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Phosphorylation-dependent regulation of cytosolic localization and oncogenic function of Skp2 by Akt/PKB

Nature Cell Biology volume 11, pages 420–432 (2009)Cite this article

Abstract

Skp2 is an F-box protein that forms the SCF complex with Skp1 and Cullin-1 to constitute an E3 ligase for ubiquitylation. Ubiquitylation and degradation of the p27 are critical for Skp2-mediated entry to the cell cycle, and overexpression and cytosolic accumulation of Skp2 have been clearly associated with tumorigenesis, although the functional significance of the latter is still unknown. Here we show that Akt/protein kinase B (PKB) interacts with and directly phosphorylates Skp2. We find that Skp2 phosphorylation by Akt triggers SCF complex formation and E3 ligase activity. A phosphorylation-defective Skp2 mutant is drastically impaired in its ability to promote cell proliferation and tumorigenesis. Furthermore, we show that Akt-mediated phosphorylation triggers 14-3-3β-dependent Skp2 relocalization to the cytosol, and we attribute a specific role to cytosolic Skp2 in the positive regulation of cell migration. Finally, we demonstrate that high levels of activation of Akt correlate with the cytosolic accumulation of Skp2 in human cancer specimens. Our results therefore define a novel proto-oncogenic Akt/PKB-dependent signalling pathway.

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Figure 1: Skp2 interacts with Akt.
Figure 2: Akt/PKB phosphorylates Skp2 at S72 in vitro and in vivo.
Figure 3: Phosphorylation of Skp2 is required for Skp2 E3 ligase activity and function.
Figure 4: Akt-mediated Skp2 phosphorylation regulates SCF complex formation.
Figure 5: Skp2 phosphorylation by Akt regulates the cytosolic localization of Skp2.
Figure 6: 14-3-3β interacts with Skp2 and is essential for Akt-mediated Skp2 relocalization in the cytosol.
Figure 7: Cytosolic Skp2 positively regulates cell migration.
Figure 8: Cytosolic Skp2 correlates with activation of the Akt kinase, PTEN loss, and metastasis in human cancer specimens.

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Acknowledgements

We thank D. Bohmann, P. Jackson, W. Wei, M. Pagano and M. H. Lee for reagents. We are also grateful to M. Asherov and I. Linkov in the Immunohistochemistry Pathology Core Laboratory, T. Matos for immunohistochemistry technical assistance, P. Bonner for data management, L. Lacomis for help with mass spectrometry, X. H. Zhu for technical advice, and S. Clohessy for flow cytometry analysis. We also thank M. C. Hung and L. Cantley for insightful comments and suggestions, and W. Wei for discussion and for sharing experimental results. Special thanks are extended to B. Carver and L. DiSantis for editing and critical reading of the manuscript, as well as to all the members of the Pandolfi laboratory for comments and discussion. This work was supported by NIH grants RO1 CA-71692 and CA-74031 to P.P.P. and by M. D. Anderson Cancer Center Trust Scholar funds to H.K.L. The Microchemistry & Proteomics Core is supported by NIH grant P30 CA-08748.

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Author notes

  1. Zhenbang Chen

    Present address: Present address: Department of Cancer Biology, Meharry Medical College, 1005 Dr D. B. Todd Jr Boulevard, Nashville, Tennessee 37067-3599, USA.,

Authors and Affiliations

  1. Department of Pathology, Cancer Biology and Genetics Program, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, 10021, New York, USA

    Hui-Kuan Lin, Guocan Wang, Zhenbang Chen, Julie Teruya-Feldstein & Pier Paolo Pandolfi

  2. Beth Israel Deaconess Cancer Center and Department of Medicine and Pathology, Cancer Genetics Program, Beth Israel Deaconess Medical Center, Harvard Medical School, 330 Brookline Avenue, Boston, 02215, Massachusetts, USA

    Guocan Wang, Zhenbang Chen & Pier Paolo Pandolfi

  3. Department of Molecular and Cellular Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, 77030, Texas, USA

    Hui-Kuan Lin, Chia-Hsin Chan & Wei-Lei Yang

  4. Molecular Pharmacology and Chemistry Program,

    Yan Liu & Stephen Nimer

  5. Molecular Biology Program, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, 10021, New York, USA

    Hediye Erdjument-Bromage & Paul Tempst

  6. Department of Molecular and Cellular Biology, Medical Institute of Bioregulation, Kyushu University, Fukuoka, 812-8582, Fukuoka, Japan

    Keiichi I. Nakayama

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Contributions

H.K.L. and P.P.P. designed the experiments and wrote the manuscript. H.K.L., G.W. Z.C., Y.L., C.H.C. and W.L.Y. performed the experiments. J.T. performed the immunohistochemistry and analysed the data. K.I.N. provided the Skp2−/− mice. S.N. provided valuable suggestions. H.E. and P.T. performed the mass spectrometry analysis.

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Correspondence to Hui-Kuan Lin or Pier Paolo Pandolfi.

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Lin, HK., Wang, G., Chen, Z. et al. Phosphorylation-dependent regulation of cytosolic localization and oncogenic function of Skp2 by Akt/PKB. Nat Cell Biol 11, 420–432 (2009). https://doi.org/10.1038/ncb1849

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