the COP9 signalosome, but greatly increases the kinetics of the neddylation reaction.

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: dcn-1(RNAi) embryos stabilize MEI-1 protein.
Figure 2: DCN-1 and Dcn1p are evolutionarily and functionally conserved and bind ubiquitin and Nedd8.
Figure 3: DCN-1 and Dcn1p proteins are required for Nedd8 modification of cullins in yeast and nematodes, and physically associate with cullins.
Figure 4: DCN-1 and Dcn1p catalyse cullin neddylation in vivo and in vitro.

Similar content being viewed by others

References

  1. Deshaies, R. J. SCF and Cullin/Ring H2-based ubiquitin ligases. Annu. Rev. Cell Dev. Biol. 15, 435–467 (1999)

    Article  CAS  PubMed  Google Scholar 

  2. Kile, B. T. et al. The SOCS box: a tale of destruction and degradation. Trends Biochem. Sci. 27, 235–241 (2002)

    Article  CAS  PubMed  Google Scholar 

  3. Pintard, L. et al. The BTB protein MEL-26 is a substrate-specific adaptor of the CUL-3 ubiquitin-ligase. Nature 425, 311–316 (2003)

    Article  ADS  CAS  PubMed  Google Scholar 

  4. Furukawa, M., Ohta, T. & Xiong, Y. Activation of UBC5 ubiquitin-conjugating enzyme by the RING finger of ROC1 and assembly of active ubiquitin ligases by all cullins. J. Biol. Chem. 277, 15758–15765 (2002)

    Article  CAS  PubMed  Google Scholar 

  5. Pan, Z. Q., Kentsis, A., Dias, D. C., Yamoah, K. & Wu, K. Nedd8 on cullin: building an expressway to protein destruction. Oncogene 23, 1985–1997 (2004)

    Article  CAS  PubMed  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  7. Pintard, L. et al. Neddylation and deneddylation of CUL-3 is required to target MEI-1/katanin for degradation at the meiosis-to-mitosis transition in C. elegans. Curr. Biol. 13, 911–921 (2003)

    Article  CAS  PubMed  Google Scholar 

  8. Kurz, T. et al. Cytoskeletal regulation by the Nedd8 ubiquitin-like protein modification pathway. Science 295, 1294–1298 (2002)

    Article  ADS  CAS  PubMed  Google Scholar 

  9. McNally, F. J. & Vale, R. D. Identification of katanin, an ATPase that severs and disassembles stable microtubules. Cell 75, 419–429 (1993)

    Article  CAS  PubMed  Google Scholar 

  10. Srayko, M., Buster, D. W., Bazirgan, O. A., McNally, F. J. & Mains, P. E. MEI-1/MEI-2 katanin-like microtubule severing activity is required for Caenorhabditis elegans meiosis. Genes Dev. 14, 1072–1084 (2000)

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Dow, M. R. & Mains, P. E. Genetic and molecular characterization of the Caenorhabditis elegans gene, mel-26, a postmeiotic negative regulator of mei-1, a meiotic-specific spindle component. Genetics 150, 119–128 (1998)

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Gonczy, P. et al. Functional genomic analysis of cell division in C. elegans using RNAi of genes on chromosome III. Nature 408, 331–336 (2000)

    Article  ADS  CAS  PubMed  Google Scholar 

  13. Fraser, A. G. et al. Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. Nature 408, 325–330 (2000)

    Article  ADS  CAS  PubMed  Google Scholar 

  14. Bucher, P., Karplus, K., Moeri, N. & Hofmann, K. A flexible motif search technique based on generalized profiles. Comput. Chem. 20, 3–23 (1996)

    Article  CAS  PubMed  Google Scholar 

  15. Meyer, H. H., Wang, Y. & Warren, G. Direct binding of ubiquitin conjugates by the mammalian p97 adaptor complexes, p47 and Ufd1-Npl4. EMBO J. 21, 5645–5652 (2002)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Liakopoulos, D., Doenges, G., Matuschewski, K. & Jentsch, S. A novel protein modification pathway related to the ubiquitin system. EMBO J. 17, 2208–2214 (1998)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Lammer, D. et al. Modification of yeast Cdc53p by the ubiquitin-related protein rub1p affects function of the SCFCdc4 complex. Genes Dev. 12, 914–926 (1998)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Uetz, P. et al. A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisiae. Nature 403, 623–627 (2000)

    Article  ADS  CAS  PubMed  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  20. Cope, G. A. et al. Role of predicted metalloprotease motif of Jab1/Csn5 in cleavage of Nedd8 from Cul1. Science 298, 608–611 (2002)

    Article  ADS  CAS  PubMed  Google Scholar 

  21. Morimoto, M., Nishida, T., Nagayama, Y. & Yasuda, H. Nedd8-modification of Cul1 is promoted by Roc1 as a Nedd8–E3 ligase and regulates its stability. Biochem. Biophys. Res. Commun. 301, 392–398 (2003)

