Development of molecular bio-dosimeiry and monitor mice for the detection of radiation dose exposed by tritium water

我们试图建立从全身到分子水平的氚水生物剂量测定系统。1)建立对辐射敏感的Revl转基因小鼠我们研究了小鼠和人的Revl基因，该基因是UmuC/DinB/XPV基因家族的成员，在自发突变中起重要作用。小鼠Revl蛋白的生化分析表明，小鼠Revl蛋白具有与人REV1蛋白相同的脱氧胞基转移酶活性。辐照诱导小鼠胚胎成纤维细胞表达Revl。在此基础上，我们尝试建立Revl转基因小鼠，建立辐射过敏小鼠模型。我们将携带mRevl的MT-1质粒从金属蛋白启动子下游引入c57bl小鼠。我们已经有36只老鼠携带了mRevl。我们将检查mRevl的表达。在DNA辐射双链断裂的修复机制中，历史H2AX、Ku70、Ku80和Tip60被认为起着重要作用。我们观察到H2AX在辐照后被磷酸化（γ-H2AX）并形成病灶。为了将焦点形成用于dna双链断裂的剂量测定，我们纯化了H2AX复合物，并通过MS/MS光谱分析鉴定了其新组分。我们制备了针对这些成分的抗体，并进行了免疫组织化学分析，以检测dna双链断裂处的病灶。此外，我们发现NBS1在辐射暴露后定位于γ- h2ax焦点。目前正在研究病灶数目与dna双链断裂之间的关系。

We tried to develop bio-dosimetry system of tritium water at the level from whole body to molecule.1) Development of Revl transgenic mice which are sensitive to radiation exposureWe characterized mouse and human Revl gene which was a member of the UmuC/DinB/XPV gene family, played important roles in spontaneous mutations. Biochemical analysis of the mouse Revl protein revealed that the mouse Revl protein possessed a deoxycytidyl transferase activity as human REV1 protein. The expression of mouse Revl in embryonic fibroblasts was induced by radiation exposure. Based on these knowledge, we try to establish Revl transgenic mouse to develop the hypersensitive mouse model to radiation exposure. We introduced MT-1 plasmid carrying mRevl down stream the metallotuionein promoter into C57BLmice. We already got 36 mice carrying mRevl. We will check the expression of mRevl.2) Development of molecular bio-dosimetry system for tritium waterIn repair machinery of DNA double trand break by radiation, Historic H2AX, Ku70, Ku80 and Tip60 are thought to play an important role. We observed that H2AX was phosphorylated (γ-H2AX) and formed foci after irradiation. In order to use the foci formation for the dosimetry of DNAdouble strand breaks, we purify the constitutive H2AX complex and identify its new components by MS/MS spectrometric analysis. We prepared antibody against these components and performed the immunohistochemical analysis to detect the foci at DNAdouble strand breaks. Furthermore, we found that NBS1 localized toγ-H2AX foci after radiation exposure. Relationship between numbers of foci and DNAdouble strand breaks is under investigation.