Dissection of a fundamental step in gene activation using the HTLV-1-encoded Tax

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DESCRIPTION (provided by applicant): The human T-cell leukemia virus type-1 (HTLV-1) is a retrovirus etiologically associated with Adult T-cell Leukemia (ATL). ATL is a malignant cancer associated with poor prognosis. Although 15-25 million people are estimated to be infected with HTLV, only 2-5 percent of these infections develop into ATL (for review see [2]). A major player in the malignant transformation of infected cells is the virally expressed oncoprotein, Tax. Tax is a potent activator of transcription from the HTLV-1 promoter. Tax recruits the cellular proteins cAMP-response element binding protein (CREB) and p300/CREB-binding protein (CBP) to viral CRE sites located in the HTLV-1 LTR (for review see [1]). CBP/p300 is involved in numerous cellular pathways and understandably, deficiencies in CBP/p300 cause many types of cancer. Despite the participation of CBP/p300 in the regulation of many cellular pathways, little is known about the mechanism or its role in coactivator-mediated gene activation. We propose that a major activity of CBP/p300 is to recruit general transcriptional machinery to the promoter and that Tax stimulates this activity by binding to CBP/p300. This binding likely initiates structural changes in distinct domains of CBP/p300 that favor recruitment of general transcriptional machinery. Overall, the proposed studies will utilize interaction assays with targeted point mutants, individual domain isolations and deletions that target the interactions between Tax-CBP/p300. Competition and activity assays will enable us to functionally and biophysically characterize Tax-CBP/p300 interactions. Tax and CBP/p300 mutants developed will also be used to investigate structural changes in various domains of CBP/p300 and how these binding- induced changes relate to transcriptional machinery recruitment and activity. These studies have implications for viral and cellular mechanisms of activation, adding to the generally weak knowledge base on the role of coactivator contribution to this process. PUBLIC HEALTH RELEVANCE: Human T-cell Leukemia virus type-1 (HTLV-1) causes several diseases including aggressive Adult T-cell Leukemia (ATL). This application investigates mechanisms of viral gene activation mediated by the viral oncoprotein, Tax. The project has many diverse and unique training opportunities designed to further my education amongst highly experienced and supportive personal.