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p53 dependent S checkpoint in early mouse embryos and radiation induction of genomic instability

早期小鼠胚胎中p53依赖性S检查点和基因组不稳定性的辐射诱导

基本信息

批准号：
14208067
负责人：
NIWA Ohtsura
金额：
$ 13.73万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-14208067/
关键词：
p53 dependent S checkpoint suppression of replication fork progression transactivation independent DNA binding domain ATM dependency sperm irradiated zygotes pink eyed unstable allele reversion mutation ゲノム防御機構マウス初期胚遅延アポトーシス遅延細胞周期

项目摘要

Elevated levels of mutations were found at the maternally derived alleles of the Ms6hm minisatellite locus and the pink-eyed unstable locus in F1 mice born to male mice irradiated at the spermatozoa stage. These mutations demonstrated the operation of untargeted and delayed recombinational mutations due to genomic instability which was induced during early mouse development by radiation DNA damage introduced by the irradiated sperm. In order to elucidate the molecular mechanism of genomic instability, we have undertaken studies of damage response in sperm irradiated mouse embryos.A p53 responsible reporter plasmid microinjected into zygotes was transcriptionally activated when they zygotes were fertilized with irradiated sperm. This pronuclear cross-talk was found to suppress DNA synthesis of female pronucleus as well as of male pronucleus in sperm irradiated zygotes. This suppression of DNA synthesis was not observed in p53-/-zygotes which was restored when the sperm irradiated p53-/-zygotes were microinjected with p53 protein, suggesting a novel pathway of S checkpoints. Microinjection of mutant p53 proteins into sperm irradiated p53-/-zygotes demonstrated that this p53 dependent S checkpoint required DNA binding domain of p53,but the transactivation domain.The p53 dependent S checkpoint was analyzed in primary mouse and found operate at doses below 2 Gy. In addition, analyses of DNA fiber elongation by double labeling the cells with Id U and Cld U showedthatthe p53 dependent S checkpoint suppressed DNA synthesis by slowing down of replication fork progression. This slowing down of replication was associated with a higher rate of recombination between sister chromatids which can explain the untargeted recombination of minisatellite. Delayed recombination observed at the pink-eyed unstable allele requires yet another mechanism of damage memory for which much more to be studied yet.

在精子期辐照的雄性小鼠所生的F1小鼠中，发现Ms6hm小卫星位点和粉红眼不稳定位点的母系等位基因突变水平升高。这些突变表明，在小鼠早期发育过程中，受辐照精子引起的辐射DNA损伤导致了基因组不稳定，从而导致了非靶向和延迟重组突变的发生。为了阐明基因组不稳定性的分子机制，我们对精子辐照小鼠胚胎的损伤反应进行了研究。将p53负责报告质粒微注射到受精卵中，受精卵与辐照精子受精后转录激活。在精子辐照受精卵中，发现这种原核串扰抑制了雌性原核和雄性原核的DNA合成。在p53-/-受精卵中没有观察到这种DNA合成的抑制，当精子照射p53-/-受精卵并微量注射p53蛋白时，这种抑制得以恢复，这表明存在一种新的S检查点途径。将突变型p53蛋白显微注射到辐照的p53-/-受精卵中，结果表明，这种p53依赖的S检查点需要p53的DNA结合域，而不是反激活域。在原代小鼠中分析了p53依赖性S检查点，发现在低于2 Gy的剂量下起作用。此外，通过双标记Id U和ld U细胞对DNA纤维伸长的分析表明，p53依赖的S检查点通过减缓复制叉的进展来抑制DNA合成。这种复制的减慢与姐妹染色单体之间较高的重组率有关，这可以解释小卫星的非靶向重组。在红眼不稳定等位基因中观察到的延迟重组需要另一种损伤记忆的机制，对此还需要进行更多的研究。

项目成果

期刊论文数量（17）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Predisposition to mouse thymic lymphomas in response to ionizing radiation depends on variant alleles encoding metal-responsive transcription factor-1 (Mtf-1).

电离辐射对小鼠胸腺淋巴瘤的易感性取决于编码金属反应转录因子 1 (Mtf-1) 的变异等位基因。

DOI：
发表时间：
2005
期刊：
Oncogene 24
影响因子：
0
作者：
Hashimoto;T.;Yamaoka;K.;Sakai;Y.;A.OOTSUYAMA;Y.Tamura
通讯作者：
Y.Tamura

Generation of multipotent stem cells from postnatal mouse testis.

从出生后小鼠睾丸中产生多能干细胞。

DOI：
发表时间：
2004
期刊：
Cell 119
影响因子：
0
作者：
Kanatsu-Shinohara M.;Inoue K.;Lee J.;Yoshimoto M.;Ogonuki N.;Miki H.;Baba T.;Kazuki Y.;Toyokuni S.;Oshimura M.;Heike T.;Nakahata T.;Ishino F.;Ogura A.;Shinohara T
通讯作者：
Shinohara T

Induced genomic instability in irradiated germ cells and in the offspring; reconciling discrepancies among the human and animal studies