Regulation of Ionizing Radiation Induced DNA Damage Response

描述（由申请人提供）： 电离辐射诱导DNA双链断裂，激活ATM/ATR激酶启动的信号级联反应。损伤信号停止细胞周期进程，并为DNA损伤修复提供足够的时间。在这里，我们已经确定了一个重要的调节器，RNF 8（环指蛋白8），参与DNA损伤反应的激活。在DNA损伤后，RNF 8的FHA结构域识别磷酸化的MDC 1，其是DNA损伤反应中组蛋白H2 AX的结合伴侣，其将RNF 8靶向DNA损伤位点。组蛋白H2 AX是一种支架蛋白，可将DNA损伤反应蛋白保留在DNA损伤位点。由于RNF 8是E3泛素连接酶，我们发现RNF 8控制DNA损伤后组蛋白H2 AX泛素化。RNF 8依赖的H2 AX泛素化可以调节下游53 BP 1（p53结合蛋白1）和BRCA 1（乳腺癌易感基因1）易位到DNA损伤位点。BRCA 1和53 BP 1不仅是ATM/ATR激酶的下游靶点，而且也是重要的肿瘤抑制因子。此外，我们已经产生了RNF 8敲除小鼠。RNF 8缺失小鼠是存活的。但是雄性RNF 8-/-小鼠是不育的。RNF 8-/-小鼠对电离辐射过敏。这些表型与组蛋白H2 AX和MDC 1敲除小鼠的表型非常相似。因此，基于我们的初步研究，我们假设RNF 8在DNA损伤反应的激活中起主要作用，并参与肿瘤预防。我们提出以下实验来检验我们的假设。目的1：探讨RNF 8依赖的组蛋白泛素化在DNA损伤反应中的作用。目的2：研究RNF 8在DNA损伤后组蛋白甲基化中的作用。目的3：研究RNF 8在肿瘤抑制中的作用。总之，我们提出的研究不仅将提供对DNA损伤反应激活机制的见解，而且有助于我们理解基因组完整性的维护和肿瘤预防。 公共卫生相关性： DNA损伤反应途径的功能障碍导致了诱导肿瘤发生的巨大风险。在我们的初步数据中，我们确定了一个重要的调节因子-RNF 8在电离辐射诱导的DNA损伤反应。在这个提议中，我们计划不仅剖析RNF 8强调电离辐射诱导的DNA损伤反应的分子机制，而且还检查RNF 8在体内肿瘤发生中的作用。

DESCRIPTION (provided by applicant): Ionizing radiation induces DNA double stand breaks, which activates a signal cascade initiated by ATM/ATR kinases. The damage signals stop the cell cycle progression and allow enough time for DNA damage repair. Here, we have identified an important regulator, RNF8 (Ring Finger protein 8) that participates in the activation of DNA damage response. Following DNA damage, the FHA domain of RNF8 recognizes phosphorylated MDC1, a binding partner of histone H2AX in DNA damage response, which targets RNF8 to DNA damage sites. Histone H2AX is a scaffold protein that retains DNA damage response proteins at the DNA damage sites. Since RNF8 is an E3 ubiquitin ligase, we have found that RNF8 controls histone H2AX ubiquitination following DNA damage. RNF8-dependent H2AX ubiquitination may regulate downstream 53BP1 (p53 Binding Protein 1) and BRCA1 (Breast Cancer Susceptibility Gene 1) translocation to DNA damage sites. Both BRCA1 and 53BP1 are not only downstream targets of ATM/ATR kinases, but also important tumor suppressors. Moreover, we have generated RNF8 knock-out mice. The RNF8 null mice are viable. But male RNF8 -/- mice are sterile. RNF8 -/- mice are hypersensitive to ionizing radiation. These phenotypes are very similar to those of histone H2AX and MDC1 knock-out mice. Thus, based on our preliminary studies, we hypothesize that RNF8 plays a major role in activation of DNA damage response and participates in tumor prevention. We propose following experiments to examine our hypotheses. Aim1: To investigate the role of RNF8-dependent histone ubiquitination in DNA damage response. Aim2: To study the role of RNF8 in histone methylation in response to DNA damage. Aim3: To examine the role of RNF8 in tumor suppression. In summary, our proposed studies will not only provide insights into the mechanisms of the activation of DNA damage response, but also contribute to our understanding of the maintenance of genomic integrity and tumor prevention. PUBLIC HEALTH RELEVANCE: Dysfunctions in DNA damage response pathways cause great risks to induce tumorigenesis. In our preliminary data, we identified an important regulator - RNF8 in the ionizing radiation-induced DNA damage response. In this proposal, we plan to not only dissect the molecular mechanism of RNF8 underlining ionizing radiation-induced DNA damage response, but also examine the role of RNF8 in tumorigenesis in vivo.