Dynamics of higher order nuclear architecture upon DNA damage

DNA 损伤后高阶核结构的动力学

• 批准号: 17510045
• 负责人: TASHIRO Satoshi
• 金额: $ 2.3万
• 依托单位: HIROSHIMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17510045/
• 关键词: ionizing irradiation; higher order nuclear structure; bioimaging; DNA double strand breaks

DNA double strand breaks (DSBs) are most serious cell damage by ionizing irradiation because failure to properly repair them may lead to tumorigenetic chromosomal translocations. Several repair proteins are shown to form higher order nuclear structures, radiation induced repair foci (RIRF), after gamma irradiation. However, how nuclear structure is reorganized upon DNA damage in human cells is poorly understood. To investigate the dynamic reorganization of nuclear structure after ionizing irradiation, we examined the topological and chronological relationships of RIRF. For this purpose, we applied multicolor immunofluorescence analysis to study the localization of more than 4 proteins in the same nucleus. Using this technique, we studied the localization of RIRF formed by γ-H2AX, RAD51, 53BP1 and ubiquitinated proteins (FK2 foci) in human fibroblast cell lines at various time points after gamma irradiation. As a result, we found that FK2 foci are formed at nuclear sites where g-H2AX foci are formed at 30 min after gamma irradiation at 2 Gy in normal cells. 53BP1 foci are then formed at γ-H2AX/FK2 foci more than one hour after induction of DSBs by 2 Gy ionizing irradiation. The g-H2AX/FK253BP1 foci formation was observed more than one hour after 12 Gy ionizing irradiation. To study the role of ATM and ligase IV in RIRF formation, we performed ionizing irradiation of ATM deficient cells. Formation of RIRF both after 2 and 12 Gy irradiation was significantly repressed in ATM deficient cells suggesting the involvement of ATM in RIRF formation after ionizing irradiation. On the other hand, ligase IV deficient cells, carrying the disturbed end-joining repair system of DSBs, showed significantly higher numbers of γ-H2AX/FK2 foci both at 2 and 12 Gy irradiation suggesting the involvement of ubiquitination system in recombinational repair pathway of DSBs.

DNA双链断裂(DSB)是电离辐射对细胞造成的最严重的损伤，因为如果不能适当地修复它们，可能会导致肿瘤的染色体易位。几种修复蛋白在伽玛射线照射后形成高阶核结构，即辐射诱导修复焦点(RIRF)。然而，人类细胞DNA损伤后核结构是如何重组的，目前还知之甚少。为了研究电离辐射后核结构的动态重组，我们研究了RIRF的拓扑和时间关系。为此，我们应用多色免疫荧光分析方法研究了4种以上蛋白质在同一细胞核中的定位。利用这一技术，我们研究了γ-H_2AX、RAD51、53BP1和泛素化蛋白(FK2焦点)在人成纤维细胞系中的定位。结果发现，在正常细胞中，2Gy伽玛射线照射后30min，在g-H_2AX焦点形成的核部位形成了FK2焦点。γ-H_2AX/FK_2灶经2Gy射线照射诱导后1小时以上即可形成53BP1灶。12Gy射线照射后1小时以上可观察到g-H_2AX/FK253BP1细胞的病灶形成。为了研究ATM和连接酶IV在RIRF形成中的作用，我们对ATM缺陷细胞进行了电离辐射。在ATM缺陷细胞中，2和12Gy射线照射后RIRF的形成均受到显着抑制，提示ATM参与了电离辐射后RIRF的形成。另一方面，携带DSB末端连接修复系统紊乱的连接酶IV缺陷细胞，在2Gy2和12Gy射线照射下，γ-H_2AX/FK2病灶数目均显著增加，提示泛素化系统参与了DSB的重组修复途径。

项目成果

Nuclear architecture : topology and function of chromatin-and non-chroamtin nuclear domains. In "Nuclear dynamics : Approaches from Biochemistry, molecular Cell biology and visual Biology".

核结构：染色质和非染色质核结构域的拓扑和功能。

• 发表时间: 2007
• 作者: Satoshi Tashiro;et al.
• 通讯作者: et al.

Nuclear positioning of the BACH2 gene in BCR-ABL positive leukemic cells

• DOI: 10.1002/gcc.20390
• 发表时间: 2007-01-01
• 期刊: GENES CHROMOSOMES & CANCER
• 影响因子: 3.7
• 作者: Ono, Atsushi;Kono, Kazuteru;Tashiro, Satoshi
• 通讯作者: Tashiro, Satoshi

Plasmacytic transcription factor Blimp-1 is repressed by Bach2 in B cells

• DOI: 10.1074/jbc.m607592200
• 发表时间: 2006-12-15
• 期刊: JOURNAL OF BIOLOGICAL CHEMISTRY
• 影响因子: 4.8
• 作者: Ochiai, Kyoko;Katoh, Yasutake;Igarashi, Kazuhiko
• 通讯作者: Igarashi, Kazuhiko

Nuclear arch i tecture : topology and function of chroma t in-and non-chroamtin nuc1ear domains. In Nuclear dynamics : Approaches from Biochemistry, molecular Cell biology and visual Biology"

核结构：染色质核域和非染色质核域的拓扑结构和功能。

• 发表时间: 2007
• 期刊: Springer
• 作者: Satoshi Tashiro;et al.
• 通讯作者: et al.