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.

  • Article
  • Published:

SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27

Nature Cell Biology volume 1, pages 193–199 (1999)Cite this article

Abstract

Degradation of the mammalian cyclin-dependent kinase (CDK) inhibitor p27 is required for the cellular transition from quiescence to the proliferative state. The ubiquitination and subsequent degradation of p27 depend on its phosphorylation by cyclin–CDK complexes. However, the ubiquitin–protein ligase necessary for p27 ubiquitination has not been identified. Here we show that the F-box protein SKP2 specifically recognizes p27 in a phosphorylation-dependent manner that is characteristic of an F-box-protein–substrate interaction. Furthermore, both in vivo and in vitro, SKP2 is a rate-limiting component of the machinery that ubiquitinates and degrades phosphorylated p27. Thus, p27 degradation is subject to dual control by the accumulation of both SKP2 and cyclins following mitogenic stimulation.

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

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

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

Figure 1: Binding of phosphorylated p27 to SKP2.
Figure 2: In vivo binding of SKP2 to p27.
Figure 3: SKP2 and cyclin E–CDK2 are rate limiting for p27 ubiquitination in G1 extracts.
Figure 4: SKP2 is required for p27–ubiquitin ligation activity.
Figure 5: In vivo role of SKP2 in p27 degradation.
Figure 6: Stabilization of cellular p27 by antisense oligonucleotides targeting SKP2 mRNA.
Figure 7: Timing of SKP2 action in the process of p27 degradation.

Similar content being viewed by others

References

  1. Sherr, C. & Roberts, J. Cdk inhibitors: positive and negative regulators of G1-phase progression. Genes Dev. 13, 1501–1514 (1999).

    Article  CAS  Google Scholar 

  2. Elledge, S. J., Winston, J. & Harper, J. W. A question of balance: the roles of cyclin kinase inhibitors in development and tumorigenesis. Trends Cell Biol. 6, 388–392 ( 1996).

    Article  CAS  Google Scholar 

  3. Sheaff, R. J. & Roberts, J. M. in Cell Cycle Control, Results Probl. Cell Differ. Vol. 22 (ed. Pagano, M.) 1– 34 (Springer, New York, 1998).

    Book  Google Scholar 

  4. Hershko, A. Roles of ubiquitin-mediated proteolysis in cell cycle control. Curr. Opin. Cell Biol. 9, 788–799 (1997).

    Article  CAS  Google Scholar 

  5. Pagano, M. Regulation of cell cycle regulatory proteins by the ubiquitin pathway. FASEB J. 11, 1067–1075 ( 1997).

    Article  CAS  Google Scholar 

  6. Patton, E., Willems, A. & Tyers, M. Combinatorial control in ubiquitin-dependent proteolysis: don"t Skp the F-box hypothesis. Trends Genet. 14, 6–14 (1998).

    Article  Google Scholar 

  7. Loda, M. et al. Increased proteasome-dependent degradation of the cyclin-dependent kinase inhibitor p27 in aggressive colorectal carcinomas. Nature Med. 3, 231–234 ( 1997).

    Article  CAS  Google Scholar 

  8. Esposito, V. et al. Prognostic role of the cell cycle inhibitor p27 in non small cell lung cancer. Cancer Res. 57, 3381– 3385 (1997).

    CAS  PubMed  Google Scholar 

  9. Mayor, S. Protein marker linked with poor cancer outcome. Br. Med. J. 314, 323 (1997).

    Article  Google Scholar 

  10. Steeg, P. & Abrams, J. Cancer prognostic: past, present and p27. Nature Med. 3, 152– 154 (1997).

    Article  CAS  Google Scholar 

  11. Ciechanover, A. The ubiquitin-proteasome pathway: on protein death and cell life. EMBO J. 17, 7151–7160 ( 1998).

    Article  CAS  Google Scholar 

  12. Budel, L. et al. Characterization of p21, p27, p53 and E2F-1 cell cycle regulators in mantle cell lymphoma: increased ubiquitin-proteasome mediated degradation of p27. Blood (in the press).

  13. Piva, R. et al. Increased proteasome-dependent degradation of p27 in malignant gliomas. J. Neuropathol. Exp. Neurol. (in the press).

  14. Pagano, M. et al. Role of the ubiquitin-proteasome pathway in regulating abundance of the cyclin-dependent kinase inhibitor p27. Science 269, 682–685 (1995).

    Article  CAS  Google Scholar 

  15. Montagnoli, A. et al. Ubiquitination of p27 is regulated by Cdk-dependent phosphorylation and trimeric complex formation. Genes Dev. 13, 1181–1189 (1999).

