Identification of ESE1 SAR-Domain Binding Proteins

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Transcription factors such as, those in the Ets family, regulate oncogene and tumor suppressor expression. The over-expression of several Ets factors, including ESE1, is highly associated with breast cancer. Using anchorage independent colony formation in soft agar assays, we have shown that stable expression in the cytoplasm of a 40AA highly acidic domain, referred to as the Serine and Aspartic Rich (SAR) domain of ESE1, is necessary and sufficient for MCF12-A mammary cell transformation. The MCF-12A cell-line is immortalized but not fully transformed and does not express ESE1 endogenously. These data suggest ESE1 plays a role in mammary cell transformation due to its cytoplasmic expression. Thus, the overall hypothesis of this proposal is that the SAR domain of ESE1 acts by binding to specific cytoplasmic regulatory proteins that govern the transformed breast cell phenotype. My specific aims are: 1) Purify cytoplasmic binding partner(s) of the SAR domain using GST-SAR affinity binding assays, 2) Identify the protein(s) purified in specific aim 1 using mass spectrometry, 3) Determine functional relevance of SAR binding protein(s) (SBP) in mammary cell transformation by loss of function and gain of function assays.