A novel regulator of p53

描述(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。在生物化学方面，P53作为一种序列特异性转录激活剂，诱导参与DNA修复、细胞周期停滞和细胞凋亡的基因，以响应遗传毒性应激。这些反应对于防止基因组不稳定的细胞的出现至关重要，因为基因组不稳定的细胞容易发生致癌转化。因此，特定基因的反式激活对P53的功能至关重要。共激活子复合体将转录激活子与特定启动子结合，连接到组蛋白乙酰化和基本转录机制，对于序列特异性转录因子的功能是必不可少的。我们已经确定人类ADA3(激活改变/缺陷)蛋白是一种新的P53结合伙伴。在酵母中，ADAS是ADA共激活复合体的重要组成部分，包括ADA2和组蛋白乙酰转移酶(HAT)GCN5(一般控制的非抑制5)。直到最近我们和其他人的研究才开始描述哺乳动物的ADA复合体。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反式激活功能。ShRNA介导的ADAS被敲除表明，它是P53乙酰化和DNA损伤后稳定所必需的。不能在主要p3OO乙酰化位点上乙酰化的突变型p53不被ADAS稳定。综上所述，这些发现使我们假设HADAs是一个中心共激活成分，它招募p3OO/CBP和其他HAT来乙酰化和稳定其靶基因启动子上的P53，从而增强P53介导的功能。ADA3依赖的组蛋白和P53乙酰化共同为P53介导的DNA损伤反应提供了关键机制。为了验证这些假说，我们将研究ADAS在主要HAT蛋白(p3OO/CBP、hGCNS和PCAF)向P53募集中的作用。此外，我们将确定ADAS在p53介导的细胞DNA损伤反应中的相对作用(UV。使用我们培育的细胞培养模型和ADAS条件性基因敲除小鼠(用于紫外线和致癌物诱导的肿瘤形成)。这些分析可能定义了一种新的生化途径来调节遗传毒性应激后P53介导的细胞反应，以及该途径作为致癌转化的屏障的相关性。阐明这一新的生化途径的作用及其成分的定义可能为未来基于基因和蛋白质的人类癌症诊断和治疗策略的发展提供新的靶点。

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.