Control of p27 Localization and Degradation by the PI3 Kinase Akt/PKB pathway in MCF-7 Breast Cancer Cells

Author

Marissa J. Bradbury, University of Tennessee, Knoxville

Date of Award

12-2004

Degree Type

Dissertation

Degree Name

Master of Science

Major

Comparative and Experimental Medicine

Major Professor

Jay Wimalasena

Committee Members

Hwa-Chain Robert Wang, Sundar Venkatachalam

Abstract

Breast cancer is caused by a variety of environmental and genetic factors that influence cell growth and survival. Changes in the level or function of cell cycle regulatory proteins are often associated with breast cancer. Low expression of the cell cycle inhibitor, p27^Kip1, is associated with aggressive breast tumors. Clinical observations have inspired studies to analyze p27 as a potential target for signal transduction pathways in cancer cells. The stability of p27 determines it’s ability to regulate the cell cycle, and changes in transcription and translation of p27 are less influential. The regulation of the cellular localization of p27 by signal transduction pathways has become a focus of recent research. This study attempts to clarify how Akt regulates p27 localization and stability, and what implications this data may have on tumor growth and metastasis. Cells were transfected with MryAkt plasmid to artificially activate Akt. Immunoflorescence and cell fractionation results verified that MryAkt causes cytoplasmic retention of p27 in MCF-7 breast cancer cells. Labeling experiments with ³⁵S methionine demonstrated that MryAkt causes nascent p27 to accumulate in the cytoplasm .Metabolic pulse chase labeling and cyclohexamide experiments also determined that p27 was stabilized by MryAkt. Degradation in the nuclear and cytoplasmic compartments was unaffected by Akt, which supports the hypothesis that MryAkt effects only p27 localization, and the stabilization of p27 by Akt is a result of its cytoplasmic retention. Lastly, the mitogen estradiol has similar effects on p27 as MryAkt, proving that estrogen can influence p27 through the Akt pathway.

Recommended Citation

Bradbury, Marissa J., "Control of p27 Localization and Degradation by the PI3 Kinase Akt/PKB pathway in MCF-7 Breast Cancer Cells. " PhD diss., University of Tennessee, 2004.
https://trace.tennessee.edu/utk_graddiss/1917