Characterization of CDKN1A (p21) binding to sites of heavy-ion-induced damage: colocalization with proteins involved in DNA repair
Characterization of CDKN1A (p21) binding to sites of heavy-ion-induced damage: colocalization with proteins involved in DNA repair
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DOI：
10.1080/09553000110090007
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发表时间：
2002-02-01
影响因子：
2.6
通讯作者：
Taucher-Scholz, G
中科院分区：
文献类型：
--
作者：
Jakob, B;Scholz, M;Taucher-Scholz, G
用途：为了确定在重离子穿越位点的局部积累的CDKN 1A和DNA修复蛋白的协会材料和方法：CDKN 1A，PCNA，DNA-PK，hMre 11和Rad 50被研究了重离子照射后的亚核定位，使用免疫细胞化学染色和共聚焦激光扫描显微镜。结果：CDKN 1A在G（0）/G（1）正常人成纤维细胞的颗粒穿越部位形成核灶;病灶持续数小时，用DNase-I治疗后消失。病灶的形成也发生在NBS 1-或ATM-缺陷系和TP 53功能废除的细胞中。在正常成纤维细胞中，CDKN 1A病灶与hMre 11和Rad 50蛋白的颗粒诱导病灶共定位。然而，只有CDKN 1A再定位观察到辐射NBS 1细胞。重离子暴露后，正常成纤维细胞中也检测到与CDKN 1A暂时共定位的PCNA灶。与此相反，没有辐射诱导的亚核再定位被发现DNA-PK.Conclusions：CDKN 1A灶迅速出现在本地化的DNA损伤的网站由重离子诱导，并与染色质。有证据表明，CDKN 1A的本地化灶是不依赖于功能性TP 53和独立的hMre 11/Rad 50/NBS 1复合物的形成。这些数据支持CDKN 1A在辐射诱导的DNA损伤的感知或早期处理中的未知作用。
Purpose: To determine an association of locally accumulated CDKN1A and DNA repair proteins at the sites of heavy-ion traversals.Materials and methods: CDKN1A, PCNA, DNA-PK, hMre11 and Rad50 were investigated for their subnuclear localization after irradiation with heavy-ions using immunocytochemical staining and confocal laser-scanning microscopy. Human fibroblasts (normal diploid or XPA, ATM- or NBS1-deficient lines and HPV16 E6-transfected cells) were used.Results: CDKN1A formed nuclear foci in G(0)/G(1) normal human fibroblasts at the sites of particle traversal. Foci were persistent over hours and vanished after treatment with DNase-I. Formation of foci also occurred in NBS1- or ATM-deficient lines and in cells functionally abrogated for TP53. In normal fibroblasts, CDKN1A foci colocalized with particle-induced foci of the hMre11 and Rad50 proteins. However, only CDKN1A relocalization was observed in irradiated NBS1 cells. PCNA foci temporarily colocalizing with CDKN1A were also detected in normal fibroblasts after exposure to heavy-ions. In contrast, no radiation-induced subnuclear relocalization was found for DNA-PK.Conclusions : CDKN1A foci arise rapidly at sites of localized DNA damage induced by heavy-ions and are associated with the chromatin. Evidence is provided that localization of CDKN1A to foci is not dependent on functional TP53 and occurs independently of the formation of the hMre11/Rad50/NBS1 complex. The data support a yet unknown role of CDKN1A in sensing or early processing of radiation-induced DNA lesions.