    Article  CAS  Google Scholar 

  16. Shirane, M. et al. Down-regulation of p27 by two mechanisms, ubiquitin-mediated degradation and proteolytic processing. J. Biol Chem. 274, 13886–13893 (1999).

    Article  CAS  Google Scholar 

  17. Muller, D. et al. Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes. Oncogene 15, 2561–2576 (1997).

    Article  CAS  Google Scholar 

  18. Sheaff, R., Groudine, M., Gordon, M., Roberts, J. & Clurman, B. Cyclin E-Cdk2 is a regulator of p27Kip1. Genes Dev. 11, 1464–1478 (1997).

    Article  CAS  Google Scholar 

  19. Vlach, J., Hennecke, S. & Amati, B. Phosphorylation-dependent of the cyclin-dependent kinase inhibitor p27Kip1. EMBO J. 16, 5334– 5344 (1997).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  21. Koepp, D., Harper, J. W. & Elledge, S. J. How the cyclin became a cyclin: regulated proteolysis in the cell cycle. Cell 97, 431– 433 (1999).

    Article  CAS  Google Scholar 

  22. Latres, E., Chiaur, D. S. & Pagano, M. The human F-box protein β-Trcp associates with the Cul1/Skp1 complex and regulates the stability of β-catenin. Oncogene 18, 849–855 ( 1999).

    Article  CAS  Google Scholar 

  23. Winston, J. T. et al. The SCFβ-TRCP ubiquitin ligase complex associates specifically with phosphorylated destruction motifs in IκBα and β-catenin and stimulates IκBα ubiquitination in vitro. Genes Dev. 13, 270–283 ( 1999).

    Article  CAS  Google Scholar 

  24. Laney, J. & Hochstrasser, M. Substrates targeting in the system. Cell 97, 427–430 (1999).

    Article  CAS  Google Scholar 

  25. Zhang, H., Kobayashi, R., Galaktionov, K. & Beach, D. p19Skp-1 and p45Skp-2 are essential elements of the cyclin A-Cdk2 S phase kinase. Cell 82, 915–925 (1995).

    Article  CAS  Google Scholar 

  26. Yaron, A. et al. Identification of the receptor component of the IkappaBalpha-ubiquitin ligase. Nature 396, 590– 594 (1998).

    Article  CAS  Google Scholar 

  27. Lisztwan, J. et al. Association of human CUL-1 and ubiquitin-conjugating enzyme CDC34 with the F-box protein p45(SKP2): evidence for evolutionary conservation in the subunit composition of the CDC34-SCF pathway. EMBO J. 17, 368–383 (1998).

    Article  CAS  Google Scholar 

  28. Michel, J. J. & Xiong, Y. Human Cul1, but not other cullin family members, selectively interacts with SKP1 to form a complex with SKP2 and cyclin A. Cell Growth Differ. 9, 435– 449 (1998).

    CAS  PubMed  Google Scholar 

  29. Hart, M. et al. The F-box protein β-TrCP associates with phosphorylated beta-catenin and regulates its activity in the cell. Curr. Biol. 9, 207–210 (1999).

    Article  CAS  Google Scholar 

  30. Spencer, E., Jiang, J. & Chen, Z. J. Signal-induced ubiquitination of IB by the F-box protein Slimb/β-TrCP. Genes Dev. 13, 284– 294 (1999).

    Article  CAS  Google Scholar 

  31. Feldman, R. M., Correll, C. C., Kaplan, K. B. & Deshaies, R. J. A complex of Cdc4p, Skp1p, and Cdc53p/Cullin catalyzes ubiquitination of the phosphorylated Cdk inhibitor Sic1p. Cell 91, 221–230 (1997).

    Article  CAS  Google Scholar 

  32. Kominami, K. & Toda, T. Fission yeast WD-repeat protein pop1 regulates genome ploidy through ubiquitin-proteasome-mediated degradation of the CDK inhibitor Rum1 and the S-phase initiator Cdc18. Genes Dev. 11, 1548–1560 ( 1997).

    Article  CAS  Google Scholar 

  33. Skowyra, D., Craig, K. L., Tyers, M., Elledge, S. J. & Harper, J. W. F-box proteins are receptors that recruit phosphorylated substrates to the SCF ubiquitin-ligase complex. Cell 91, 209–219 (1997).

    Article  CAS  Google Scholar 

  34. Kominami, K., Ochotorena, I. & Toda, T. Two F-box/WD-repeat proteins Pop1 and Pop2 form hetero- and homo- complexes together with cullin-1 in the fission yeast SCF (Skp1-Cullin1-F-box) ubiquitin ligase. Genes Cells 3, 721– 735 (1998).

