A Ligand-Binding Study of G-Protein Coupled Estrogen Receptors in MCF-7 Breast Cancer Cells and 17ß-Estradiol

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2015-05
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Okudara, Rance
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Ng, Leung Ho
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Biology
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University of Hawaii at Manoa
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It is known that some cancer cells have receptors for hormone ligands. For example, breast cancer cells many times have receptors for estrogens. When estrogens bind to the receptors of these cells, a response—such as growth—is generated within the cells. Recently, a new trans-membrane protein has been found to be associated with estrogenic pathways, called G-protein-coupled estrogen receptor (GPER). It is believed that GPERs are embedded in the membranes portions of cells. Study of GPER raises exciting new questions because roles for GPER have already been implicated for almost every system of the body. Previous research has already demonstrated the binding of estrogens to other estrogen receptors. Several assays demonstrating the biological effects of estrogen in cells expressing GPERs suggest that GPERs are likely receptors for estrogen. However, direct molecular evidence of GPER’s ability to bind estrogen is still lacking. This project uses the MCF-7 breast cancer cell line, affinity chromatography, and Western Blotting techniques in an effort to provide direct molecular evidence showing that GPERs bind to 17β-estradiol (E2). The objectives of the experiment were to grow and lyse the cells to release the GPER proteins, purify the proteins by using an estradiol affinity chromatography column, and detect whether GPERs binds to E2 based on which fraction it is found in using a Western Blot. Currently, we are working on optimizing the estradiol affinity chromatography column and Western Blot.
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32 pages
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