    Article  CAS  PubMed  Google Scholar 

  22. Zheng, J. et al. CAND1 binds to unneddylated CUL1 and regulates the formation of SCF ubiquitin E3 ligase complex. Mol. Cell 10, 1519–1526 (2002)

    Article  CAS  PubMed  Google Scholar 

  23. Liu, J., Furukawa, M., Matsumoto, T. & Xiong, Y. NEDD8 modification of CUL1 dissociates p120(CAND1), an inhibitor of CUL1–SKP1 binding and SCF ligases. Mol. Cell 10, 1511–1518 (2002)

    Article  CAS  PubMed  Google Scholar 

  24. Kawakami, T. et al. NEDD8 recruits E2-ubiquitin to SCF E3 ligase. EMBO J. 20, 4003–4012 (2001)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Dharmasiri, S., Dharmasiri, N., Hellmann, H. & Estelle, M. The RUB/Nedd8 conjugation pathway is required for early development in Arabidopsis. EMBO J. 22, 1762–1770 (2003)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Guthrie, C. & Fink, G. R. in Guide to Yeast Genetics and Molecular Biology (eds Abelson, J. N. & Simon, M. I.) (Academic, San Diego, California, 1991)

    Google Scholar 

  27. Hofmann, K. Sensitive protein comparisons with profiles and hidden Markov models. Brief. Bioinform. 1, 167–178 (2000)

    Article  CAS  PubMed  Google Scholar 

  28. Jaquenoud, M., Gulli, M. P., Peter, K. & Peter, M. The Cdc42p effector Gic2p is targeted for ubiquitin-dependent degradation by the SCFGrr1 complex. EMBO J. 17, 5360–5373 (1998)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Verma, R., Chi, Y. & Deshaies, R. J. Cell-free ubiquitination of cell cycle regulators in budding yeast extracts. Methods Enzymol. 283, 366–376 (1997)

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank M. Srayko for providing DCN-1 cDNA; M. Tyers and X. Tang for purified Cdc53p–Rbx1p complex; S. Rybina for help with antibody purification; and J. C. Labbé and L. Pintard for critical reading of the manuscript. T.K. was supported by an American Heart Association Pre-doctoral Fellowship and an EMBO Long-Term Fellowship, S.M.O. by the Damon Runyon Cancer Research Foundation, B.B. by the NIH, and M.P. by the ETHZ and the Swiss National Science Foundation.

Author information

Authors and Affiliations

  1. Institute of Biochemistry, ETH Hönggerberg, 8093, Zürich, Switzerland

    Thimo Kurz, Fabian Rudolf, Brian Luke & Matthias Peter

  2. Institute of Molecular Biology, University of Oregon, Eugene, Oregon, 97403, USA

    Thimo Kurz, Sean M. O'Rourke & Bruce Bowerman

  3. Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, 01307, Germany

    Nurhan Özlü & Anthony A. Hyman

  4. Memorec Biotec GmbH, 50829, Köln, Germany

    Kay Hofmann

Authors
  1. Thimo Kurz
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nurhan Özlü
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Fabian Rudolf
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sean M. O'Rourke
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Brian Luke
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Kay Hofmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Anthony A. Hyman
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Bruce Bowerman
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Matthias Peter
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Bruce Bowerman or Matthias Peter.

Ethics declarations

Competing interests

Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Video S1

This time-lapse movie shows the first mitotic division of a Caernorhabditis elegans embryo expressing a GFP::MEI-1 fusion protein. It is evident from this movie that under wildtype conditions MEI-1 protein does not localize to the mitotic spindle. (MOV 9187 kb)

Supplementary Video 2

This time-lapse movie shows the first mitotic division of a C. elegans embryo expressing a GFP::MEI-1 fusion protein, in which dcn-1 has been inactivated by RNAi. It is evident from this movie that loss of DCN-1 results in the stabilization of MEI-1 protein and its ectopic association with the mitotic spindle. (MOV 2982 kb)

Supplementary Video Legends

Legends to accompany the above Supplementary Videos (DOC 20 kb)

Supplementary Figures

Supplementary Figure S1, multiple alignment of DUF298 domains; Supplementary Figure S2, multiple alignment of UBA-like domains; Supplementary Figure S3, drawing of the two C. elegans DCN-1 spliceforms; Supplementary Figure S4, subcellular localization of CeDCN-1; Supplementary Figure S5, neddylation state of cullin Rtt101p. (DOC 878 kb)

Supplementary Table S1

Table of yeast strains used in this study. (DOC 32 kb)

Supplementary Methods

This file contains additional Methods that could not be displayed in the print version. (DOC 23 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kurz, T., Özlü, N., Rudolf, F. et al. The conserved protein DCN-1/Dcn1p is required for cullin neddylation in C. elegans and S. cerevisiae. Nature 435, 1257–1261 (2005). https://doi.org/10.1038/nature03662

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature03662

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