    Article  CAS  Google Scholar 

  35. Jallepalli, P. V., Tien, D. & Kelly, T. J. sud1(+) targets cyclin-dependent kinase-phosphorylated Cdc18 and Rum1 proteins for degradation and stops unwanted diploidization in fission yeast. Proc. Natl Acad. Sci. USA 95, 8159–8164 (1998).

    Article  CAS  Google Scholar 

  36. Maekawa, H., Kitamura, K. & Shimoda, C. The Ste16 WD-repeat protein regulates cell-cycle progression under starvation through the Rum1 protein in Schizosaccharomyces pombe. Curr. Genet. 33, 29– 37 (1998).

    Article  CAS  Google Scholar 

  37. Tomoda, K., Kubota, Y. & Kato, J. Degradation of the cyclin-dependent-kinase inhibitor p27 is instigated by Jab1. Nature 398, 160– 164 (1999).

    Article  CAS  Google Scholar 

  38. Yu, Z. K., Gervais, J. & Zhang, H. Human Cul1 associates with the Skp1/Skp2 complex and regulates p21(CIP1/WAF1) and cyclin D proteins. Proc. Natl Acad. Sci. USA 95, 11324–11329 (1998).

    Article  CAS  Google Scholar 

  39. Marti, A., Wirbelauer, C., Scheffner, M. & Krek, W. Interaction between ubiquitin–protein ligase SCFSKP2 and E2F-1 underlies the regulation of E2F-1 degradation. Nature Cell Biol. 1, 14–19 (1999).

    Article  CAS  Google Scholar 

  40. Pagano, M. et al. Regulation of the human cell cycle by the Cdk2 protein kinase . J. Cell Biol. 121, 101– 111 (1993).

    Article  CAS  Google Scholar 

  41. Pagano, M., Pepperkok, R., Verde, F., Ansorge, W. & Draetta, G. Cyclin A is required at two points in the human cell cycle. EMBO J. 11, 761– 771 (1992).

    Article  Google Scholar 

  42. O’Connor, P. & Jackman, J. in Cell Cycle: Materials and Methods (ed. Pagano, M.) 63–74 (Springer, New York, 1995).

    Google Scholar 

  43. Pagano, M. in Cell Cycle: Materials and Methods (ed. Pagano, M.) 271– 280 (Springer, New York, 1995).

    Google Scholar 

  44. Harlow, E. & Lane, D. in Using Antibodies. A Laboratory Manual (eds Harlow, E. & Lane, D.) 187–233 (Cold Spring Harb. Lab. Press, Cold Spring Harbor, 1998).

    Google Scholar 

  45. Faha, B., Harlow, E. & Lees, E. The adenovirus E1A-associated kinase consists of cyclin E-p33cdk2 and Cyclin A-p33cdk2. J.Virol. 67, 2456–2465 (1993).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Pagano, M., Draetta, G. & Jansen-Dürr, P. Association of cdk2 kinase with the transcription factor E2F during S phase. Science 255, 1144– 1147 (1992).

    Article  CAS  Google Scholar 

  47. Desai, D., Gu, Y. & Morgan, D. O. Activation of human cyclin-dependent kinase in vitro. Mol. Biol. Cell 3, 571– 582 (1992).

    Article  CAS  Google Scholar 

  48. Hochstrasser, M. There’s the rub: a novel ubiquitin-like modification linked to cell cycle regulation. Genes Dev. 12, 901– 907 (1998).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank G. Draetta, M. Garabedian, E. Harlow, A. Koff, E. Lees, D. Morgan, A. Pause, Y. Xiong and H. Zhang for reagents; M. Chao, J. Lukas and L. Yamasaki for critically reading the manuscript; and E. Latres and other members of M.P.’s laboratory for their contribution to this work. M.P. thanks L. Yamasaki and T.B. Balduur for their continuous support. A.H. is supported in part by grants from the Israel Ministry of Science and The Council for Tobacco Research, USA, and by a Human Frontier Science Program Organization (HFSPO) grant (RG0229); M.P. is supported by an HFSPO grant (RG0229) and by NIH RO1 grants CA76584 and GM57587.

Correspondence and requests for materials should be addressed to M.P.

Author information

Author notes

  1. Andrea C. Carrano, Esther Eytan and Avram Hershko: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Pathology and Kaplan Comprehensive Cancer Center, MSB 548, New York University Medical Center, 550 First Avenue, New York, 10016, New York, USA

    Andrea C. Carrano & Michele Pagano

  2. Unit of Biochemistry, The B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, 31096 , Israel

    Esther Eytan & Avram Hershko

Authors
  1. Andrea C. Carrano
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Esther Eytan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Avram Hershko
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Michele Pagano
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Michele Pagano.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Carrano, A., Eytan, E., Hershko, A. et al. SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27. Nat Cell Biol 1, 193–199 (1999). https://doi.org/10.1038/12013

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/12013

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