G₁/S Cell Cycle Arrest Provides Anoikis Resistance through Erk-Mediated Bim Suppression

Abstract

Proper attachment to the extracellular matrix is essential for cell survival. Detachment from the extracellular matrix results in an apoptotic process termed anoikis. Anoikis induction in MCF-10A mammary epithelial cells is due not only to loss of survival signals following integrin disengagement, but also to consequent downregulation of epidermal growth factor (EGFR) and loss of EGFR-induced survival signals. Here we demonstrate that G₁/S arrest by overexpression of the cyclin-dependent kinase inhibitors p16^INK4a, p21^Cip1, or p27^Kip1 or by treatment with mimosine or aphidicolin confers anoikis resistance in MCF-10A cells. G₁/S arrest-mediated anoikis resistance involves suppression of the BH3-only protein Bim. Furthermore, in G₁/S-arrested cells, Erk phosphorylation is maintained in suspension and is necessary for Bim suppression. Following G₁/S arrest, known proteins upstream of Erk, including Raf and Mek, are not activated. However, retained Erk activation under conditions in which Raf and Mek activation is lost is observed, suggesting that G₁/S arrest acts at the level of Erk dephosphorylation. Thus, anoikis resistance by G₁/S arrest is mediated by a mechanism involving Bim suppression through maintenance of Erk activation. These results provide a novel link between cell cycle arrest and survival, and this mechanism could contribute to the survival of nonreplicating, dormant tumor cells that avert apoptosis during early stages of metastasis.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/.

ACKNOWLEDGMENTS

We thank D. Livingston, R. Weinberg, M. Ewen, S. Muthuswamy, J. Ruderman, P. Hinds, W. Sellers, and members of the Brugge lab for stimulating scientific discussions and feedback. We also thank A. Witt for preparation of some of the viral vectors, H. Irie and M. Overholtzer for scientific and technical advice, S. Isakoff for critical reading of the manuscript, P. Hinds, M.-C. Hung, and K. Polyak for plasmids, and R. Mulligan for VSV-GPG cells.

This work was supported by grants from the National Cancer Institute (CA89393 and CA80111) to J.S.B., a National Science Foundation predoctoral fellowship to N.L.C., a Susan Komen Breast Cancer Postdoctoral Fellowship to M.J.R., and support from the Breast Cancer Research Foundation to J.L. and J.S.B. J.S.B. was an American Cancer Society Research Professor during the period when this research was performed.