Date of Award

8-2009

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Comparative and Experimental Medicine

Major Professor

Howard K. Plummer III

Committee Members

Madhu Dhar, Karla J. Matteson, Seung J. Baek

Abstract

Breast cancer is a leading cause of cancer death and in 2009, the American Cancer Society estimates that over 192,000 new cases of breast cancer will be diagnosed, and over 40,000 women will die from breast cancer. Estrogen (E2) is required for normal female development and reproduction, but long-term exposure is carcinogenic and considered a risk factor for breast cancer. Membrane ion channels are essential for cell proliferation and are suggested to have a role in cancer, especially potassium channels. In the present study, we investigate the effects of estrogen and the estrogen antagonist ICI182780 on G-protein inwardly rectifying potassium channels (GIRK) in estrogen responsive MCF-7 breast cancer cells. GIRK1 and GIRK2 specific channels are thought to play a major role in rapid channel activation. We found increases in GIRK1 and GIRK2 membrane protein levels in response to estrogen treatment, as well as increases in intracellular potassium concentrations and cellular proliferation. ICI182780 treatment increased GIRK1, GIRK2 and GIRK4 membrane protein levels but resulted in an initial decrease in intracellular potassium concentration and decreased cell proliferation. GIRK1 RNAi knockdown decreased estrogen receptor alpha protein levels and activation. In addition, estrogen treatment resulted in increased phosphorylation of specific members of GPCR and MAPK signaling pathways that have been shown to be responsive to GIRK1 knockdown. Using microarray analysis of nontreated and E2 treated MCF7 cells, we observed 489 differentially expressed genes (283 upregulated and 206 downregulated) that were comprised largely of transcription and cell cycle associated genes. This study identified several human cell cycle associated genes that are both responsive to E2 treatment and are functionally correlated with GIRKs. Five genes were selected for further analysis by real time PCR. Our data suggests that specific GIRK channel composition at the cell membrane may be stimulated by estrogen exposure or downstream targets of estrogen signaling and may contribute to increased cell proliferation in MCF-7 breast cancer cells. Taken together, these data add further support of GIRK involvement in cancer progression and identify some potential biological roles of GIRKs in cancer biology beyond the initilal findings of GIRK1 assciation in metastatic breast cancer.

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