A novel regulator of p53

描述（由申请人提供）：在超过一半的人类癌症中，p53肿瘤抑制基因因错义突变或缺失而失活。从生化角度看，p53作为序列特异性转录激活因子，在基因毒性应激反应中诱导参与DNA修复、细胞周期阻滞和细胞凋亡的基因。这些反应对于防止具有不稳定基因组的细胞的出现至关重要，不稳定基因组容易发生致癌转化。因此，特定基因的反激活对p53的功能至关重要。连接转录激活因子，结合特定启动子，组蛋白乙酰化和基础转录机制的共激活因子复合物对于序列特异性转录因子的功能至关重要。我们已经确定了人类ADA3（激活的改变/缺陷）蛋白作为一种新的p53结合伙伴。在酵母中，ADAS是ADA共激活因子复合体的重要组成部分，该复合体包括ADA2和组蛋白乙酰转移酶（HAT） GCN5（一般控制非抑制5）。直到最近，我们和其他人的研究才开始描述哺乳动物ADA复合物的特征。我们已经证明ADAS直接与p53相互作用，并通过促进其乙酰化和稳定性来增强其转激活功能。shrna介导的ADAS敲低表明它是p53乙酰化和DNA损伤稳定所必需的。在主要的p300乙酰化位点上不能乙酰化的突变p53不能被ADAS稳定。综上所述，这些发现使我们假设hADAS是一个中心的共激活因子成分，它招募p300 /CBP和其他HATs来乙酰化和稳定p53在其靶基因启动子上，从而增强p53介导的功能。ada3依赖性组蛋白和p53乙酰化共同提供了p53介导的DNA损伤反应的关键机制。为了验证这些假设，我们将研究ADAS在主要HAT蛋白（p300 /CBP、hGCNS和PCAF）向p53募集中的作用。此外，我们将定义ADAS在p53介导的DNA损伤（UV）细胞反应中的相对作用。辐射和化学物质)使用细胞培养模型和ADAS条件敲除小鼠（用于紫外线和致癌物诱导的肿瘤发生）。这些分析可能定义了一种新的生化途径来调节基因毒性应激后p53介导的细胞反应，以及该途径作为致癌转化障碍的相关性。阐明这一新的生化途径的作用及其组分的定义可能为未来开发基于基因和蛋白质的人类癌症诊断和治疗策略提供新的靶点。

DESCRIPTION (provided by applicant): The p53 tumor suppressor gene is inactivated by missense mutations or deletion in over half of all human cancers. Biochemically, p53 functions as a sequence-specific transcriptional activator to induce genes involved in DNA repair, cell cycle arrest and apoptosis in response to genotoxic stress. These responses are essential to prevent the emergence of cells with unstable genomes, which are prone to oncogenic transformation. Thus, transactivation of specific genes is critical for p53 function. Coactivator complexes which link transcriptional activators, bound to specific promoters, to histone acetylation and basal transcriptional machinery, are essential for the function by sequence-specific transcription factors. We have identified the human ADA3 (alteration/deficiency in activation) protein as a novel p53-binding partner. In the yeast, ADAS is an essential component of the ADA coactivator complex that include ADA2 and GCN5 (general control non-repressed 5), a histone acetyl transferase (HAT). Only recent studies by us and others have begun to characterize the mammalian ADA complexes. We have demonstrated that ADAS directly interacts with p53 and enhances its transactivation function by promoting its acetylation and stability. shRNA-mediated knockdown of ADAS indicates that it is required for p53 acetylation and stabilization upon DNA damage. Mutant p53 that can not be acetylated on major p3OO acetylation sites is not stabilized by ADAS. Collectively, these findings lead us to hypothesize that hADAS is a central coactivator component that recruits p3OO/CBP and other HATs to acetylate and stabilize p53 on its target gene promoters leading to enhancement of p53-mediated function. Together, ADA3-dependent histone and p53 acetylation provide critical mechanisms for p53-mediated DNA damage response. To test these hypotheses, we will examine the role of ADAS in the recruitment of major HAT proteins (p3OO/CBP, hGCNS and PCAF) to p53. Furthermore, we will define the relative role of ADAS in p53-mediated cellular responses upon DNA damage (UV. radiation and chemicals) using cell culture models and ADAS conditional knockout mice (for UV and carcinogen-induced tumorigenesis) that we have generated. These analyses are likely to define a novel biochemical pathway to regulate p53-mediated cellular responses following genotoxic stress, and the relevance of the pathway as a barrier to oncogenic transformation. Elucidation of the role of this new biochemical pathway and definition of its components is likely to provide new targets for future development of gene- and protein-based diagnostic and therapeutic strategies for human cancer.