Role of p300/CBP in SV40 Large T Antigen Transformation

Dana-Farber Cancer Institute, Boston MA

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Abstract

DESCRIPTION (provided by applicant): This proposed project will examine whether activities of p300 and CBP contribute to the transforming function of SV40 large T antigen (LT). A requirement for p300/CBP association with LT will be studied using a model of human carcinogenesis. In this multi-step system, expression of LT and the active telomerase subunit promotes immortalization. Full tumorigenic conversion is then conferred through expression of H-Ras and SV40 small T antigen. Since LT activity is required for the initial steps of transformation, this provides an excellent model to expand current understanding of cellular pathways targeted by LT. We have identified an interaction between LT and the p300/CBP transcriptional coactivators and implicate a bridging function of p53. Our first goal is to determine whether the acetylation of LT that we have found under cellular conditions can be attributed to p300/CBP binding and whether this requires an intermediary role of p53. This will be addressed using p53-/- MEF cells or purified proteins. We will then determine whether p300/CBP activity contributes to the overall transforming properties of LT. The use of competitive p300 fragments to disrupt LT binding to endogenous p300/CBP, with or without a fused acetyltransferase domain, will be used to determine whether LT binding recruits p300/CBP acetyltransferase activity or sequesters p300/CBP from normal cellular targets. Finally, we will determine whether the stabilization of inactive p53 can further contribute to the transformation process through a gain of function that serves to sequester p300/CBP from normal cellular functioning.