Mechanisms of DNA damage-induced, p53-dependent repression of the Cdc25C Phosphat

描述（由申请人提供）：肿瘤抑制基因p53是人类癌症中最常见的突变基因[3]。它在保护哺乳动物细胞免受肿瘤发生方面起着至关重要的作用[22，23]。Cdc 25 C磷酸酶在G2/M转换的细胞周期进展中起重要作用[7-10]。其活性在响应DNA损伤时被抑制，从而促进G2检查点响应[7-10]。Cdc 25 C是p53转录抑制的靶点。Cdc 25 C的过表达使癌细胞对DNA损伤诱导的细胞死亡敏感[11，12]。有趣的是，正常的未转化细胞对这种治疗不敏感[12]。因此，Cdc 25 C阻遏抑制可能导致与DNA损伤组合的新的治疗方法。为了解决这种可能性，一个明确的分子基础p53依赖性抑制Cdc 25 C必须加以描述。到目前为止，已经提出了三种不同的抑制机制，一种涉及p53与DNA的直接结合，两种涉及不同元件的间接结合。每一种都可以提供用于抑制的分子靶标。本研究的目的是剖析p53依赖性抑制Cdc 25 C的分子基础，并探讨其相关性作为一种工具，可以利用，以提高癌症患者的化疗反应。假设：假设p53应答元件是p53与Cdc 25 C启动子结合和募集共阻遏物所必需的。其他因素，如p21，可能需要这种抑制。此外，在癌细胞中抑制这种机制可以消除DNA损伤诱导的Cdc 25 C抑制，这导致对DNA损伤诱导的细胞死亡的敏感性提高。特定目的：目的1：分析DNA损伤诱导的Cdc 25 C p53依赖性抑制的分子基础。 A）表征组蛋白脱乙酰酶（HDAC）和DNA甲基转移酶（DNMT）在Cdc 25 C阻遏中的作用。 B）研究CCAAT盒结合因子NF-γ在Cdc 25 C的p53依赖性阻遏中的作用。目的2：Cdc 25 C启动子不同元件的功能分析。 A）表征Cdc 25 C启动子中的相关元件以鉴定潜在的治疗靶标。 B）探索使用Cdc 25 C阻遏抑制作为新的治疗方法的方法。目的3：研究p21在p53抑制Cdc 25 C中的作用。A）研究p53和共阻遏物募集到Cdc 25 C启动子的p21依赖性。B）研究Rb家族蛋白在Cdc 25 C介导的响应于DNA损伤的阻遏中的作用。 公共卫生相关性：本研究的目的是剖析p53依赖性抑制Cdc 25 C的分子基础，并探讨其相关性作为一种工具，可以利用，以提高癌症患者的化疗反应。

DESCRIPTION (provided by applicant): The tumor suppressor p53 is the most commonly mutated gene found in human cancers [3]. It plays a vital role in guarding mammalian cells against tumorigenesis [22,23]. The Cdc25C phosphatase plays an important role in cell cycle progression at the G2/M transition [7-10]. Its activity is inhibited in response to DNA damage thereby contributing to the G2 checkpoint response [7-10]. Cdc25C is a target for transcriptional repression by p53. Overexpression of Cdc25C sensitizes cancer cells to DNA damage induced cell death [11, 12]. Interestingly, normal untransformed cells are not sensitized to this treatment [12]. Therefore, Cdc25C repression inhibition could lead to a novel therapeutic approach in combination with DNA damage. In order to address this possibility, a clear molecular basis for p53-dependent repression of Cdc25C must be described. As of yet, three distinct mechanisms of repression have been proposed, one involving direct binding to DNA by p53 and two indirect involving different elements. Each of which may provide molecular targets for inhibition. The present study aims to dissect the molecular basis for p53-dependent repression of Cdc25C and to explore its relevance as a tool that can be exploited in order to enhance chemotherapeutic responses in cancer patients. Hypothesis: It is hypothesized that the p53 response element is required in order for p53 to bind to the Cdc25C promoter and for the recruitment of co-repressors. Additional factors, such as p21, may be required for this repression. Furthermore, inhibition of this mechanism in cancer cells can abrogate DNA damage-induced Cdc25C repression, which results in heightened sensitivity to DNA damage-induced cell death. Specific Aims: Aim 1: Analysis of the molecular basis for DNA damage-induced p53-dependent repression of Cdc25C. A) Characterize the role of Histone Deacetylases (HDACs) and DNA methyltransferases (DNMTs) in Cdc25C repression. B) Study the role of CCAAT box binding factor, NF-Y in p53-dependent repression of Cdc25C. Aim 2: Functional analysis of different elements in the Cdc25C promoter. A) Characterize relevant elements in the Cdc25C promoter in order to identify potential therapeutic targets. B) Explore methods to use Cdc25C repression inhibition as a novel therapeutic approach. Aim 3: Study of the role of p21 in the p53-induced repression of Cdc25C. A) Investigate the p21 dependence of p53 and co-repressor recruitment to the Cdc25C promoter. B) Study the role of Rb family proteins in Cdc25C mediated repression in response to DNA damage. PUBLIC HEALTH RELEVANCE: The present study aims to dissect the molecular basis for p53-dependent repression of Cdc25C and to explore its relevance as a tool that can be exploited in order to enhance chemotherapeutic responses in cancer patients.