REGULATION OF CBP/P300 IN NUCLEAR RECEPTOR FUNCTION

类固醇激素和维生素，包括糖皮质激素和维生素A衍生物，通过直接调节基因转录来诱导细胞分化，并且通常与细胞生长的抑制有关。 核受体与激素和维生素结合，并通过招募乙酰化酶共激活复合物来激活激素应答基因。 该提案的广泛的长期目标是：（a）破译涉及受体和共激活因子功能的新的调节机制;（B）鉴定受体共激活复合物的新组分;和（c）剖析将激素信号翻译成转录和生理反应的复合物的调节特性。 为了实现上述目标，将追求以下具体目的：（I）检验CBP/p300的两个核受体相互作用结构域对于核受体介导的基因激活是必需的这一假设。 (II)确定CBP/p300的组蛋白乙酰转移酶（HAT）相关结构域在转录激活中的作用，并检验HAT介导的转录激活所需的其他蛋白质的假设。 (III)验证腺病毒癌蛋白E1 A的恒定区2（CR2）和恒定区3（CR 3）是HAT抑制结构域的假设，并探讨E1 A抑制转录的机制。将使用体外结合、定点诱变、缺失分析和基于哺乳动物细胞转染的体内试验的组合来解决上述特定目的。酵母双杂交筛选将用于鉴定和表征新的CBP/p300调节蛋白。 这些研究具有临床意义，因为：（a）核受体激动剂和拮抗剂可用于治疗乳腺癌、前列腺癌、白血病、糖尿病和心血管疾病;和（B）染色体易位和核受体及其辅因子的扩增与白血病和乳腺癌有关，表明受体或其辅激活因子的活性的异常靶向或调节可能起重要作用在白血病和肿瘤发生中的作用。 上述具体目标的实现应有助于更好地理解核受体和辅激活因子在转录和人类疾病（包括癌症）中的作用。

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.