Genetic and Molecular Analysis of DNA Damage induced Apoptosis and Cell Cycle Arrest in C. elegans

我们对DNA损伤的感知和程序性细胞死亡机制的激活之间的分子联系还知之甚少，因此以线虫生殖系为模型系统来研究离子辐射诱导的细胞死亡和细胞周期停滞。正如GA 703/1-1中提出的那样，我们克隆了线虫rad-5检查点基因，并对其进行了遗传特征分析，结果证明该基因在进化上是保守的2。此外，作为使用功能基因组学和基于RNAi的方法的合作努力的一部分，我们可以证明以前在酵母检查点信号中隐含的保守基因也在线虫DNA损伤反应途径中发挥作用。此外，我们进行了正向遗传学筛查，通过筛查我们发现了与DNA损伤诱导的细胞凋亡有关的新的候选突变(A.Gartner，未发表)。最后，我们可以定义哺乳动物P53肿瘤抑制基因的功能蠕虫同源物。作为该提案(GA 703/1-1项目的扩展)的一部分，我们旨在进一步描述新的rad-5-检查点基因(TASK-1)。此外，我们计划定位克隆与gt11突变相对应的检查点基因，这是我们在试点基因筛查中确定的(TASK-2)。通过继续我们的基因筛查，我们的目标是定义新的DNA损伤检查点突变，我们希望开始定位克隆相应的基因(任务3)。最后，我们计划开始使用生化、遗传学和细胞学的方法，将新的和已知的损伤反应基因放入遗传和生化途径(任务4)。综上所述，我们相信有助于更深入地了解DNA损伤反应。

We still know relatively little about the molecular connection between the sensing of DNA damage and the activation of the programmed cell death machinery and therefore use the C. elegans germ line as model system to study ionic radiation induced cell death and cell cycle arrest. As proposed in Ga 703/1-1 we cloned and genetically characterized the C. elegans rad-5 checkpoint gene that turned out to be evolutionarily conserved2. Furthermore, as part of a collaborative effort using functional-genomics and RNAi-based approaches we could show that conserved genes, which were previously implied in yeast checkpoint signaling, also function in C. elegans DNA damage response pathways3. Furthermore, we performed a forward genetics screen, through which we found novel candidate mutations involved in DNA damage induced apoptosis, (A. Gartner, unpublished). Finally, we could define a functional worm homolog of the mammalian p53 tumor suppressor4 . As part of this proposal (extension of the project Ga 703/1-1) we aim at further characterizing the novel rad-5-checkpoint gene (task-1). In addition, we plan to positionally clone the checkpoint gene corresponding to the gt11 mutation, which we have identified in a pilot genetic screen (task-2). By continuing with our genetic screen, we aim at defining new DNA damage checkpoint mutations and we want to start to positionally clone the corresponding genes (task-3). Finally we plan to start to put novel and already known damage response genes into genetic and biochemical pathways (task-4) using biochemical genetic and cytological methods. In summary we believe to contribute to a deeper understanding of DNA damage response.