The ATM/E2F1 Pathway in DNA Damage and Growth Control

描述（由申请人提供）：本项目的总体目标是研究E2 F1稳定性的调节和E2 F1对DNA损伤的特定作用。虽然E2 F在细胞周期进程中的功能已经得到了很好的确立，但我们最近的工作表明E2 F1在DNA损伤反应中起着重要作用。ATM/ATR可使E2 F1磷酸化并导致E2 F1的稳定化。DNA损伤诱导的细胞凋亡需要E2 F1的诱导。因此，E2 F1似乎参与DNA损伤检查点控制，这在人类癌症中经常丢失。为了进一步了解E2 F1如何参与检查点控制，我们确定了三种与E2 F1相互作用的蛋白质，并可能在DNA损伤期间调节E2 F1。我们建议研究这些蛋白质如何调节E2 F1。 目的1：探讨TopBP 1/E2 F相互作用在转录和复制调控中的作用。 为了验证这一假设：TopBP 1对E2 F1的调节是适当的DNA损伤反应所必需的。 目的2：检测14-3-3在E2 F稳定性和活性调节中的作用。 为了检验假设：14-3-3与E2 F1的结合诱导E2 F1的稳定性和活性。 目的3：测试hHYD对于E2 F降解的作用。 为了检验假设：hHYD靶向E2 F1进行泛素化，14-3-3的结合保护E2 F1免于降解。 我们将采用生物化学，遗传学和荧光显微镜方法以及DNA微阵列分析来解决这些问题。从这项研究中获得的知识将是重要的理解正常细胞如何应对遗传毒性应激。失去适当的反应可能导致肿瘤的发展。

DESCRIPTION (provided by applicant): The overall goal of this project is to study the regulation of E2F1 stability and specific role of E2F1 for DNA damage. While the function of E2F in cell cycle progression is well established, our recent works demonstrate an important role of E2F1 for DNA damage response. ATM/ATR can phosphorylate E2F1 and lead to stabilization of E2F1. The induction of E2F1 is required for DNA damage-induced apoptosis. Thus E2F1 appears to participate in the DNA damage checkpoint control, which is often lost in human cancers. To further understand how E2F1 participates in the checkpoint control, we identified three proteins that interact with E2F1 and may regulate E2F1 during DNA damage. We propose to study how these proteins regulate E2F1. Aim 1: Explore the role of TopBP1/E2F interaction in transcription and replication control. To test the hypothesis: Regulation of E2F1 by TopBP1 is required for proper DNA damage response. Aim 2: Test the role of 14-3-3 in the regulation of E2F stability and activity. To test the hypothesis: Binding of 14-3-3 to E2F1 induces the stability and activity of E2F1. Aim 3: Test the role of hHYD for E2F degradation. To test the hypothesis: hHYD targets E2F1 for ubiquitination and binding of 14-3-3 protects E2F1 from degradation. We will employ biochemical, genetic and fluorescent microscope methodology as well as DNA microarray analysis to address these questions. The knowledge obtained from this study will be important in the understanding of how normal cells respond to genotoxic stress. A loss of the proper response may lead to tumor development.