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Study of cellular responses to single-strand break

细胞对单链断裂反应的研究

基本信息

批准号：
10480131
负责人：
YASUI Akira
金额：
$ 7.55万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10480131/
关键词：
DNA damage DNA repair DNA break circadian rhythm photo repair excision repair 塩基損傷 DNA単鎖切断 PARP XRCC1 塩基除去修復単鎖切断紫外線損傷

项目摘要

Although single-strand breaks (SSBs) occur frequently, the cellular responses and repair of SSB are not well understood. To address this, we established mammalian cell lines expressing Neurospora crassa UV damage endonuclease (UVDE), which introduces a SSB with a 3'-OH, immediately 5' to UV-induced cycrobutane, pyrimidine dimers or 6-4 photoproducts and initiates an alternative excision repair process. Xeroderma pigmentosum group A cells expressing UVDE show UV resistance of almost the wild-type level. In these cells SSBs are produced upon UV irradiation and then efficiently repaired. The repair patch size is about 7 nucleotides and repair synthesis is decreased to 30% by aphidicoline, suggesting the involvement of a DNA polymerase δ/ε-dependent long-patch repair. Immediately after UV irradiation cellular proteins are poly(ADP-ribosyl)ated. The UV resistance of the cells is decreased in the presence of 3-aminobenzamide (3AB), an inhibitor of poly- (ADP-ribose) polymerase (PARP). Expression of UVDE in XRCC1-defective EM9, a Chinese hamster ovary cell line, greatly sensitizes the host cells to UV and addition of 3AB results in almost no further sensitization of the cells to UV. Thus, we show that XRCC1 and PARP are involved in the same pathway for the repair of SSBs.

虽然单链断裂（SSBs）经常发生，但细胞对单链断裂的反应和修复尚不清楚。为了解决这个问题，我们建立了表达粗神经孢子虫紫外线损伤内切酶（UVDE）的哺乳动物细胞系，该细胞系将具有3‘-OH，立即5’的SSB引入紫外线诱导的环丁烷，嘧啶二聚体或6-4光产物，并启动替代切除修复过程。表达UVDE的着色性干皮病A组细胞抗紫外线能力接近野生型水平。在这些细胞中，ssb在紫外线照射下产生，然后有效地修复。修复补丁大小约为7个核苷酸，修复合成减少到30%，这表明参与了DNA聚合酶δ/ε依赖的长补丁修复。紫外线照射后，细胞蛋白立即发生聚（adp -核糖基）化。3-氨基苯甲酰胺（3AB）是一种聚（adp -核糖）聚合酶（PARP）的抑制剂，它的存在降低了细胞对紫外线的抵抗力。在xrcc1缺陷的中国仓鼠卵巢细胞系EM9中表达UVDE可使宿主细胞对紫外线显著增敏，添加3AB后，细胞对紫外线几乎不再增敏。因此，我们发现XRCC1和PARP参与了ssb修复的相同途径。