Epstein-Barr virus (EBV) is a gamma herpesvirus that causes infectious mononucleosis and is associated with a variety of human lymphoid and epithelial cancers. Immunocompromise is a risk factor for EBV associated disease with malignancies such as post-transplant lymphoproliferative disease arising in transplant patients and central nervous system lymphoma and a proportion of systemic lymphomas being EBV associated in AIDS patients. EBV is also associated with Burkitt's lymphoma, nasopharyngeal carcinoma, nasal T cell lymphoma, and a subset of Hodgkin's lymphomas and gastric carcinomas. After primary infection an individual will remain latently infected with EBV and it is this life-long reservoir of latently infected cells that is a factor in the development of subsequent malignant disease. EBV infection of B cells in culture leads to the outgrowth of immortalized B cell lines and EBNA2 is one of the EBV encoded proteins essential for this process. EBNA2 alters cellular gene expression by targeting aDMA binding protein, CBF1/RBPJk, and switching CBF1 repressed promoters to an activated state. In its targeting of CBF1, EBNA2 mimics activated Notch signaling. It is proposed to re-evaluate the model of EBNA2 as a Notch mimic by comparing EBNA2 and NotchIC interactions with cellular proteins to identify protein partners that are shared by both EBNA2 and NotchIC versus protein partners that are unique to EBNA2 or NotchIC. High throughput screening assays provide new tools for identifying protein-protein interactions and can provide insight both into individual protein functions and into interactions between the protein of interest and cell signaling networks. Our approach will be to screen proteomic chips generated in house by the Johns Hopkins High Throughput Biology Center with EBNA2 and NotchllC and Notch2IC. Two different proteomic chips will be utilized initially: One printed with 5000 human proteins and the second a unique transcription factor specific chip containing 1003 known and predicted human transcription factors. In follow-up studies, the proteomic chips will be used as a high throughput drug screening platform to identify compounds that differentially block EBNA2 versus NotchIC interactions with cell proteins and that hence would have anti-EBV therapeutic potential.
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