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Cytoplasmic relocalization and inhibition of the cyclin-dependent kinase inhibitor p27Kip1 by PKB/Akt-mediated phosphorylation in breast cancer

Abstract

The cyclin-dependent kinase inhibitor p27kip1 is a putative tumor suppressor for human cancer. The mechanism underlying p27kip1 deregulation in human cancer is, however, poorly understood. We demonstrate that the serine/threonine kinase Akt regulates cell proliferation in breast cancer cells by preventing p27kip1-mediated growth arrest. Threonine 157 (T157), which maps within the nuclear localization signal of p27kip1, is a predicted Akt-phosphorylation site. Akt-induced T157 phosphorylation causes retention of p27kip1 in the cytoplasm, precluding p27kip1-induced G1 arrest. Conversely, the p27kip1-T157A mutant accumulates in cell nuclei and Akt does not affect p27kip1–T157A-mediated cell cycle arrest. Lastly, T157-phosphorylated p27kip1 accumulates in the cytoplasm of primary human breast cancer cells coincident with Akt activation. Thus, cytoplasmic relocalization of p27kip1, secondary to Akt-mediated phosphorylation, is a novel mechanism whereby the growth inhibitory properties of p27kip1 are functionally inactivated and the proliferation of breast cancer cells is sustained.

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Figure 1: PI3K-Akt inhibition induces G1 arrest of breast cancer cells and nuclear accumulation of p27kip1.
Figure 2: Akt phosphorylates p27kip1 on threonine 157 in vitro and in vivo.
Figure 3: Akt-dependent phosphorylation of T157 promotes cytosolic localization of p27kip1 and rescues S-phase entry of p27kip1-transfected cells.
Figure 4: In breast cancer cells the PI3K-Akt pathway promotes T157 phosphorylation and cytosolic retention of p27kip1.
Figure 5: The PI3K-Akt pathway regulates T157 phosphorylation and nuclear accumulation of p27kip1 in normal mammary cells.
Figure 6: Activation of Akt correlates with T157 phosphorylation and cytosolic accumulation of p27kip1 in breast cancer.

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Acknowledgements

We thank G. Baldassarre and M. Napolitano for flow-cytometry analysis; G. Tallini for help in the statistical analysis; and J.A. Gilder for editing the text. This work was supported by grants from the Associazione Italiana Ricerca sul Cancro (AIRC), Agenzia 2000 of the Consiglio Nazionale delle Ricerche (CNR) and the Progetto Finalizzato Biotecnologie of the CNR. M.L.M. and F.V. are supported by FIRC fellowships. P.B. was the recipient of a BIOGEM fellowship.

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Correspondence to Giuseppe Viglietto.

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Viglietto, G., Motti, M., Bruni, P. et al. Cytoplasmic relocalization and inhibition of the cyclin-dependent kinase inhibitor p27Kip1 by PKB/Akt-mediated phosphorylation in breast cancer. Nat Med 8, 1136–1144 (2002). https://doi.org/10.1038/nm762

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