REGULATION OF CBP/P300 IN NUCLEAR RECEPTOR FUNCTION

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Steroid hormones and vitamins including glucocorticoids and vitamin A derivatives act to induce cellular differentiation by directly modulating gene transcription and are often associated with the inhibition of cell growth. Nuclear receptors bind to hormones and vitamins and activate hormone responsive genes by recruiting an acetylase co-activation complex. The broad long- term objectives of this proposal are: (a) to decipher novel regulatory mechanisms involved in receptor and coactivator function; (b) to identify new components of the receptor coactivation-complex; and (c) to dissect the regulatory properties of the complex that translate a hormonal signal into transcriptional and physiological responses. To achieve the above goals, the following Specific Aims will be pursued: (I) To test the hypothesis that both nuclear receptor interaction domains of CBP/p300 are necessary for nuclear receptor mediated gene activation. (II) To determine the roles of the histone acetyltransferase (HAT)- associated domains of CBP/p300 in transcriptional activation and to test the hypothesis that additional proteins are necessary for HAT-mediated transactivation. (III) To test the hypothesis that constant region 2 (CR2) and constant region 3 (CR3) of adenoviral oncoprotein E1A function as the HAT-inhibitory domains and to determine the mechanism of transcription inhibition by E 1 A. A combination of in vitro binding, site directed mutagenesis, deletion analysis, and mammalian cell transfection based in vivo assays will be utilized to address the above Specific Aims. Yeast two-hybrid screens will be employed to identify and characterize novel CBP/p300 regulatory proteins. These studies are clinically significant because: (a) nuclear receptor agonists and antagonists are useful in treatment of breast, and prostate cancer, leukemia, diabetes, and cardiovascular diseases; and (b) chromosomal translocations and amplification of nuclear receptors and their cofactors have been implicated in leukemia and breast cancer suggesting abnormal targeting or regulation of the activities of receptors or their coactivators may play important roles in leukemia and oncogenesis. The accomplishment of the Specific Aims described above geared towards achieving the long- term goals should provide a better understanding of the role of nuclear receptors and coactivators in transcription and in human diseases including cancer.