TULANE CANCER GENETICS COBRE: ATR KINASE AND UV-INDUCED CELL CYCLE CHECKPOINTS

细胞周期检查点是真核细胞对电离辐射(IR)和紫外线(UV)等DNA损伤物质做出最佳反应的机制之一。这些细胞周期进程的控制器可能会增强遗传稳定性，限制生物体内的肿瘤发生。对这些信号通路中生化步骤的描述将有助于我们更好地理解：1)正常的细胞周期控制；以及2)DNA损伤后细胞转化的机制。虽然IR诱导的细胞周期检查点已经得到了广泛的研究，但关于UV诱导的细胞周期停滞的研究还很少。我们的长期目标是阐明控制紫外线诱导的细胞周期检查点的信号转导途径。我们的初步观察表明，ATR是紫外线诱导的S期和G2/M期检查点所必需的ATR。ATR是磷脂酰肌醇激酶相关蛋白家族中的一员，它对信号的存在是必不可少的。 DNA损伤或复制阻断并激活细胞周期检查点。由于大多数DNA损伤诱导的信号通路包括调节细胞周期进程的激酶依赖的信号级联，我们推测ATR通过其下游靶点的磷酸化在UV诱导的细胞周期检查点的激活中发挥核心作用。我们提出了三个具体目标来剖析ATR在紫外线诱导的周期检查点中的作用。在……里面 具体目的1，我们将研究BRCA1、NBS1和SMC1三个蛋白是否参与了ATR依赖的S期阻滞，以及ATR是否使这些蛋白磷酸化以促进这种阻滞。在特定目标2中，我们将研究紫外线诱导的G2细胞停滞过程中ATR和p38 MAP激酶的生化联系。在具体目标3中，我们将调查RAD 17是否是一个DNA损伤传感器，用于招募ATR来调节检查点。这些研究将为ATR激酶的作用和机制提供新的见解，并在此过程中揭示 环境DNA损伤后细胞周期调控机制的研究。

Cell cycle checkpoints are among the mechanisms developed by eukaryotic cells to optimally respond to DNA damage agents, such as ionizing radiation (IR) and ultraviolet radiation (UV). These controllers of cell cycle progression presumably enhance genetic stability and limit tumorigenesis in the organism. Characterization of the biochemical steps in these signaling pathways will help improve our understanding of: 1) normal cell cycle control; and 2) mechanisms of cellular transformation following DNA damage. While IR-induced cell cycle checkpoints have been extensively studied, less is known about UV-induced cell cycle arrest. Our long term goal is to elucidate the signal transduction pathways that control UV-induced cell cycle checkpoints. Our preliminary observations suggest that the ATM-RAD3 related kinase, ATR, is required for UV-induced S-phase and G2/M checkpoints. ATR is a member of the phosphatidylinositol kinase related protein family that is essential for signaling the presence of DNA damage or replication blocks and activating cell cycle checkpoints. Since most of the DNA damage-induced signaling pathways consist of kinase-dependent signaling cascades that regulate cell cycle progression, we reason that ATR plays a central role in activation of UV-induced cell cycle checkpoints through phosphorylation of its downstream targets. We propose three specific aims to dissect the role of ATR in UV-induced cycle checkpoints. In Specific Aim 1, we will investigate whether BRCA1, NBS1 and SMC1 are involved in ATR-dependent S-phase arrest, and whether ATR phosphorylates these proteins to facilitate the arrest. In Specific Aim 2, we will investigate the biochemical linkage of ATR kinase and p38 MAP kinase in UV-induced G2 arrest. In specific Aim 3, we will investigate whether RAD 17 is a DNA damage sensor for recruiting ATR to regulate the checkpoints. These studies will provide novel insights into the roles and mechanisms of the ATR kinase and, in the process, should shed light on mechanisms involved in cell cycle control after environmental DNA damage.